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Ready, SET, Go: Post-translational regulation of the histone lysine methylation network in budding yeast

Open AccessPublished:July 02, 2021DOI:https://doi.org/10.1016/j.jbc.2021.100939
      Histone lysine methylation is a key epigenetic modification that regulates eukaryotic transcription. Here, we comprehensively review the function and regulation of the histone methylation network in the budding yeast and model eukaryote, Saccharomyces cerevisiae. First, we outline the lysine methylation sites that are found on histone proteins in yeast (H3K4me1/2/3, H3K36me1/2/3, H3K79me1/2/3, and H4K5/8/12me1) and discuss their biological and cellular roles. Next, we detail the reduced but evolutionarily conserved suite of methyltransferase (Set1p, Set2p, Dot1p, and Set5p) and demethylase (Jhd1p, Jhd2p, Rph1p, and Gis1p) enzymes that are known to control histone lysine methylation in budding yeast cells. Specifically, we illustrate the domain architecture of the methylation enzymes and highlight the structural features that are required for their respective functions and molecular interactions. Finally, we discuss the prevalence of post-translational modifications on yeast histone methylation enzymes and how phosphorylation, acetylation, and ubiquitination in particular are emerging as key regulators of enzyme function. We note that it will be possible to completely connect the histone methylation network to the cell’s signaling system, given that all methylation sites and cognate enzymes are known, most phosphosites on the enzymes are known, and the mapping of kinases to phosphosites is tractable owing to the modest set of protein kinases in yeast. Moving forward, we expect that the rich variety of post-translational modifications that decorates the histone methylation machinery will explain many of the unresolved questions surrounding the function and dynamics of this intricate epigenetic network.

      Keywords

      The abbreviations used are:

      AdoMet (S-adenosyl-L-methionine), AID (autoinhibitory domain), AWS (associated with SET), BAH (bromo-adjacent homology), COMPASS (complex of proteins associated with Set1), C2H2 (Cys2-His2), CTD (C-terminal domain), DSB (double strand break), Dot1p (disruptor of telomeric silencing 1), Gis1p (Gig1-2 suppressor 1), HAT (histone acetyltransferase), HDAC (histone deacetylase), Jhd1p (JmjC domain–containing histone demethylase 1), Jhd2p (JmjC domain–containing histone demethylase 2), JmjC (Jumonji C), JmjN (Jumonji N), LSD (lysine-specific demethylase), MYND (myeloid translocation protein, Nervy, Deaf), NHEJ (nonhomologous end joining), PHD (plant homeodomain), PRMT (protein arginine methyltransferase), PTM (post-translational modification), RNAPII (RNA polymerase II), Rph1 (regulator of Phr1), RRM (RNA recognition motif), SET (Su(var)3-9, Enhancer of Zeste, Trithorax), Sir (silent information regulator), SRI (Set2 Rbp1 interacting), ZF (zinc finger), 7βS (seven-β-strand)
      Within the eukaryotic cell, genetic material is packaged into chromatin. The basic repeating unit of chromatin is the nucleosome, which comprises 146 base pairs of linear DNA wrapped approximately 1.6 times around an octamer of core histone proteins (two copies each of H2A, H2B, H3, and H4) (
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      Chromatin structure: A repeating unit of histones and DNA.
      ). Each of these four histone families possesses a highly conserved and structured histone fold domain toward the center of the nucleosome, as well as a disordered N-terminal tail that protrudes from the nucleosomal core (
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      The significance, development and progress of high-throughput combinatorial histone code analysis.
      ). The spatial accessibility of histone tails makes them available for post-translational modification (PTM), and indeed a number of different modification types have been identified in these unstructured regions, including methylation (
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      The occurrence of iε-N-methyl lysine in histones.
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      Acetylation and methylation of histones and their possible role in the regulation of RNA synthesis.
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      ). Such modifications are known to regulate gene expression by either affecting chromatin compaction or by serving as binding platforms for transcriptional coregulators that harbor domains to specifically recognize modified histone residues. Given their central role in transcription, it is unsurprising that aberrant modification of histones has been linked to the pathogenesis of human cancers (
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      ), and autoimmune diseases (
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      Epigenetics in inflammatory rheumatic diseases.
      ). In the budding yeast Saccharomyces cerevisiae, dysregulated histone PTMs are associated with deleterious growth phenotypes (
      • Briggs S.D.
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      Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae.
      ,
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      Understanding histone H3 lysine 36 methylation and its deregulation in disease.
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      ) as well as altered apoptotic cell death and lifespan-resetting pathways (
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      Histone modifications as regulators of life and death in Saccharomyces cerevisiae.
      ).
      Histone methylation is a key epigenetic modification that regulates many nuclear processes, including transcription (
      • Black J.C.
      • Van Rechem C.
      • Whetstine J.R.
      Histone lysine methylation dynamics: Establishment, regulation, and biological impact.
      ), DNA replication (
      • Fu H.
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      • Zhao K.
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      Methylation of histone H3 on lysine 79 associates with a group of replication origins and helps limit DNA replication once per cell cycle.
      ), and DNA repair (
      • Gong F.
      • Miller K.M.
      Histone methylation and the DNA damage response.
      ). It refers to the covalent attachment of methyl (CH3) group(s) to the amino acid side chains of lysine or arginine residues on histone proteins. Lysine residues can be mono-, di-, or trimethylated on their ε-amino group (
      • Murray K.
      The occurrence of iε-N-methyl lysine in histones.
      ), whereas arginine residues can be mono-, asymmetrically di-, or symmetrically dimethylated on their terminal guanidinium group (
      • Bedford M.T.
      • Richard S.
      Arginine methylation: An emerging regulatorof protein function.
      ). Unlike other modifications that affect chromatin folding through an electrostatic mechanism (e.g., acetylation, phosphorylation), methylation does not alter the charge of lysine or arginine side chains. Instead, methylated residues constitute recognition sites for a range of transcription factors and associated regulatory proteins, which in turn elicit downstream changes in gene expression (
      • Martin C.
      • Zhang Y.
      The diverse functions of histone lysine methylation.
      ). These effector proteins carry “reader” interaction interfaces, such as chromodomains (
      • Nielsen P.R.
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      • Mott H.R.
      • Callaghan J.
      • Bannister A.
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      • Murzin A.G.
      • Murzina N.V.
      • Laue E.D.
      Structure of the HP1 chromodomain bound to histone H3 methylated at lysine 9.
      ), PHD (
      • Shi X.
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      • Walter K.L.
      • Ewalt M.
      • Michishita E.
      • Hung T.
      • Carney D.
      • Pena P.
      • Lan F.
      • Kaadige M.R.
      ING2 PHD domain links histone H3 lysine 4 methylation to active gene repression.
      ), and Tudor domains (
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      • Lake A.
      • Krishna M.
      • Xia L.
      • Zhang Y.
      • Bedford M.T.
      Tudor, MBT and chromo domains gauge the degree of lysine methylation.
      ), which specifically bind methyl-lysine and methyl-arginine residues. Although some histone modifications simply denote either an open or closed chromatin conformation, methylation has a more nuanced role. Accordingly, specific methyl-lysine sites on histones can have either activating or repressive effects on transcription depending on their position and methylation state (
      • Black J.C.
      • Van Rechem C.
      • Whetstine J.R.
      Histone lysine methylation dynamics: Establishment, regulation, and biological impact.
      ). In budding yeast, lysine methylation sites at H3K4, H3K36, and H3K79 are enriched within transcriptionally active euchromatin and are predominantly associated with gene expression. Strikingly, these methyl marks can also promote a repressed chromatin landscape depending on their location within transcriptional units, thus highlighting their functional diversity. All eukaryotic histone methylation sites display unique chromosomal signatures, both throughout gene bodies and within noncoding and regulatory elements (e.g., promoters, enhancers) (
      • Ernst J.
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      • Mikkelsen T.S.
      • Shoresh N.
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      • Epstein C.B.
      • Zhang X.
      • Wang L.
      • Issner R.
      • Coyne M.
      Mapping and analysis of chromatin state dynamics in nine human cell types.
      ,
      • Heintzman N.D.
      • Stuart R.K.
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      • Fu Y.
      • Ching C.W.
      • Hawkins R.D.
      • Barrera L.O.
      • Van Calcar S.
      • Qu C.
      • Ching K.A.
      Distinct and predictive chromatin signatures of transcriptional promoters and enhancers in the human genome.
      ). Crucially, the abundance and distribution of specific methyl marks changes markedly during cellular growth (
      • Briggs S.D.
      • Bryk M.
      • Strahl B.D.
      • Cheung W.L.
      • Davie J.K.
      • Dent S.Y.
      • Winston F.
      • Allis C.D.
      Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae.
      ), differentiation (
      • Li E.
      Chromatin modification and epigenetic reprogramming in mammalian development.
      ), and in response to exogenous perturbation (
      • Weiner A.
      • Hsieh T.-H.S.
      • Appleboim A.
      • Chen H.V.
      • Rahat A.
      • Amit I.
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      • Friedman N.
      High-resolution chromatin dynamics during a yeast stress response.
      ), to bring about widespread transcriptional reprogramming.
      The histone methylation network is exquisitely conserved across eukaryotes, from the methylation sites themselves to the enzymatic machinery responsible for their regulation (
      • Woo Y.H.
      • Li W.-H.
      Evolutionary conservation of histone modifications in mammals.
      ,
      • Roguev A.
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      • Shevchenko A.
      • Stewart A.F.
      High conservation of the Set1/Rad6 axis of histone 3 lysine 4 methylation in budding and fission yeasts.
      ,
      • Fuchs J.
      • Demidov D.
      • Houben A.
      • Schubert I.
      Chromosomal histone modification patterns–from conservation to diversity.
      ). In the human cell, there are seven histone lysine methylation sites that are controlled by the counteracting activities of 30 methyltransferases and 22 demethylases (
      • Højfeldt J.W.
      • Agger K.
      • Helin K.
      Histone lysine demethylases as targets for anticancer therapy.
      ,
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      Histone lysine methyltransferases in biology and disease.
      ,
      • Separovich R.J.
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      Controlling the controllers: Regulation of histone methylation by phosphosignalling.
      ). This system is highly complex and challenging to interrogate experimentally given the number, redundancy, and overlapping site specificity of its constituent members. By contrast, S. cerevisiae is a eukaryotic model organism in which many of the foundational discoveries of histones and chromatin biology have been made (
      • Rando O.J.
      • Winston F.
      Chromatin and transcription in yeast.
      ). In yeast, the histone methylation system is substantially simplified, comprising six histone lysine methyl marks and only four methyltransferases and four demethylases (Fig. 1). Crucially, almost all yeast histone methyl marks and enzymes have a mammalian counterpart, thus underscoring the high degree of evolutionary conservation of this system (Table 1). Deciphering the function and regulation of the histone methylation network in yeast will therefore be of relevance to all eukaryotes and will provide new insights into epigenetic processes in higher organisms. This is of pharmacological importance given the association of human histone methylation enzymes with disease etiology (
      • Donaldson-Collier M.C.
      • Sungalee S.
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      • Tavernari D.
      • Katanayeva N.
      • Battistello E.
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      EZH2 oncogenic mutations drive epigenetic, transcriptional, and structural changes within chromatin domains.
      ,
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      Histone methylation: A dynamic mark in health, disease and inheritance.
      ,
      • Hyun K.
      • Jeon J.
      • Park K.
      • Kim J.
      Writing, erasing and reading histone lysine methylations.
      ) and their emergence as promising therapeutic targets for anticancer drug development (
      • Liu Y.
      • Liu K.
      • Qin S.
      • Xu C.
      • Min J.
      Epigenetic targets and drug discovery: Part 1: Histone methylation.
      ,
      • McGrath J.
      • Trojer P.
      Targeting histone lysine methylation in cancer.
      ,
      • Morera L.
      • Lübbert M.
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      Targeting histone methyltransferases and demethylases in clinical trials for cancer therapy.
      ,
      • Song Y.
      • Wu F.
      • Wu J.
      Targeting histone methylation for cancer therapy: Enzymes, inhibitors, biological activity and perspectives.
      ). Histone methylation sites and enzymes are also conserved, albeit to varying extents, in lower eukaryotes that impact on human health, such as protozoans (
      • Croken M.M.
      • Nardelli S.C.
      • Kim K.
      Chromatin modifications, epigenetics, and how protozoan parasites regulate their lives.
      ,
      • Emery-Corbin S.J.
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      • Balan B.
      • Rojas-López L.
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      • Jex A.R.
      Eukaryote-conserved histone post-translational modification landscape in Giardia duodenalis revealed by mass spectrometry.
      ,
      • Jiang L.
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      • Ni T.
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      • Rayavara K.
      • Yang W.
      • Turner L.
      • Lavstsen T.
      • Theander T.G.
      PfSETvs methylation of histone H3K36 represses virulence genes in Plasmodium falciparum.
      ). Budding yeast also serves as an excellent model for epigenetic gene regulation in these species and may therefore provide insights into the design of antiparasitic drugs.
      Figure thumbnail gr1
      Figure 1Histone lysine methylation network in budding yeast. All histone lysine methylation sites in yeast are depicted along histone proteins as yellow diamonds. The upstream methyltransferase and demethylase enzymes that control these sites are shown in green and pink, respectively. Histone methyl marks are recognized by downstream effector proteins that harbor methyl-reader domains and are colored according to the key (bottom). Methylation can also inhibit binding of proteins to chromatin, notably the bromo-adjacent homology (BAH) domain of Sir3p (magenta), which is blocked by H3K79 methylation. The functional outcomes of histone methylation sites and the recruitment of specific effector proteins and complexes are shown in gray boxes. For ease of visualization, only a single copy of histones H3 and H4 has been illustrated, whereas both copies of histones H2A and H2B have been omitted.
      Table 1Histone lysine methylation enzymes in budding yeast
      Type
      Methyltransferase (MTase) or demethylase (DMase).
      EnzymeUniProt IDSGD IDAliasSpecificityEC
      Enzyme Commission number.
      ChromosomeCoordinatesCopies/cell
      Under log-phase growth in synthetic defined medium (346).
      Human ortholog
      From Alliance of Genome Resources Release 4.0.0 (347).
      MTaseSet1pP38827YHR119WKMT2H3K4me1/2/32.1.1.354VIII346043–349285172SETD1A
      Set2pP46995YJL168CKMT3H3K36me1/2/32.1.1.359X102227–104428217SETD2
      Dot1pQ04089YDR440WKMT4H3K79me1/2/32.1.1.360IV1342493–13442412160DOT1L
      Set5pP38890YHR207C-H4K5/8/12me1-VIII514905–5164855000SMYD3
      DMaseJhd1pP40034YER051WKDM2H3K36me1/21.14.11.27V254656–256134784FBXL11
      Jhd2pP47156YJR119CKDM5H3K4me1/2/31.14.11.67X644304–646490290JARID1C
      Rph1pP39956YER169WKDM4H3K36me2/31.14.11.27V523369–5257592229JMJD2A
      Gis1pQ03833YDR096WKDM4H3K36me1/21.14.11.27IV637139–639823432JMJD2A
      a Methyltransferase (MTase) or demethylase (DMase).
      b Enzyme Commission number.
      c Under log-phase growth in synthetic defined medium (
      • Ghaemmaghami S.
      • Huh W.-K.
      • Bower K.
      • Howson R.W.
      • Belle A.
      • Dephoure N.
      • O'Shea E.K.
      • Weissman J.S.
      Global analysis of protein expression in yeast.
      ).
      d From Alliance of Genome Resources Release 4.0.0 (
      The Alliance of Genome Resources Consortium
      Alliance of genome Resources Portal: Unified model organism research platform.
      ).
      Despite the importance that S. cerevisiae has played in our understanding of histone methylation, and the high conservation of these processes in higher eukaryotes, there has been no systematic analysis of the literature to date for this key epigenetic modification in yeast. To this end, here we present a comprehensive review of the histone lysine methylation system in yeast. We first outline the histone methylation sites and their contribution to both molecular and cellular processes. Next, we provide a detailed examination of the histone methyltransferases and demethylases that control these marks, with a particular focus on their structure, function, and regulation. Finally, we discuss the PTMs that are known to exist on histone methylation enzymes and how they are emerging as key regulators of enzyme function. We expect the latter will ultimately explain many of the intricacies of the histone methylation network in due course.

      Histone methylation sites

      Many of the histone methylation sites were first discovered through Edman sequencing of bulk histones after metabolic labeling (
      • Zhang Y.
      • Reinberg D.
      Transcription regulation by histone methylation: Interplay between different covalent modifications of the core histone tails.
      ), following which their roles in transcriptional regulation and other cellular processes have been established (
      • Margueron R.
      • Trojer P.
      • Reinberg D.
      The key to development: Interpreting the histone code?.
      ,
      • Shi Y.
      • Whetstine J.R.
      Dynamic regulation of histone lysine methylation by demethylases.
      ). Strikingly, many of the major activating methylation sites are conserved among eukaryotes, whereas the repressive marks are more variable in their evolutionary conservation (
      • Fuchs J.
      • Demidov D.
      • Houben A.
      • Schubert I.
      Chromosomal histone modification patterns–from conservation to diversity.
      ). This is exemplified in S. cerevisiae which carries the transcriptionally activating histone lysine methylations at H3K4, H3K36, and H3K79 but not the repressive H3K9, H3K27, and H4K20 methylation sites (Fig. 1) (
      • Zhao Y.
      • Garcia B.A.
      Comprehensive catalog of currently documented histone modifications.
      ). More recently, monomethylation of H4K5, H4K8, and H4K12 has been identified in yeast (
      • Green E.M.
      • Mas G.
      • Young N.L.
      • Garcia B.A.
      • Gozani O.
      Methylation of H4 lysines 5, 8 and 12 by yeast Set5 calibrates chromatin stress responses.
      ). Here, we summarize the histone lysine methyl marks in S. cerevisiae, their genomic distribution, biological function, and how they cross talk with other histone PTMs.

      H3K4me1/2/3

      Histone H3 lysine 4 (H3K4) exhibits three methylation states, each of which have distinct functions and positional signatures across the yeast genome. Functionally, all three states are associated with transcriptional activation in a variety of eukaryotic species (
      • Ruthenburg A.J.
      • Allis C.D.
      • Wysocka J.
      Methylation of lysine 4 on histone H3: Intricacy of writing and reading a single epigenetic mark.
      ). Trimethylation of H3K4 (H3K4me3) is concentrated within promoter regions and toward the 5′-ends of actively transcribed genes (Fig. 2A) (
      • Rando O.J.
      Global patterns of histone modifications.
      ). H3K4me3 is highly correlated with transcription rates, active RNA polymerase II (RNAPII) occupancy, and histone acetylation (
      • Pokholok D.K.
      • Harbison C.T.
      • Levine S.
      • Cole M.
      • Hannett N.M.
      • Lee T.I.
      • Bell G.W.
      • Walker K.
      • Rolfe P.A.
      • Herbolsheimer E.
      Genome-wide map of nucleosome acetylation and methylation in yeast.
      ), and its enrichment at transcription start sites is highly conserved across eukaryotes (
      • Bernstein B.E.
      • Kamal M.
      • Lindblad-Toh K.
      • Bekiranov S.
      • Bailey D.K.
      • Huebert D.J.
      • McMahon S.
      • Karlsson E.K.
      • Kulbokas Iii, E.J.
      • Gingeras T.R.
      Genomic maps and comparative analysis of histone modifications in human and mouse.
      ,
      • Bernstein B.E.
      • Mikkelsen T.S.
      • Xie X.
      • Kamal M.
      • Huebert D.J.
      • Cuff J.
      • Fry B.
      • Meissner A.
      • Wernig M.
      • Plath K.
      A bivalent chromatin structure marks key developmental genes in embryonic stem cells.
      ). By contrast, the distribution and function of dimethylated H3K4 (H3K4me2) is variable between yeast and vertebrates (
      • Ruthenburg A.J.
      • Allis C.D.
      • Wysocka J.
      Methylation of lysine 4 on histone H3: Intricacy of writing and reading a single epigenetic mark.
      ). In S. cerevisiae, H3K4me2 is most abundant toward the middle of coding regions and is associated with both transcriptionally poised and active regions (Fig. 2A). The majority of H3K4me2 in vertebrates, however, colocalizes with H3K4me3 in discrete zones (~5–20 nucleosomes in length) proximal to highly transcribed genes (
      • Bernstein B.E.
      • Kamal M.
      • Lindblad-Toh K.
      • Bekiranov S.
      • Bailey D.K.
      • Huebert D.J.
      • McMahon S.
      • Karlsson E.K.
      • Kulbokas Iii, E.J.
      • Gingeras T.R.
      Genomic maps and comparative analysis of histone modifications in human and mouse.
      ). Monomethylated H3K4 (H3K4me1) peaks toward the 3′-ends of transcriptional units (Fig. 2A) and is considered a hallmark of active enhancers in metazoans (
      • Heintzman N.D.
      • Stuart R.K.
      • Hon G.
      • Fu Y.
      • Ching C.W.
      • Hawkins R.D.
      • Barrera L.O.
      • Van Calcar S.
      • Qu C.
      • Ching K.A.
      Distinct and predictive chromatin signatures of transcriptional promoters and enhancers in the human genome.
      ,
      • Heintzman N.D.
      • Ren B.
      Finding distal regulatory elements in the human genome.
      ). Of importance, patterning of all H3K4 methylation states along active genes is not static or universal and changes dynamically depending on the frequency and rate of transcription elongation (
      • Soares L.M.
      • He P.C.
      • Chun Y.
      • Suh H.
      • Kim T.
      • Buratowski S.
      Determinants of histone H3K4 methylation patterns.
      ) and in response to transcriptional stress (
      • Zhang L.
      • Schroeder S.
      • Fong N.
      • Bentley D.L.
      Altered nucleosome occupancy and histone H3K4 methylation in response to ‘transcriptional stress’.
      ) in a gene-specific manner.
      Figure thumbnail gr2
      Figure 2Genomic distribution of histone lysine methylation sites in budding yeast and their regulation by other histone residues and modifications. A, the abundance of the mono-, di-, and trimethylated forms of H3K4 (top), H3K36 (middle), and H3K79 (bottom) along an active transcriptional unit is depicted by a color intensity gradient (indigo). B, regulation of budding yeast histone methylation sites by other histone residues and PTMs including acetylation (Ac; orange), ubiquitination (Ub; purple), and methylation (Me; yellow). The upstream modifying enzymes responsible for these PTMs are colored according to the key (bottom). The effects of these histone residues and PTMs are tabulated for each major lysine methylation site, where red arrows denote a stimulatory effect and blue arrows indicate an inhibitory effect. For both panels, histone H4 monomethylation sites at K5, K8, and K12 have been omitted given that little is known about the distribution of these modifications along genes and their cross talk with other features of the chromatin landscape. PTM, post-translational modification; TSS, transcription start site; TTS, transcription termination site.
      H3K4 methylation is a nuanced epigenetic modification that can participate in both gene activation and repression (
      • Malik S.
      • Bhaumik S.R.
      Mixed lineage leukemia: Histone H3 lysine 4 methyltransferases from yeast to human.
      ). In its canonical role, methylation of H3K4 is most commonly associated with transcriptional activation and is found abundantly within genes being actively transcribed by RNAPII (
      • Soares L.M.
      • Radman-Livaja M.
      • Lin S.G.
      • Rando O.J.
      • Buratowski S.
      Feedback control of Set1 protein levels is important for proper H3K4 methylation patterns.
      ). It is required for the normal induction of transcription in yeast (
      • Bhaumik S.R.
      • Smith E.
      • Shilatifard A.
      Covalent modifications of histones during development and disease pathogenesis.
      ,
      • Kouzarides T.
      Chromatin modifications and their function.
      ,
      • Shilatifard A.
      Chromatin modifications by methylation and ubiquitination: Implications in the regulation of gene expression.
      ), which is achieved by the recruitment of specific chromatin modifiers (e.g., Chd1p, Isw1p, Yng1p, Pho23p, Set3p (
      • Chong S.Y.
      • Cutler S.
      • Lin J.-J.
      • Tsai C.-H.
      • Tsai H.-K.
      • Biggins S.
      • Tsukiyama T.
      • Lo Y.-C.
      • Kao C.-F.
      H3K4 methylation at active genes mitigates transcription-replication conflicts during replication stress.
      )) via their methyl “reader” domains (Fig. 1). These effectors have varying degrees of preference for distinct methylation states of H3K4 (
      • Sims R.J.
      • Reinberg D.
      Histone H3 Lys 4 methylation: Caught in a bind?.
      ). For example, Yng1p binds H3K4 trimethylated chromatin using its plant homeodomain (PHD) finger (
      • Martin B.J.
      • McBurney K.L.
      • Maltby V.E.
      • Jensen K.N.
      • Brind’Amour J.
      • Howe L.J.
      Histone H3K4 and H3K36 methylation independently recruit the NuA3 histone acetyltransferase in Saccharomyces cerevisiae.
      ) and subsequently associates with the yeast NuA3 histone acetyltransferase (HAT) complex to catalyze histone acetylation at the promoter and thus activate transcription. As an additional layer of regulation, H3K4me2 predominantly recruits histone deacetylases (HDACs, e.g., Set3p) throughout the body of a gene to prevent cryptic transcriptional initiation sites (
      • Kim T.
      • Buratowski S.
      Dimethylation of H3K4 by Set1 recruits the Set3 histone deacetylase complex to 5′ transcribed regions.
      ). Little is known about the molecular functions of H3K4me1 in transcriptional activation in yeast; however, it has been speculated that its presence is simply a transitional state between unmodified and dimethylated H3K4 (
      • Ramakrishnan S.
      • Pokhrel S.
      • Palani S.
      • Pflueger C.
      • Parnell T.J.
      • Cairns B.R.
      • Bhaskara S.
      • Chandrasekharan M.B.
      Counteracting H3K4 methylation modulators Set1 and Jhd2 co-regulate chromatin dynamics and gene transcription.
      ). With respect to repression, H3K4 methylation has been found to be involved in silencing several heterochromatic genomic regions (e.g., telomeres (
      • Krogan N.J.
      • Dover J.
      • Khorrami S.
      • Greenblatt J.F.
      • Schneider J.
      • Johnston M.
      • Shilatifard A.
      COMPASS, a histone H3 (Lysine 4) methyltransferase required for telomeric silencing of gene expression.
      ), ribosomal DNA clusters (
      • Briggs S.D.
      • Bryk M.
      • Strahl B.D.
      • Cheung W.L.
      • Davie J.K.
      • Dent S.Y.
      • Winston F.
      • Allis C.D.
      Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae.
      ), HML mating-type locus (
      • Nislow C.
      • Ray E.
      • Pillus L.
      SET1, a yeast member of the trithorax family, functions in transcriptional silencing and diverse cellular processes.
      )). Accordingly, several ribosomal biosynthesis genes are downregulated by H3K4 methylation during multiple stresses (
      • Weiner A.
      • Chen H.V.
      • Liu C.L.
      • Rahat A.
      • Klien A.
      • Soares L.
      • Gudipati M.
      • Pfeffner J.
      • Regev A.
      • Buratowski S.
      Systematic dissection of roles for chromatin regulators in a yeast stress response.
      ). Simultaneous loss of H3K4me3 and H3K4me2 in yeast results in increased steady-state mRNA levels and delayed repression kinetics for certain gene groups in vivo (
      • Margaritis T.
      • Oreal V.
      • Brabers N.
      • Maestroni L.
      • Vitaliano-Prunier A.
      • Benschop J.J.
      • van Hooff S.
      • van Leenen D.
      • Dargemont C.
      • Geli V.
      Two distinct repressive mechanisms for histone 3 lysine 4 methylation through promoting 3′-end antisense transcription.
      ), whereas H3K4me1 specifically inhibits gene expression induced by osmotic stress (
      • Nadal-Ribelles M.
      • Mas G.
      • Millán-Zambrano G.
      • Solé C.
      • Ammerer G.
      • Chávez S.
      • Posas F.
      • de Nadal E.
      H3K4 monomethylation dictates nucleosome dynamics and chromatin remodeling at stress-responsive genes.
      ). These repressive effects are also mediated by chromatin modifiers and their cognate reader domains; Pho23p recognizes H3K4me3 and recruits the transcriptionally repressive Rpd3L HDAC complex (Fig. 1) (
      • Wang S.-S.
      • Zhou B.O.
      • Zhou J.-Q.
      Histone H3 lysine 4 hypermethylation prevents aberrant nucleosome remodeling at the PHO5 promoter.
      ).
      At the cellular level, H3K4 methylation is involved in several key cellular processes and its dysregulation contributes to deleterious phenotypes. This modification has been shown to mediate yeast cell cycle progression and assembly of the mitotic spindle (
      • Beilharz T.H.
      • Harrison P.F.
      • Miles D.M.
      • See M.M.
      • Le U.M.M.
      • Kalanon M.
      • Curtis M.J.
      • Hasan Q.
      • Saksouk J.
      • Margaritis T.
      Coordination of cell cycle progression and mitotic spindle assembly involves histone H3 lysine 4 methylation by Set1/COMPASS.
      ), DNA damage response and genomic stability (
      • Faucher D.
      • Wellinger R.J.
      Methylated H3K4, a transcription-associated histone modification, is involved in the DNA damage response pathway.
      ), and mRNA splicing (
      • Hérissant L.
      • Moehle E.A.
      • Bertaccini D.
      • Van Dorsselaer A.
      • Schaeffer-Reiss C.
      • Guthrie C.
      • Dargemont C.
      H2B ubiquitylation modulates spliceosome assembly and function in budding yeast.
      ), and serves as an important molecular trigger for cell death (
      • Walter D.
      • Matter A.
      • Fahrenkrog B.
      Loss of histone H3 methylation at lysine 4 triggers apoptosis in Saccharomyces cerevisiae.
      ). Of interest, increased levels of overall H3K4 methylation have been reported to act as a memory of recent transcriptional activity that allows genes to be rapidly switched on or off in response to stimuli (
      • Ng H.H.
      • Robert F.
      • Young R.A.
      • Struhl K.
      Targeted recruitment of Set1 histone methylase by elongating Pol II provides a localized mark and memory of recent transcriptional activity.
      ). Yeast cells with defective H3K4 methylation have decreased viability owing to improper repair of double-strand DNA breaks (DSBs) by nonhomologous end-joining (NHEJ) (
      • Faucher D.
      • Wellinger R.J.
      Methylated H3K4, a transcription-associated histone modification, is involved in the DNA damage response pathway.
      ). They also display increased cell death during chronological aging (
      • Walter D.
      • Matter A.
      • Fahrenkrog B.
      Loss of histone H3 methylation at lysine 4 triggers apoptosis in Saccharomyces cerevisiae.
      ) and are sensitive to certain antifungal drugs (e.g., Brefeldin A) owing to abnormal expression of ergosterol biosynthesis enzymes (e.g., HMGCR) (
      • South P.F.
      • Harmeyer K.M.
      • Serratore N.D.
      • Briggs S.D.
      H3K4 methyltransferase Set1 is involved in maintenance of ergosterol homeostasis and resistance to Brefeldin A.
      ).
      Methylation at H3K4 is known to cross talk with multiple other histone modifications. Perhaps the most well-studied example of eukaryotic PTM cross talk is the interplay between H3K4 methylation and H2B ubiquitination (Fig. 2B). Studies in S. cerevisiae first demonstrated that monoubiquitination at H2BK123 is required for subsequent H3K4 methylation (
      • Briggs S.D.
      • Xiao T.
      • Sun Z.-W.
      • Caldwell J.A.
      • Shabanowitz J.
      • Hunt D.F.
      • Allis C.D.
      • Strahl B.D.
      Trans-histone regulatory pathway in chromatin.
      ,
      • Dover J.
      • Schneider J.
      • Tawiah-Boateng M.A.
      • Wood A.
      • Dean K.
      • Johnston M.
      • Shilatifard A.
      Methylation of histone H3 by COMPASS requires ubiquitination of histone H2B by Rad6.
      ,
      • Sun Z.-W.
      • Allis C.D.
      Ubiquitination of histone H2B regulates H3 methylation and gene silencing in yeast.
      ), a trans-regulatory mechanism that has since been elucidated for several human H3K4 methyltransferase complexes (
      • Kim J.
      • Guermah M.
      • McGinty R.K.
      • Lee J.-S.
      • Tang Z.
      • Milne T.A.
      • Shilatifard A.
      • Muir T.W.
      • Roeder R.G.
      RAD6-Mediated transcription-coupled H2B ubiquitylation directly stimulates H3K4 methylation in human cells.
      ,
      • Kim J.
      • Kim J.-A.
      • McGinty R.K.
      • Nguyen U.T.
      • Muir T.W.
      • Allis C.D.
      • Roeder R.G.
      The n-SET domain of Set1 regulates H2B ubiquitylation-dependent H3K4 methylation.
      ). Histone H2BK123 monoubiquitination, and thus H3K4 methylation, requires the yeast E2 ubiquitin-conjugating enzyme, Rad6p, and its cognate E3 ligase, Bre1p (
      • Lee J.-S.
      • Shukla A.
      • Schneider J.
      • Swanson S.K.
      • Washburn M.P.
      • Florens L.
      • Bhaumik S.R.
      • Shilatifard A.
      Histone crosstalk between H2B monoubiquitination and H3 methylation mediated by COMPASS.
      ), and is involved in the transcriptional silencing of telomeric genes (
      • Dover J.
      • Schneider J.
      • Tawiah-Boateng M.A.
      • Wood A.
      • Dean K.
      • Johnston M.
      • Shilatifard A.
      Methylation of histone H3 by COMPASS requires ubiquitination of histone H2B by Rad6.
      ). The mechanism of cross talk will be discussed in greater detail later in the article (see “Histone methyltransferase” section). Curiously, despite conservation in mammalian cells, this histone methylation/ubiquitination interplay is absent in the fission yeast, Schizosaccharomyces pombe (
      • Mikheyeva I.V.
      • Grady P.J.
      • Tamburini F.B.
      • Lorenz D.R.
      • Cam H.P.
      Multifaceted genome control by Set1 dependent and independent of H3K4 methylation and the Set1C/COMPASS complex.
      ). Other examples of H3K4 methyl cross talk include its negative regulation by adjacent arginine asymmetric dimethylation at H3R2 (Fig. 2B), which abrogates H3K4me3 via spatial occlusion of the methylation machinery (
      • Kirmizis A.
      • Santos-Rosa H.
      • Penkett C.J.
      • Singer M.A.
      • Vermeulen M.
      • Mann M.
      • Bähler J.
      • Green R.D.
      • Kouzarides T.
      Arginine methylation at histone H3R2 controls deposition of H3K4 trimethylation.
      ). A systematic histone mutagenesis screen revealed that H3K14, which is known to be acetylated by Gcn5p and Sas3p in vivo, is a critical residue for H3K4me3 levels, suggesting a potential cis-regulatory cross talk between histone methylation and acetylation through an unknown mechanism (
      • Nakanishi S.
      • Sanderson B.W.
      • Delventhal K.M.
      • Bradford W.D.
      • Staehling-Hampton K.
      • Shilatifard A.
      A comprehensive library of histone mutants identifies nucleosomal residues required for H3K4 methylation.
      ). Finally, antagonism between acetylation and methylation of H3K4 serves to fine-tune the deposition of these competing modifications, whereby H3K4 methylation limits H3K4 acetylation at promoters, and vice versa (
      • Guillemette B.
      • Drogaris P.
      • Lin H.-H.S.
      • Armstrong H.
      • Hiragami-Hamada K.
      • Imhof A.
      • Bonneil E.
      • Thibault P.
      • Verreault A.
      • Festenstein R.J.
      H3 lysine 4 is acetylated at active gene promoters and is regulated by H3 lysine 4 methylation.
      ).

      H3K36me1/2/3

      Histone H3 lysine 36 (H3K36) can be co-transcriptionally modified by the addition of one (H3K36me1), two (H3K36me2), or three (H3K36me3) methyl groups in budding yeast and other eukaryotes (
      • Schmähling S.
      • Meiler A.
      • Lee Y.
      • Mohammed A.
      • Finkl K.
      • Tauscher K.
      • Israel L.
      • Wirth M.
      • Philippou-Massier J.
      • Blum H.
      Regulation and function of H3K36 di-methylation by the trithorax-group protein complex AMC.
      ,
      • Venkatesh S.
      • Smolle M.
      • Li H.
      • Gogol M.M.
      • Saint M.
      • Kumar S.
      • Natarajan K.
      • Workman J.L.
      Set2 methylation of histone H3 lysine 36 suppresses histone exchange on transcribed genes.
      ). All three methylation states of H3K36 accumulate in transcribed regions, making this modification a hallmark of active transcriptional elongation (
      • Li J.
      • Ahn J.H.
      • Wang G.G.
      Understanding histone H3 lysine 36 methylation and its deregulation in disease.
      ). Genome-wide localization studies in S. cerevisiae have shown that levels of H3K36 methylation increase in a 5′-to-3′ gradient along active transcriptional units, the direct converse of H3K4me patterning (Fig. 2A). Consequently, H3K36me1 is predominantly found at the 5′-end of gene bodies, and H3K36me2 and H3K36me3 are concentrated at their 3′-ends (
      • Pokholok D.K.
      • Harbison C.T.
      • Levine S.
      • Cole M.
      • Hannett N.M.
      • Lee T.I.
      • Bell G.W.
      • Walker K.
      • Rolfe P.A.
      • Herbolsheimer E.
      Genome-wide map of nucleosome acetylation and methylation in yeast.
      ,
      • Bell O.
      • Wirbelauer C.
      • Hild M.
      • Scharf A.N.
      • Schwaiger M.
      • MacAlpine D.M.
      • Zilbermann F.
      • Van Leeuwen F.
      • Bell S.P.
      • Imhof A.
      Localized H3K36 methylation states define histone H4K16 acetylation during transcriptional elongation in Drosophila.
      ,
      • Rao B.
      • Shibata Y.
      • Strahl B.D.
      • Lieb J.D.
      Dimethylation of histone H3 at lysine 36 demarcates regulatory and nonregulatory chromatin genome-wide.
      ). Trimethylation at H3K36 in particular is highly correlated with active transcription and recruits distinct reader proteins to maintain a permissive transcriptional landscape (Fig. 1) (
      • Wozniak G.G.
      • Strahl B.D.
      Hitting the ‘mark’: Interpreting lysine methylation in the context of active transcription.
      ). Indeed, H3K36 methylation does not spread to adjacent loci downstream of transcription termination sites and exhibits a relatively short epigenetic memory for recent transcriptional activity (
      • Sein H.
      • Värv S.
      • Kristjuhan A.
      Distribution and maintenance of histone H3 lysine 36 trimethylation in transcribed locus.
      ). Methylation at H3K36 is also conserved in lower eukaryotic microbes where it plays a critical role in the regulation of chromatin-templated processes. For example, in the pathogenic protozoan Plasmodium falciparum, H3K36 trimethylation by PfSETvs represses transcription of virtually all virulence genes in infected erythrocytes (
      • Jiang L.
      • Mu J.
      • Zhang Q.
      • Ni T.
      • Srinivasan P.
      • Rayavara K.
      • Yang W.
      • Turner L.
      • Lavstsen T.
      • Theander T.G.
      PfSETvs methylation of histone H3K36 represses virulence genes in Plasmodium falciparum.
      ).
      In budding yeast, H3K36 methylation regulates chromatin structure and transcriptional fidelity through the recruitment of specific chromatin modifiers involved in diverse cellular pathways (
      • Lerner A.M.
      • Hepperla A.J.
      • Keele G.R.
      • Meriesh H.A.
      • Yumerefendi H.
      • Restrepo D.
      • Zimmerman S.
      • Bear J.E.
      • Kuhlman B.
      • Davis I.J.
      An optogenetic switch for the Set2 methyltransferase provides evidence for transcription-dependent and-independent dynamics of H3K36 methylation.
      ). Within the context of transcription, distinct methylation states of H3K36 are known to differentially engage three major macromolecular complexes: (1) Rpd3S, (2) Isw1b, and (3) NuA3 (Fig. 1). With respect to the former, the Rpd3S HDAC complex specifically binds H3K36 methylated chromatin via the chromodomain of its constituent member, Eaf3p (
      • Carrozza M.J.
      • Li B.
      • Florens L.
      • Suganuma T.
      • Swanson S.K.
      • Lee K.K.
      • Shia W.-J.
      • Anderson S.
      • Yates J.
      • Washburn M.P.
      Histone H3 methylation by Set2 directs deacetylation of coding regions by Rpd3S to suppress spurious intragenic transcription.
      ,
      • Joshi A.A.
      • Struhl K.
      Eaf3 chromodomain interaction with methylated H3-K36 links histone deacetylation to Pol II elongation.
      ,
      • Keogh M.-C.
      • Kurdistani S.K.
      • Morris S.A.
      • Ahn S.H.
      • Podolny V.
      • Collins S.R.
      • Schuldiner M.
      • Chin K.
      • Punna T.
      • Thompson N.J.
      Cotranscriptional set2 methylation of histone H3 lysine 36 recruits a repressive Rpd3 complex.
      ). This interaction is further stabilized by the PHD domain of another Rpd3S subunit, Rco1p (
      • Li B.
      • Gogol M.
      • Carey M.
      • Lee D.
      • Seidel C.
      • Workman J.L.
      Combined action of PHD and chromo domains directs the Rpd3S HDAC to transcribed chromatin.
      ). Rpd3S is thus preferentially targeted to H3K36me throughout coding regions where it promotes widespread histone deacetylation in the wake of transcription (
      • Govind C.K.
      • Qiu H.
      • Ginsburg D.S.
      • Ruan C.
      • Hofmeyer K.
      • Hu C.
      • Swaminathan V.
      • Workman J.L.
      • Li B.
      • Hinnebusch A.G.
      Phosphorylated Pol II CTD recruits multiple HDACs, including Rpd3C (S), for methylation-dependent deacetylation of ORF nucleosomes.
      ,
      • Li B.
      • Jackson J.
      • Simon M.D.
      • Fleharty B.
      • Gogol M.
      • Seidel C.
      • Workman J.L.
      • Shilatifard A.
      Histone H3 lysine 36 dimethylation (H3K36me2) is sufficient to recruit the Rpd3s histone deacetylase complex and to repress spurious transcription.
      ,
      • Youdell M.L.
      • Kizer K.O.
      • Kisseleva-Romanova E.
      • Fuchs S.M.
      • Duro E.
      • Strahl B.D.
      • Mellor J.
      Roles for Ctk1 and Spt6 in regulating the different methylation states of histone H3 lysine 36.
      ). This is required for the suppression of spurious transcriptional initiation from cryptic internal promoters (
      • Wagner E.J.
      • Carpenter P.B.
      Understanding the language of Lys36 methylation at histone H3.
      ). A similar regulatory mechanism is retained, although embellished, in human cells, where the mammalian ortholog of Eaf3p, MRG15, binds trimethylated H3K36 and can interact with a mammalian Rpd3S-like complex or the H3K4me2/3 demethylation machinery to coordinate transcription (
      • Xie L.
      • Pelz C.
      • Wang W.
      • Bashar A.
      • Varlamova O.
      • Shadle S.
      • Impey S.
      KDM5B regulates embryonic stem cell self-renewal and represses cryptic intragenic transcription.
      ,
      • Zhang P.
      • Du J.
      • Sun B.
      • Dong X.
      • Xu G.
      • Zhou J.
      • Huang Q.
      • Liu Q.
      • Hao Q.
      • Ding J.
      Structure of human MRG15 chromo domain and its binding to Lys36-methylated histone H3.
      ). Second, the Isw1b chromatin remodeling complex is recruited to H3K36 methylation by the proline-tryptophan-tryptophan-proline (PWWP) domain–containing subunit, Ioc4p (
      • Maltby V.E.
      • Martin B.J.
      • Schulze J.M.
      • Johnson I.
      • Hentrich T.
      • Sharma A.
      • Kobor M.S.
      • Howe L.
      Histone H3 lysine 36 methylation targets the Isw1b remodeling complex to chromatin.
      ,
      • Smolle M.
      • Venkatesh S.
      • Gogol M.M.
      • Li H.
      • Zhang Y.
      • Florens L.
      • Washburn M.P.
      • Workman J.L.
      Chromatin remodelers Isw1 and Chd1 maintain chromatin structure during transcription by preventing histone exchange.
      ), as evidenced by the co-localization of Isw1b with H3K36me at the mid- and 3′ regions of transcribed genes (Fig. 1). This complex works cooperatively with Rdp3S to re-establish a heterochromatic conformation following transcription in order to prevent production of intragenic transcripts. Third, in terms of the NuA3 HAT complex, the PWWP domain within its Pdp3p subunit selectively recognizes H3K36 trimethylation (Fig. 1) (
      • Gilbert T.M.
      • McDaniel S.L.
      • Byrum S.D.
      • Cades J.A.
      • Dancy B.C.
      • Wade H.
      • Tackett A.J.
      • Strahl B.D.
      • Taverna S.D.
      A PWWP domain-containing protein targets the NuA3 acetyltransferase complex via histone H3 lysine 36 trimethylation to coordinate transcriptional elongation at coding regions.
      ). In addition, the PHD finger in Nto1p of this complex has been shown to bind H3K36me3 in vitro (
      • Shi X.
      • Kachirskaia I.
      • Walter K.L.
      • Kuo J.-H.A.
      • Lake A.
      • Davrazou F.
      • Chan S.M.
      • Martin D.G.
      • Fingerman I.M.
      • Briggs S.D.
      Proteome-wide analysis in Saccharomyces cerevisiae identifies several PHD fingers as novel direct and selective binding modules of histone H3 methylated at either lysine 4 or lysine 36.
      ), and H3K36 methylation is necessary for NuA3 chromatin binding (
      • Martin D.G.
      • Grimes D.E.
      • Baetz K.
      • Howe L.
      Methylation of histone H3 mediates the association of the NuA3 histone acetyltransferase with chromatin.
      ). Taken together, these observations hint at a regulatory role for NuA3-mediated histone acetylation at actively transcribed gene bodies; however, this remains to be elucidated experimentally (
      • Wozniak G.G.
      • Strahl B.D.
      Hitting the ‘mark’: Interpreting lysine methylation in the context of active transcription.
      ).
      In normal yeast cells, H3K36 methylation is involved in the regulation of many genomic and transcriptomic processes, including DNA replication and repair (
      • Wagner E.J.
      • Carpenter P.B.
      Understanding the language of Lys36 methylation at histone H3.
      ,
      • Jha D.K.
      • Strahl B.D.
      An RNA polymerase II-coupled function for histone H3K36 methylation in checkpoint activation and DSB repair.
      ,
      • Pryde F.
      • Jain D.
      • Kerr A.
      • Curley R.
      • Mariotti F.R.
      • Vogelauer M.
      H3 k36 methylation helps determine the timing of cdc45 association with replication origins.
      ), 5-methylcytosine deposition (
      • Morselli M.
      • Pastor W.A.
      • Montanini B.
      • Nee K.
      • Ferrari R.
      • Fu K.
      • Bonora G.
      • Rubbi L.
      • Clark A.T.
      • Ottonello S.
      In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse.
      ), and pre-mRNA splicing (
      • Hérissant L.
      • Moehle E.A.
      • Bertaccini D.
      • Van Dorsselaer A.
      • Schaeffer-Reiss C.
      • Guthrie C.
      • Dargemont C.
      H2B ubiquitylation modulates spliceosome assembly and function in budding yeast.
      ,
      • Leung C.S.
      • Douglass S.M.
      • Morselli M.
      • Obusan M.B.
      • Pavlyukov M.S.
      • Pellegrini M.
      • Johnson T.L.
      H3K36 methylation and the chromodomain protein Eaf3 are required for proper cotranscriptional spliceosome assembly.
      ,
      • Sorenson M.R.
      • Jha D.K.
      • Ucles S.A.
      • Flood D.M.
      • Strahl B.D.
      • Stevens S.W.
      • Kress T.L.
      Histone H3K36 methylation regulates pre-mRNA splicing in Saccharomyces cerevisiae.
      ). Of interest, these distinct functions are linked to unique methylation states of H3K36. Appropriate temporal patterning of H3K36me3 and H3K36me2 around DSBs is required for maintenance and repair of chromatin structure at DNA damage sites (
      • Jha D.K.
      • Strahl B.D.
      An RNA polymerase II-coupled function for histone H3K36 methylation in checkpoint activation and DSB repair.
      ), whereas H3K36me1 has been reported to regulate the formation of DNA replication origins (
      • Pryde F.
      • Jain D.
      • Kerr A.
      • Curley R.
      • Mariotti F.R.
      • Vogelauer M.
      H3 k36 methylation helps determine the timing of cdc45 association with replication origins.
      ) via an unknown reader protein. It has also been shown that H3K36 methylation is associated with yeast cellular aging (
      • Ryu H.-Y.
      • Rhie B.-H.
      • Ahn S.H.
      Loss of the Set2 histone methyltransferase increases cellular lifespan in yeast cells.
      ), and deficits in sustaining this modification over time are related to increased cryptic transcription at certain loci in older cells (
      • Sen P.
      • Dang W.
      • Donahue G.
      • Dai J.
      • Dorsey J.
      • Cao X.
      • Liu W.
      • Cao K.
      • Perry R.
      • Lee J.Y.
      H3K36 methylation promotes longevity by enhancing transcriptional fidelity.
      ). As with other histone methyl marks, disruption of H3K36me results in mild-to-severe growth and nutritional phenotypes in S. cerevisiae. H3K36me-null cells are sensitive to nutrient stress (
      • McDaniel S.L.
      • Hepperla A.J.
      • Huang J.
      • Dronamraju R.
      • Adams A.T.
      • Kulkarni V.G.
      • Davis I.J.
      • Strahl B.D.
      H3K36 methylation regulates nutrient stress response in Saccharomyces cerevisiae by enforcing transcriptional fidelity.
      ) and have a shortened life span due to aberrant initiation of cryptic transcription within gene bodies (
      • Li J.
      • Ahn J.H.
      • Wang G.G.
      Understanding histone H3 lysine 36 methylation and its deregulation in disease.
      ). Antisense transcripts produced from such spurious events impair gene expression from the sense strand (
      • McDaniel S.L.
      • Hepperla A.J.
      • Huang J.
      • Dronamraju R.
      • Adams A.T.
      • Kulkarni V.G.
      • Davis I.J.
      • Strahl B.D.
      H3K36 methylation regulates nutrient stress response in Saccharomyces cerevisiae by enforcing transcriptional fidelity.
      ,
      • Venkatesh S.
      • Li H.
      • Gogol M.M.
      • Workman J.L.
      Selective suppression of antisense transcription by Set2-mediated H3K36 methylation.
      ), and thus several yeast strains deficient in H3K36 methylation are synthetically sick or lethal with transcriptional elongation mutants (
      • Fuchs S.M.
      • Kizer K.O.
      • Braberg H.
      • Krogan N.J.
      • Strahl B.D.
      RNA polymerase II carboxyl-terminal domain phosphorylation regulates protein stability of the Set2 methyltransferase and histone H3 di-and trimethylation at lysine 36.
      ).
      Cross talk between H3K36 methylation and other epigenetic modifications occurs in budding yeast cells. Chromatin immunoprecipitation sequencing experiments have shown that genome-wide occupancy of H3K36ac and H3K36me are inversely related (
      • Morris S.A.
      • Rao B.
      • Garcia B.A.
      • Hake S.B.
      • Diaz R.L.
      • Shabanowitz J.
      • Hunt D.F.
      • Allis C.D.
      • Lieb J.D.
      • Strahl B.D.
      Identification of histone H3 lysine 36 acetylation as a highly conserved histone modification.
      ), raising the fascinating prospect that acetylation and methylation of a single histone lysine residue may display functional interplay to mediate chromatin-templated processes. Indeed, an acetyl/methyl switch at H3K36 has been found to control DSB repair pathway choice in fission yeast whereby trimethylation, in contrast to its role in transcriptional activation, compacts chromatin and promotes NHEJ, while counteracting Gcn5-dependent acetylation enhances chromatin accessibility and encourages repair by homologous recombination (
      • Pai C.-C.
      • Deegan R.S.
      • Subramanian L.
      • Gal C.
      • Sarkar S.
      • Blaikley E.J.
      • Walker C.
      • Hulme L.
      • Bernhard E.
      • Codlin S.
      A histone H3K36 chromatin switch coordinates DNA double-strand break repair pathway choice.
      ). Other examples of interplay include the trans-regulation of H3K36me deposition by H4K44 (
      • Du H.-N.
      • Fingerman I.M.
      • Briggs S.D.
      Histone H3 K36 methylation is mediated by a trans-histone methylation pathway involving an interaction between Set2 and histone H4.
      ) and H2AL116/L117 (
      • Du H.-N.
      • Briggs S.D.
      A nucleosome surface formed by histone H4, H2A, and H3 residues is needed for proper histone H3 Lys36 methylation, histone acetylation, and repression of cryptic transcription.
      ) residues (Fig. 2B). In these cases, the unmodified H4 and H2A amino acids are required for the correct positioning of the H3K36 methylation machinery on nucleosomal substrates. Unlike methylation of H3K4 and H3K79, which require prior H2BK123ub, ubiquitination acts as a negative effector of H3K36 methylation (
      • Batta K.
      • Zhang Z.
      • Yen K.
      • Goffman D.B.
      • Pugh B.F.
      Genome-wide function of H2B ubiquitylation in promoter and genic regions.
      ). This occurs indirectly through the Ctk1p kinase, which needs H2BK123 to be deubiquitinated by SAGA-associated Ubp8p in order to phosphorylate the C-terminal domain (CTD) of RNAPII (
      • Wyce A.
      • Xiao T.
      • Whelan K.A.
      • Kosman C.
      • Walter W.
      • Eick D.
      • Hughes T.R.
      • Krogan N.J.
      • Strahl B.D.
      • Berger S.L.
      H2B ubiquitylation acts as a barrier to Ctk1 nucleosomal recruitment prior to removal by Ubp8 within a SAGA-related complex.
      ). Another negative regulator of H3K36 methylation is the histone proline isomerase, Fpr4p, which alters the adjacent H3P38 residue into a configuration that renders H3K36 unsuitable for trimethylation (Fig. 2B) (
      • Nelson C.J.
      • Santos-Rosa H.
      • Kouzarides T.
      Proline isomerization of histone H3 regulates lysine methylation and gene expression.
      ).

      H3K79me1/2/3

      Histone H3 lysine 79 (H3K79) is a conserved eukaryotic methylation site that is uniquely located within the globular core of the nucleosomal architecture (
      • Ng H.H.
      • Feng Q.
      • Wang H.
      • Erdjument-Bromage H.
      • Tempst P.
      • Zhang Y.
      • Struhl K.
      Lysine methylation within the globular domain of histone H3 by Dot1 is important for telomeric silencing and Sir protein association.
      ), as opposed to on a histone tail (Fig. 1). H3K79 exists in three methylation states; monomethyl (H3K79me1), dimethyl (H3K79me2), and trimethyl (H3K79me3), the latter of which is the most prevalent in vivo (~50%) (
      • van Leeuwen F.
      • Gafken P.R.
      • Gottschling D.E.
      Dot1p modulates silencing in yeast by methylation of the nucleosome core.
      ). The distinct functions and genomic distributions of these methylation states, however, are poorly understood. Genome-wide maps have revealed that H3K79 methylation broadly occurs in a uniform fashion throughout the coding region of actively transcribed genes (Fig. 2A) (
      • Pokholok D.K.
      • Harbison C.T.
      • Levine S.
      • Cole M.
      • Hannett N.M.
      • Lee T.I.
      • Bell G.W.
      • Walker K.
      • Rolfe P.A.
      • Herbolsheimer E.
      Genome-wide map of nucleosome acetylation and methylation in yeast.
      • Ng H.H.
      • Ciccone D.N.
      • Morshead K.B.
      • Oettinger M.A.
      • Struhl K.
      Lysine-79 of histone H3 is hypomethylated at silenced loci in yeast and mammalian cells: A potential mechanism for position-effect variegation.
      ,
      • Shahbazian M.D.
      • Zhang K.
      • Grunstein M.
      Histone H2B ubiquitylation controls processive methylation but not monomethylation by Dot1 and Set1.
      ) and is thus associated with transcriptional activation. Curiously, in S. cerevisiae, the vast majority (~90%) of H3K79 is methylated; however, in mammals, H3K79 is predominantly unmodified (
      • van Leeuwen F.
      • Gafken P.R.
      • Gottschling D.E.
      Dot1p modulates silencing in yeast by methylation of the nucleosome core.
      ). Akin to other epigenetic marks, H3K79 methylation patterning across the yeast genome is dynamic. Levels of H3K79me3 remain unchanged throughout the cell cycle, whereas H3K79me2 levels increase gradually through the G1/S and G2/M phase transitions (
      • Farooq Z.
      • Banday S.
      • Pandita T.K.
      • Altaf M.
      The many faces of histone H3K79 methylation.
      ).
      The precise mechanism by which H3K79 methylation regulates transcription and other biological processes is still an open question in the field of epigenetics (
      • Wozniak G.G.
      • Strahl B.D.
      Hitting the ‘mark’: Interpreting lysine methylation in the context of active transcription.
      ). This is largely due to a paucity in understanding of reader domains and proteins specific to this methylated residue (
      • Kim J.
      • Daniel J.
      • Espejo A.
      • Lake A.
      • Krishna M.
      • Xia L.
      • Zhang Y.
      • Bedford M.T.
      Tudor, MBT and chromo domains gauge the degree of lysine methylation.
      ,
      • Botuyan M.V.
      • Lee J.
      • Ward I.M.
      • Kim J.-E.
      • Thompson J.R.
      • Chen J.
      • Mer G.
      Structural basis for the methylation state-specific recognition of histone H4-K20 by 53BP1 and Crb2 in DNA repair.
      ). In budding yeast, H3K79 methylation is known to interact with two major effector proteins, which, in turn, coordinate two major cellular processes: (1) transcription and (2) DNA repair (Fig. 1). With respect to transcriptional regulation, H3K79 methylation is required for the proper formation of heterochromatin at telomeric and centromeric regions of the chromosome. This is mediated by the recruitment of Sir (silent information regulator) proteins and their cognate SIR HDAC complex (
      • Norris A.
      • Boeke J.D.
      Silent information regulator 3: The Goldilocks of the silencing complex.
      ). The presence of all three methylation states of H3K79 inhibits the binding of the bromo-adjacent homology (BAH) domain of Sir3p and prevents assembly of the repressive SIR complex and its resultant deacetylation within euchromatic regions (Fig. 1) (
      • Altaf M.
      • Utley R.T.
      • Lacoste N.
      • Tan S.
      • Briggs S.D.
      • Côté J.
      Interplay of chromatin modifiers on a short basic patch of histone H4 tail defines the boundary of telomeric heterochromatin.
      ,
      • Onishi M.
      • Liou G.-G.
      • Buchberger J.R.
      • Walz T.
      • Moazed D.
      Role of the conserved Sir3-BAH domain in nucleosome binding and silent chromatin assembly.
      ). Accordingly, defective H3K79 methylation causes aberrant redistribution of Sir proteins to actively transcribed genes and thereby impairs proper silencing of telomeres and cryptic mating-type (HML/HMR) loci (
      • van Leeuwen F.
      • Gafken P.R.
      • Gottschling D.E.
      Dot1p modulates silencing in yeast by methylation of the nucleosome core.
      ,
      • Ng H.H.
      • Xu R.-M.
      • Zhang Y.
      • Struhl K.
      Ubiquitination of histone H2B by Rad6 is required for efficient Dot1-mediated methylation of histone H3 lysine 79.
      ). The interplay between Sir proteins and histone methylation is bidirectional, as Sir3p competes with the H3K79 methylation machinery for binding to a basic patch of histone H4 (
      • Altaf M.
      • Utley R.T.
      • Lacoste N.
      • Tan S.
      • Briggs S.D.
      • Côté J.
      Interplay of chromatin modifiers on a short basic patch of histone H4 tail defines the boundary of telomeric heterochromatin.
      ,
      • Fingerman I.M.
      • Li H.-C.
      • Briggs S.D.
      A charge-based interaction between histone H4 and Dot1 is required for H3K79 methylation and telomere silencing: Identification of a new trans-histone pathway.
      ), as discussed in detail later. Of interest, the importance of fluctuations in nucleosomal occupancy of Sir proteins has been questioned (
      • Kitada T.
      • Kuryan B.G.
      • Tran N.N.H.
      • Song C.
      • Xue Y.
      • Carey M.
      • Grunstein M.
      Mechanism for epigenetic variegation of gene expression at yeast telomeric heterochromatin.
      ), suggesting that there are additional layers of complexity to be uncovered.
      In addition to its canonical role in transcription, H3K79 methylation has also been shown to mediate passage through several key checkpoints during yeast cellular growth and reproduction and in response to stress. First, this modification regulates the DNA damage checkpoints throughout the cell cycle by recruiting the checkpoint adaptor protein, Rad9p, via its methyl-binding Tudor domain (Fig. 1) (
      • Wysocki R.
      • Javaheri A.
      • Allard S.
      • Sha F.
      • Côté J.
      • Kron S.J.
      Role of Dot1-dependent histone H3 methylation in G1 and S phase DNA damage checkpoint functions of Rad9.
      ). This interaction inhibits the production of single-stranded DNA at DSBs and at uncapped telomeres, suggesting that H3K79 plays a role in the resection of damaged DNA and its repair by homologous recombination (
      • Farooq Z.
      • Banday S.
      • Pandita T.K.
      • Altaf M.
      The many faces of histone H3K79 methylation.
      ,
      • Lazzaro F.
      • Sapountzi V.
      • Granata M.
      • Pellicioli A.
      • Vaze M.
      • Haber J.E.
      • Plevani P.
      • Lydall D.
      • Muzi-Falconi M.
      Histone methyltransferase Dot1 and Rad9 inhibit single-stranded DNA accumulation at DSBs and uncapped telomeres.
      ). Moreover, Rad9p is critical in the maintenance of single-stranded DNA during NHEJ in late G2 phase (
      • Lazzaro F.
      • Sapountzi V.
      • Granata M.
      • Pellicioli A.
      • Vaze M.
      • Haber J.E.
      • Plevani P.
      • Lydall D.
      • Muzi-Falconi M.
      Histone methyltransferase Dot1 and Rad9 inhibit single-stranded DNA accumulation at DSBs and uncapped telomeres.
      ). The human ortholog of Rad9p, 53BP1, is similarly recruited to DSBs by H3K79 methylation (
      • Huyen Y.
      • Zgheib O.
      • DiTullio Jr., R.A.
      • Gorgoulis V.G.
      • Zacharatos P.
      • Petty T.J.
      • Sheston E.A.
      • Mellert H.S.
      • Stavridi E.S.
      • Halazonetis T.D.
      Methylated lysine 79 of histone H3 targets 53BP1 to DNA double-strand breaks.
      ). H3K79 methylation also plays a crucial role in progression through the pachytene checkpoint during meiosis (
      • Farooq Z.
      • Banday S.
      • Pandita T.K.
      • Altaf M.
      The many faces of histone H3K79 methylation.
      ). Although H3K79 methylation is virtually dispensable for unperturbed meiosis, it is essential in coordinating the checkpoint response to unrepaired DSBs and synapsis defects in certain yeast meiotic mutants (
      • Ontoso D.
      • Acosta I.
      • van Leeuwen F.
      • Freire R.
      • San-Segundo P.A.
      Dot1-dependent histone H3K79 methylation promotes activation of the Mek1 meiotic checkpoint effector kinase by regulating the Hop1 adaptor.
      ,
      • San-Segundo P.A.
      • Roeder G.S.
      Role for the silencing protein Dot1 in meiotic checkpoint control.
      ). This has been shown to occur through the chromosomal recruitment of Hop1p to H3K79me, which then activates Mek1p kinase in response to meiotic DNA damage (Fig. 1) (
      • Ontoso D.
      • Acosta I.
      • van Leeuwen F.
      • Freire R.
      • San-Segundo P.A.
      Dot1-dependent histone H3K79 methylation promotes activation of the Mek1 meiotic checkpoint effector kinase by regulating the Hop1 adaptor.
      ). Unsurprisingly, yeast cells with deficient H3K79 methylation exhibit increased sensitivity to ionizing radiation (
      • Bostelman L.J.
      • Keller A.M.
      • Albrecht A.M.
      • Arat A.
      • Thompson J.S.
      Methylation of histone H3 lysine-79 by Dot1p plays multiple roles in the response to UV damage in Saccharomyces cerevisiae.
      ) and are unable to initiate DNA damage repair (
      • Giannattasio M.
      • Lazzaro F.
      • Plevani P.
      • Muzi-Falconi M.
      The DNA damage checkpoint response requires histone H2B ubiquitination by Rad6-Bre1 and H3 methylation by Dot1.
      ) and meiotic recombination checkpoint functions (
      • San-Segundo P.A.
      • Roeder G.S.
      Role for the silencing protein Dot1 in meiotic checkpoint control.
      ).
      Methylation at H3K79 can be positively and negatively regulated by several histone PTMs through cross talk. Analogous to H3K4me, monoubiquitination of H2BK123 by the E2-E3 complex Rad6p/Bre1p is a prerequisite for H3K79 trimethylation in S. cerevisiae (Fig. 2B) (
      • Shahbazian M.D.
      • Zhang K.
      • Grunstein M.
      Histone H2B ubiquitylation controls processive methylation but not monomethylation by Dot1 and Set1.
      ). These modifications lie in spatial proximity on the same exposed nucleosome surface, thus providing a structural basis for their interplay (
      • Nguyen A.T.
      • Zhang Y.
      The diverse functions of Dot1 and H3K79 methylation.
      ). Of importance, this cross talk is conserved in mammalian systems wherein dimethylation of H3K79 is stimulated by ubiquitination of H2BK120, which is equivalent to yeast H2BK123 (
      • Zhu B.
      • Zheng Y.
      • Pham A.-D.
      • Mandal S.S.
      • Erdjument-Bromage H.
      • Tempst P.
      • Reinberg D.
      Monoubiquitination of human histone H2B: The factors involved and their roles in HOX gene regulation.
      ). Although this was initially considered to be a unidirectional effect (
      • Sun Z.-W.
      • Allis C.D.
      Ubiquitination of histone H2B regulates H3 methylation and gene silencing in yeast.
      ), it has since been shown that the H3K79 methyltransferase Dot1p promotes H2B ubiquitination via its N-terminal region, independent of its catalytic activity (
      • van Welsem T.
      • Korthout T.
      • Ekkebus R.
      • Morais D.
      • Molenaar T.M.
      • van Harten K.
      • Poramba-Liyanage D.W.
      • Sun S.M.
      • Lenstra T.L.
      • Srivas R.
      Dot1 promotes H2B ubiquitination by a methyltransferase-independent mechanism.
      ). Cross talk between ubiquitination and H3K79 methylation is further complicated by the input of the Rpd3L HDAC complex, which deacetylates its target genes in transcriptionally repressed regions devoid of H2BK123ub1 (
      • Vlaming H.
      • McLean C.M.
      • Korthout T.
      • Alemdehy M.F.
      • Hendriks S.
      • Lancini C.
      • Palit S.
      • Klarenbeek S.
      • Kwesi-Maliepaard E.M.
      • Molenaar T.M.
      Conserved crosstalk between histone deacetylation and H3K79 methylation generates DOT1L-dose dependency in HDAC1-deficient thymic lymphoma.
      ). As such, a subset of yeast genes have lower H3K79me3 and gene expression owing to antagonistic Rpd3L activity, a regulatory mechanism that is retained by human HDAC1 (
      • Vlaming H.
      • McLean C.M.
      • Korthout T.
      • Alemdehy M.F.
      • Hendriks S.
      • Lancini C.
      • Palit S.
      • Klarenbeek S.
      • Kwesi-Maliepaard E.M.
      • Molenaar T.M.
      Conserved crosstalk between histone deacetylation and H3K79 methylation generates DOT1L-dose dependency in HDAC1-deficient thymic lymphoma.
      ). Finally, acetylation at H4K16 in yeast by either Sas2p or Esa1p indirectly promotes H3K79 methylation through steric hindrance of Sir3p H4 binding (Fig. 2B) (
      • Altaf M.
      • Utley R.T.
      • Lacoste N.
      • Tan S.
      • Briggs S.D.
      • Côté J.
      Interplay of chromatin modifiers on a short basic patch of histone H4 tail defines the boundary of telomeric heterochromatin.
      ).

      H4K5/8/12me1

      It has recently been discovered that a cluster of lysine residues within the histone H4 N-terminal tail are subject to methylation in S. cerevisiae. In 2012, the laboratory of Or Gozani used systematic mutagenesis of the H4 tail, immunoblotting, and tandem mass spectrometry to identify monomethylation sites at H4K5, H4K8, and H4K12 in growing yeast cells (Fig. 1) (
      • Green E.M.
      • Mas G.
      • Young N.L.
      • Garcia B.A.
      • Gozani O.
      Methylation of H4 lysines 5, 8 and 12 by yeast Set5 calibrates chromatin stress responses.
      ). It is interesting that no evidence for di- or tri-methylation at these sites was found. H4K5 methylation is functionally conserved in mammalian cells where it is controlled by SMYD3 (
      • Van Aller G.S.
      • Reynoird N.
      • Barbash O.
      • Huddleston M.
      • Liu S.
      • Zmoos A.-F.
      • McDevitt P.
      • Sinnamon R.
      • Le B.
      • Mas G.
      Smyd3 regulates cancer cell phenotypes and catalyzes histone H4 lysine 5 methylation.
      ) and its dysregulation contributes to tumorigenesis (
      • Hamamoto R.
      • Furukawa Y.
      • Morita M.
      • Iimura Y.
      • Silva F.P.
      • Li M.
      • Yagyu R.
      • Nakamura Y.
      SMYD3 encodes a histone methyltransferase involved in the proliferation of cancer cells.
      ,
      • Van Aller G.S.
      • Reynoird N.
      • Barbash O.
      • Huddleston M.
      • Liu S.
      • Zmoos A.-F.
      • McDevitt P.
      • Sinnamon R.
      • Le B.
      • Mas G.
      Smyd3 regulates cancer cell phenotypes and catalyzes histone H4 lysine 5 methylation.
      ,
      • Hamamoto R.
      • Silva F.P.
      • Tsuge M.
      • Nishidate T.
      • Katagiri T.
      • Nakamura Y.
      • Furukawa Y.
      Enhanced SMYD3 expression is essential for the growth of breast cancer cells.
      ,
      • Wang S.-z.
      • Luo X.-g.
      • Shen J.
      • Zou J.-n.
      • Lu Y.-h.
      • Xi T.
      Knockdown of SMYD3 by RNA interference inhibits cervical carcinoma cell growth and invasion in vitro.
      ). Methylation sites at H4K8 and H4K12, however, are not retained in mammalian systems, suggesting they may serve specific functions in budding yeast. Although these methylation sites are newly discovered in comparison with other epigenetic modifications, their identification has added new layers of functionality to the H4 tail (
      • Green E.M.
      • Morrison A.J.
      • Gozani O.
      New marks on the block: Set5 methylates H4 lysines 5, 8 and 12.
      ). Targeted studies have shown that, although loss of H4K5/8/12me1 resulted in only minor changes in global gene expression, these modifications play an important role in determining cellular fitness and responses to environmental stress (
      • Green E.M.
      • Mas G.
      • Young N.L.
      • Garcia B.A.
      • Gozani O.
      Methylation of H4 lysines 5, 8 and 12 by yeast Set5 calibrates chromatin stress responses.
      ). Unique from other histone methylation sites, these three lysine residues can functionally compensate for one another, indicating that they are unlikely to recruit different chromatin modifiers (Fig. 1) (
      • Bird A.W.
      • David Y.Y.
      • Pray-Grant M.G.
      • Qiu Q.
      • Harmon K.E.
      • Megee P.C.
      • Grant P.A.
      • Smith M.M.
      • Christman M.F.
      Acetylation of histone H4 by Esa1 is required for DNA double-strand break repair.
      ,
      • Dion M.F.
      • Altschuler S.J.
      • Wu L.F.
      • Rando O.J.
      Genomic characterization reveals a simple histone H4 acetylation code.
      ,
      • Ma X.-J.
      • Wu J.
      • Altheim B.A.
      • Schultz M.C.
      • Grunstein M.
      Deposition-related sites K5/K12 in histone H4 are not required for nucleosome deposition in yeast.
      ). With respect to PTM cross talk, it appears that H4 methylation sites cooperatively function with methylation at H3K4 as yeast strains deficient in H3K4 methylation and methylation at any of the H4 sites are sensitive to cellular stress. Moreover, the possibility of methylation/acetylation interplay has been suggested because H4K5, H4K8, and H4K12 are also known to be acetylated by Esa1p, a constituent of the NuA4 HAT complex in yeast (
      • Allard S.
      • Utley R.T.
      • Savard J.
      • Clarke A.
      • Grant P.
      • Brandl C.J.
      • Pillus L.
      • Workman J.L.
      • Côté J.
      NuA4, an essential transcription adaptor/histone H4 acetyltransferase complex containing Esa1p and the ATM-related cofactor Tra1p.
      • Clarke A.S.
      • Lowell J.E.
      • Jacobson S.J.
      • Pillus L.
      Esa1p is an essential histone acetyltransferase required for cell cycle progression.
      ). Despite much progress, the exact molecular mechanisms underpinning H4K5/8/12me1 function and cross talk with H3K4 methylation and H4K5/8/12 acetylation are not well understood, and no reader proteins specific for this mark have been confirmed (Fig. 1). Considerable work is required to clarify the role of monomethylation sites at H4K5/8/12 in chromatin structure and function in yeast.

      Histone methyltransferases

      Despite the landmark discovery of histone methylation over 50 years ago (
      • Allfrey V.G.
      • Mirsky A.E.
      Structural modifications of histones and their possible role in the regulation of RNA synthesis.
      ), it was not until 2000 that the first histone methyltransferase was identified (
      • Rea S.
      • Eisenhaber F.
      • O'carroll D.
      • Strahl B.D.
      • Sun Z.-W.
      • Schmid M.
      • Opravil S.
      • Mechtler K.
      • Ponting C.P.
      • Allis C.D.
      Regulation of chromatin structure by site-specific histone H3 methyltransferases.
      ). This methyltransferase, SUV39H1, was shown to specifically trimethylate H3K9 in human cells where it controls the formation of repressive heterochromatin at pericentric and telomeric regions (
      • Peters A.H.
      • O'Carroll D.
      • Scherthan H.
      • Mechtler K.
      • Sauer S.
      • Schöfer C.
      • Weipoltshammer K.
      • Pagani M.
      • Lachner M.
      • Kohlmaier A.
      Loss of the Suv39h histone methyltransferases impairs mammalian heterochromatin and genome stability.
      ). Since then, numerous methyltransferases that target basic residues on histone proteins, particularly within their disordered N-terminal tails, have been identified in yeast and in other eukaryotes. Broadly, there are two evolutionarily conserved enzymatic families that catalyze the transfer of methyl group(s) from the metabolic donor S-adenosyl-L-methionine (AdoMet, also known as SAM) to the ε-amino group of lysine side chains on histone proteins (
      • Guo H.-B.
      • Guo H.
      Mechanism of histone methylation catalyzed by protein lysine methyltransferase SET7/9 and origin of product specificity.
      ). SET (Su(var)3-9, Enhancer of Zeste, Trithorax) domain methyltransferases harbor a SET catalytic domain, which forms a knot-like β-sheet structure that facilitates methyl transfer (
      • Cheng X.
      • Collins R.E.
      • Zhang X.
      Structural and sequence motifs of protein (histone) methylation enzymes.
      ,
      • Lanouette S.
      • Mongeon V.
      • Figeys D.
      • Couture J.F.
      The functional diversity of protein lysine methylation.
      ). In S. cerevisiae, there are three SET domain–containing proteins that have bona fide histone methyltransferase activity: Set1p, Set2p, and Set5p, all of which methylate histones on their N-terminal tails (Fig. 1, Table 1) (
      • Cheng X.
      • Collins R.E.
      • Zhang X.
      Structural and sequence motifs of protein (histone) methylation enzymes.
      ,
      • Lanouette S.
      • Mongeon V.
      • Figeys D.
      • Couture J.F.
      The functional diversity of protein lysine methylation.
      ). The seven-β-strand (7βS) family of methyltransferases is more diverse than the SET family, comprising both lysine and arginine protein methyltransferases, as well as DNA methyltransferases (
      • Husmann D.
      • Gozani O.
      Histone lysine methyltransferases in biology and disease.
      ). Dot1p is the sole 7βS histone lysine methyltransferase in budding yeast and uniquely methylates histone H3 within its globular core (Fig. 1, Table 1) (
      • Feng Q.
      • Wang H.
      • Ng H.H.
      • Erdjument-Bromage H.
      • Tempst P.
      • Struhl K.
      • Zhang Y.
      Methylation of H3-lysine 79 is mediated by a new family of HMTases without a SET domain.
      ,
      • Lacoste N.
      • Utley R.T.
      • Hunter J.M.
      • Poirier G.G.
      • Côté J.
      Disruptor of telomeric silencing-1 is a chromatin-specific histone H3 methyltransferase.
      ). In this section, we discuss the structure, function, and conservation of the four histone lysine methyltransferases in S. cerevisiae and highlight recent efforts to understand their regulation.

      Set1p (COMPASS)

      SET domain-containing 1 (Set1p), also known as KMT2 or YTX1, was the first histone lysine methyltransferase to be discovered in S. cerevisiae. Early studies used sequence homology approaches to identify Set1p as a yeast member of the Trithorax gene family and revealed that, although not essential for viability, it plays key roles in the regulation of transcriptional silencing at mating-type loci and telomeres, in the maintenance of telomere length (
      • Nislow C.
      • Ray E.
      • Pillus L.
      SET1, a yeast member of the trithorax family, functions in transcriptional silencing and diverse cellular processes.
      ) and in DNA repair (
      • Corda Y.
      • Schramke V.
      • Longhese M.P.
      • Smokvina T.
      • Paciotti V.
      • Brevet V.
      • Gilson E.
      • Géli V.
      Interaction between Set1p and checkpoint protein Mec3p in DNA repair and telomere functions.
      • Schramke V.
      • Neecke H.
      • Brevet V.
      • Corda Y.
      • Lucchini G.
      • Longhese M.P.
      • Gilson E.
      • Géli V.
      The set1Δ mutation unveils a novel signaling pathway relayed by the Rad53-dependent hyperphosphorylation of replication protein A that leads to transcriptional activation of repair genes.
      ). It was not until 2002 that the methyltransferase function of Set1p was investigated; Briggs et al. demonstrated that deletion of SET1 completely abolishes H3K4 methylation in vivo, manifesting in aberrant transcription at rDNA loci and a slow-growth phenotype (
      • Briggs S.D.
      • Bryk M.
      • Strahl B.D.
      • Cheung W.L.
      • Davie J.K.
      • Dent S.Y.
      • Winston F.
      • Allis C.D.
      Histone H3 lysine 4 methylation is mediated by Set1 and required for cell growth and rDNA silencing in Saccharomyces cerevisiae.
      ). A number of groups have since confirmed, both in vivo and in vitro, that Set1p is the sole enzyme responsible for all three states of H3K4 methylation (Table 1) (
      • Krogan N.J.
      • Dover J.
      • Khorrami S.
      • Greenblatt J.F.
      • Schneider J.
      • Johnston M.
      • Shilatifard A.
      COMPASS, a histone H3 (Lysine 4) methyltransferase required for telomeric silencing of gene expression.
      ,
      • Miller T.
      • Krogan N.J.
      • Dover J.
      • Erdjument-Bromage H.
      • Tempst P.
      • Johnston M.
      • Greenblatt J.F.
      • Shilatifard A.
      Compass: A complex of proteins associated with a trithorax-related SET domain protein.
      ,
      • Shilatifard A.
      The COMPASS family of histone H3K4 methylases: Mechanisms of regulation in development and disease pathogenesis.
      ). Set1p is the largest histone methyltransferase in the yeast proteome, at 1080 amino acids in length, and comprises a SET catalytic domain (residues 938–1055) and a post-SET domain (residues 1064–1080) (Fig. 3). Set1p also harbors, in addition to its catalytic regions, several regulatory domains that control its enzymatic activity and interactions. An N-SET domain (residues 752–928) has been reported to enable the cross talk between Set1p-mediated H3K4 methylation and H2B ubiquitination (
      • Kim J.
      • Kim J.-A.
      • McGinty R.K.
      • Nguyen U.T.
      • Muir T.W.
      • Allis C.D.
      • Roeder R.G.
      The n-SET domain of Set1 regulates H2B ubiquitylation-dependent H3K4 methylation.
      ), although these findings have been challenged (Fig. 3) (
      • Thornton J.L.
      • Westfield G.H.
      • Takahashi Y.-h.
      • Cook M.
      • Gao X.
      • Woodfin A.R.
      • Lee J.-S.
      • Morgan M.A.
      • Jackson J.
      • Smith E.R.
      Context dependency of Set1/COMPASS-mediated histone H3 Lys4 trimethylation.
      ). A highly conserved tandem RNA recognition motif, comprising RRM1 (residues 274–375, Protein Data Bank [PDB] ID: 2J8A (
      • Trésaugues L.
      • Dehé P.-M.
      • Guérois R.
      • Rodriguez-Gil A.
      • Varlet I.
      • Salah P.
      • Pamblanco M.
      • Luciano P.
      • Quevillon-Cheruel S.
      • Sollier J.
      Structural characterization of Set1 RNA recognition motifs and their role in histone H3 lysine 4 methylation.
      )) and RRM2 (residues 376–579) toward the N-terminus of Set1p, is required for its capacity to trimethylate H3K4 (
      • Fingerman I.M.
      • Wu C.-L.
      • Wilson B.D.
      • Briggs S.D.
      Global loss of Set1-mediated H3 Lys4 trimethylation is associated with silencing defects in Saccharomyces cerevisiae.
      ) but is dispensable for dimethylation (Fig. 3) (
      • Schlichter A.
      • Cairns B.R.
      Histone trimethylation by Set1 is coordinated by the RRM, autoinhibitory, and catalytic domains.
      ). This domain, as well as N-SET, allows Set1p to bind RNA in vitro and to interact with nascent transcripts in vivo (
      • Luciano P.
      • Jeon J.
      • El-Kaoutari A.
      • Challal D.
      • Bonnet A.
      • Barucco M.
      • Candelli T.
      • Jourquin F.
      • Lesage P.
      • Kim J.
      Binding to RNA regulates Set1 function.
      ). The positive regulatory effects of RRM are counterbalanced by a semiconserved and centrally located autoinhibitory domain (AID), which attenuates Set1p trimethyltransferase function (
      • Schlichter A.
      • Cairns B.R.
      Histone trimethylation by Set1 is coordinated by the RRM, autoinhibitory, and catalytic domains.
      ). The precise residues that comprise Set1p AID are unknown and have thus been omitted from Figure 3; however, arginine 483 within this central region has been shown to be essential for autoinhibition.
      Figure thumbnail gr3
      Figure 3Domain architecture and structural features of yeast histone methyltransferase and demethylase enzymes. Linear sequence maps of yeast histone methyltransferase (left panel) and demethylase (right panel) enzymes. Protein domains are displayed, to scale, for each enzyme. Methyltransferase and demethylase domains are shown in green and pink, respectively, whereas other regulatory and interaction domains are colored in blue. Amino acid (aa) residues that are critical for enzymatic activity are shown in crimson. To date, partial crystal structures have been resolved for the RNA recognition motif (RRM; Protein Data Bank [PDB] ID: 2J8A) and the SET methyltransferase domain (PDB ID: 6BX3) of Set1p, the tryptophan–tryptophan (WW; PDB ID: 1E0N) and Set2 Rbp1 interacting (SRI; PDB ID: 2C5Z) domains of Set2p, the DOT1 methyltransferase domain (PDB ID: 1U2Z) of Dot1p, and the JmjN and JmjC demethylase domains (PDB ID: 3OPW) of Rph1p. Structures are depicted as ribbon diagrams in inset boxes and colored according to the region of the linear sequence map to which they correspond. AID, autoinhibitory domain; AWS, associated with SET; C2H2, Cys2-His2; MYND, myeloid translocation protein, Nervy, Deaf; NLS, nuclear localization signal; PHD, plant homeodomain.
      Set1p is the only yeast histone methyltransferase that forms a catalytically active multimeric complex in vivo. It associates with seven other protein subunits, Bre2p (Cps60), Sdc1p (Cps25), Shg1p (Cps15), Spp1p (Cps40), Swd1p (Cps50), Swd2p (Cps35), and Swd3p (Cps30), to form a H3K4 methyltransferase complex known as COMPASS (complex of proteins associated with Set1) (
      • Soares L.M.
      • Buratowski S.
      Yeast Swd2 is essential because of antagonism between Set1 histone methyltransferase complex and APT (associated with Pta1) termination factor.
      ). Strikingly, of these subunits, only Swd2p is essential for yeast cell viability; however, this is likely due to its additional function within the RNA 3′-end processing and termination complex, APT (
      • Bae H.J.
      • Dubarry M.
      • Jeon J.
      • Soares L.M.
      • Dargemont C.
      • Kim J.
      • Geli V.
      • Buratowski S.
      The Set1 N-terminal domain and Swd2 interact with RNA polymerase II CTD to recruit COMPASS.
      ). Although Set1p is the catalytic constituent of COMPASS, the other subunits, with the exception of Shg1p (
      • Mersman D.P.
      • Du H.-N.
      • Fingerman I.M.
      • South P.F.
      • Briggs S.D.
      Charge-based interaction conserved within histone H3 lysine 4 (H3K4) methyltransferase complexes is needed for protein stability, histone methylation, and gene expression.
      ), each influence the stability and activity of the methyltransferase complex in distinct ways (
      • Dehé P.-M.
      • Dichtl B.
      • Schaft D.
      • Roguev A.
      • Pamblanco M.
      • Lebrun R.
      • Rodríguez-Gil A.
      • Mkandawire M.
      • Landsberg K.
      • Shevchenko A.
      Protein interactions within the Set1 complex and their roles in the regulation of histone 3 lysine 4 methylation.
      ,
      • Dehe P.-M.
      • Geli V.
      The multiple faces of Set1.
      ,
      • Schneider J.
      • Wood A.
      • Lee J.-S.
      • Schuster R.
      • Dueker J.
      • Maguire C.
      • Swanson S.K.
      • Florens L.
      • Washburn M.P.
      • Shilatifard A.
      Molecular regulation of histone H3 trimethylation by COMPASS and the regulation of gene expression.
      ). For instance, the WD40 domain–containing subunits Swd1p and Swd3p are both required for COMPASS to catalyze all three states of H3K4 methylation, whereas Spp1p and Sdc1p are only needed for trimethylation (
      • Schneider J.
      • Wood A.
      • Lee J.-S.
      • Schuster R.
      • Dueker J.
      • Maguire C.
      • Swanson S.K.
      • Florens L.
      • Washburn M.P.
      • Shilatifard A.
      Molecular regulation of histone H3 trimethylation by COMPASS and the regulation of gene expression.
      ,
      • Takahashi Y.-H.
      • Shilatifard A.
      Structural basis for H3K4 trimethylation by yeast Set1/COMPASS.
      ). Recently, structural studies have provided insights into the molecular stoichiometry and topology of COMPASS (
      • Hsu P.L.
      • Li H.
      • Lau H.-T.
      • Leonen C.
      • Dhall A.
      • Ong S.-E.
      • Chatterjee C.
      • Zheng N.
      Crystal structure of the COMPASS H3K4 methyltransferase catalytic module.
      ,
      • Qu Q.
      • Takahashi Y.-h.
      • Yang Y.
      • Hu H.
      • Zhang Y.
      • Brunzelle J.S.
      • Couture J.-F.
      • Shilatifard A.
      • Skiniotis G.
      Structure and conformational dynamics of a COMPASS histone H3K4 methyltransferase complex.
      ). The complex is scaffolded by a core subcomplex involving two heteromeric interactions between Swd1p/Swd3p and Bre2p/Sdc1p (
      • Roguev A.
      • Schaft D.
      • Shevchenko A.
      • Pijnappel W.P.
      • Wilm M.
      • Aasland R.
      • Stewart A.F.
      The Saccharomyces cerevisiae Set1 complex includes an Ash2 homologue and methylates histone 3 lysine 4.
      ). High-resolution cryogenic electron microscopy defined the three-dimensional structure of the COMPASS core, revealing a Y-shaped configuration wherein Swd1p/Swd3p localize at the top of adjacent lobes, whereas Bre2p/Sdc1p reside at the base (
      • Takahashi Y.-h.
      • Westfield G.H.
      • Oleskie A.N.
      • Trievel R.C.
      • Shilatifard A.
      • Skiniotis G.
      Structural analysis of the core COMPASS family of histone H3K4 methylases from yeast to human.
      ). The SET domain of Set1p is located at the juncture of these subunits, thus creating a central canal that may regulate catalysis and product specificity of COMPASS. This macromolecular structure has been shown to be stabilized by an electrostatic interaction between a small basic patch within the N-SET domain of Set1p and an acidic patch toward the C-terminus of Swd1p (
      • Mersman D.P.
      • Du H.-N.
      • Fingerman I.M.
      • South P.F.
      • Briggs S.D.
      Charge-based interaction conserved within histone H3 lysine 4 (H3K4) methyltransferase complexes is needed for protein stability, histone methylation, and gene expression.
      ). Crucially, the COMPASS complex forms a dimeric macromolecule in vivo, via the Sdc1p dimer interface, allowing COMPASS to efficiently deposit methylation at both copies of histone H3 within a single nucleosome (
      • Choudhury R.
      • Singh S.
      • Arumugam S.
      • Roguev A.
      • Stewart A.F.
      The Set1 complex is dimeric and acts with Jhd2 demethylation to convey symmetrical H3K4 trimethylation.
      ). This symmetric H3K4 methylation by Set1p is the only known example of such a phenomenon in budding yeast.
      Through its methylation of H3K4, Set1p is involved in the regulation of transcriptional initiation and early elongation. Set1p plays a key role in transcriptional activation; approximately 80% of S. cerevisiae genes are downregulated upon Set1p deletion (
      • Boa S.
      • Coert C.
      • Patterton H.G.
      Saccharomyces cerevisiae Set1p is a methyltransferase specific for lysine 4 of histone H3 and is required for efficient gene expression.
      ). In certain contexts, Set1p can also function as a transcriptional repressor through its recruitment of HDAC complexes to chromatin where they antagonize nucleosome acetylation and remodeling of downstream promoters (Fig. 1) (
      • Kim T.
      • Buratowski S.
      Dimethylation of H3K4 by Set1 recruits the Set3 histone deacetylase complex to 5′ transcribed regions.
      ,
      • Pinskaya M.
      • Gourvennec S.
      • Morillon A.
      H3 lysine 4 di-and tri-methylation deposited by cryptic transcription attenuates promoter activation.
      ). Strikingly, recent studies have proposed a model whereby the combined activities of Set1p and its cognate H3K4 demethylase, Jhd2p, cooperatively regulate genome-wide chromatin structure and thus gene expression, rather than opposing one another as logic may suggest (
      • Ramakrishnan S.
      • Pokhrel S.
      • Palani S.
      • Pflueger C.
      • Parnell T.J.
      • Cairns B.R.
      • Bhaskara S.
      • Chandrasekharan M.B.
      Counteracting H3K4 methylation modulators Set1 and Jhd2 co-regulate chromatin dynamics and gene transcription.
      • Choudhury R.
      • Singh S.
      • Arumugam S.
      • Roguev A.
      • Stewart A.F.
      The Set1 complex is dimeric and acts with Jhd2 demethylation to convey symmetrical H3K4 trimethylation.
      ). The mechanistic details of this coregulation are discussed later (see Jhd2p section). Although the different degrees of H3K4 methylation are known to play distinct roles in transcriptional regulation (Fig. 2A) (
      • Kim T.
      • Buratowski S.
      Dimethylation of H3K4 by Set1 recruits the Set3 histone deacetylase complex to 5′ transcribed regions.
      ,
      • Fingerman I.M.
      • Wu C.-L.
      • Wilson B.D.
      • Briggs S.D.
      Global loss of Set1-mediated H3 Lys4 trimethylation is associated with silencing defects in Saccharomyces cerevisiae.
      ), the precise molecular cues controlling the differential production of H3K4me1, me2, and me3 by Set1p are still largely unknown. Set1p contains two conserved RRMs that have been shown to bind RNA in vitro in a methyltransferase-independent manner and may affect COMPASS distribution along nascent mRNA transcripts and subsequent H3K4 methylation (Fig. 3) (
      • Luciano P.
      • Jeon J.
      • El-Kaoutari A.
      • Challal D.
      • Bonnet A.
      • Barucco M.
      • Candelli T.
      • Jourquin F.
      • Lesage P.
      • Kim J.
      Binding to RNA regulates Set1 function.
      • Bae H.J.
      • Dubarry M.
      • Jeon J.
      • Soares L.M.
      • Dargemont C.
      • Kim J.
      • Geli V.
      • Buratowski S.
      The Set1 N-terminal domain and Swd2 interact with RNA polymerase II CTD to recruit COMPASS.
      ). Set1p has also been shown to methylate the kinetochore component, Dam1p, making it the only histone methyltransferase in budding yeast to modify a nonhistone substrate (
      • Zhang K.
      • Lin W.
      • Latham J.A.
      • Riefler G.M.
      • Schumacher J.M.
      • Chan C.
      • Tatchell K.
      • Hawke D.H.
      • Kobayashi R.
      • Dent S.Y.
      The Set1 methyltransferase opposes Ipl1 aurora kinase functions in chromosome segregation.
      ). Here, Dam1p methylation negatively regulates its subsequent phosphorylation by the Aurora kinase, Ipl1p, in order to control chromosome segregation and cell viability. It remains unclear how Set1p selects its desired substrate (histone H3 or Dam1p) for methylation, and it is not known whether the activities of Set1p in transcriptional regulation and mitosis functionally interact.
      There are two primary means by which the function of Set1p is regulated in the context of transcription. First, H3K4 methylation is controlled by H2B ubiquitination. Although H2BK123 ubiquitination is dispensable for H3K4 monomethylation, it is a requirement for COMPASS to catalyze both dimethylation and trimethylation (Fig. 2B) (
      • Shahbazian M.D.
      • Zhang K.
      • Grunstein M.
      Histone H2B ubiquitylation controls processive methylation but not monomethylation by Dot1 and Set1.
      ), suggesting that it may modulate Set1p processivity. The mechanisms underpinning this PTM cross talk have been intensely studied and rigorously debated in the literature. Initial studies proposed that Swd2p, the only essential COMPASS subunit, recognizes H2BK123-ubiquitinated chromatin and is thus required for the assembly of trimethylation-competent COMPASS (
      • Lee J.-S.
      • Shukla A.
      • Schneider J.
      • Swanson S.K.
      • Washburn M.P.
      • Florens L.
      • Bhaumik S.R.
      • Shilatifard A.
      Histone crosstalk between H2B monoubiquitination and H3 methylation mediated by COMPASS.
      ,
      • Wu M.
      • Wang P.F.
      • Lee J.S.
      • Martin-Brown S.
      • Florens L.
      • Washburn M.
      • Shilatifard A.
      Molecular regulation of H3K4 trimethylation by Wdr82, a component of human Set1/COMPASS.
      ). It was later clarified that the N-SET domain of Set1p serves as a novel sensor of H2BK123ub and that this cross talk conditionally involves Spp1p but not Swd2p (
      • Kim J.
      • Kim J.-A.
      • McGinty R.K.
      • Nguyen U.T.
      • Muir T.W.
      • Allis C.D.
      • Roeder R.G.
      The n-SET domain of Set1 regulates H2B ubiquitylation-dependent H3K4 methylation.
      ). Indeed, in vitro analysis of reconstituted COMPASS and H2Bub chromatin showed that the Spp1p PHDL domain, in conjunction with N-SET, interacts with Swd1p/Swd3p to facilitate H2Bub-dependent H3K4 methylation (
      • Jeon J.
      • McGinty R.K.
      • Muir T.W.
      • Kim J.-A.
      • Kim J.
      Crosstalk among Set1 complex subunits involved in H2B ubiquitylation-dependent H3K4 methylation.
      ). However, in light of the lack of evidence showing a direct physical interaction between Spp1p or N-SET and H2BK123-ubiquitinated chromatin, the precise mechanisms that govern this interplay remain elusive. Second, Set1p is recruited to sites of transcriptional elongation through its phosphorylation-dependent interaction with the CTD of Rbp1p, a component of the RNAPII holoenzyme. In S. cerevisiae, the CTD of Rbp1p consists of 26 repeats of an evolutionarily conserved heptapeptide, of consensus sequence YSPTSPS (
      • Corden J.L.
      Tails of RNA polymerase II.
      ). Both serine 2 and serine 5 within this repeat can be phosphorylated, and these modification isoforms show unique spatiotemporal profiles; serine 5–phosphorylated RNAPII is localized to promoter regions at initiation/early elongation stages, whereas its serine 2–phosphorylated counterpart is found throughout coding regions during elongation (
      • Komarnitsky P.
      • Cho E.-J.
      • Buratowski S.
      Different phosphorylated forms of RNA polymerase II and associated mRNA processing factors during transcription.
      ,
      • Licatalosi D.D.
      • Geiger G.
      • Minet M.
      • Schroeder S.
      • Cilli K.
      • McNeil J.B.
      • Bentley D.L.
      Functional interaction of yeast pre-mRNA 3′ end processing factors with RNA polymerase II.
      ). Set1p co-transcriptionally associates with serine 5–phosphorylated RNAPII (
      • Ng H.H.
      • Robert F.
      • Young R.A.
      • Struhl K.
      Targeted recruitment of Set1 histone methylase by elongating Pol II provides a localized mark and memory of recent transcriptional activity.
      ) to establish a gradient of H3K4 methylation that peaks near the promoter and decreases throughout a gene’s body (Fig. 2A) (
      • Kouzarides T.
      Chromatin modifications and their function.
      ). This 5′ concentration of H3K4 methylation is mediated by Kin28p, a TFIIH-associated kinase, which phosphorylates serine 5 of Rbp1p and thus triggers the transition between transcriptional initiation and elongation in response to cellular cues via COMPASS (
      • Ng H.H.
      • Robert F.
      • Young R.A.
      • Struhl K.
      Targeted recruitment of Set1 histone methylase by elongating Pol II provides a localized mark and memory of recent transcriptional activity.
      ).
      In addition to regulation by H2B ubiquitination and RNAPII phosphorylation, Set1p function is fine-tuned through a number of mechanisms. In one of the few examples of transcriptional control of genes encoding histone methylation proteins in S. cerevisiae, Gcn5p promotes the expression of SET1 and thus indirectly increases H3K4 trimethylation levels but not H3K4me2 and H3K4me1 (
      • Gong X.
      • Yu Q.
      • Duan K.
      • Tong Y.
      • Zhang X.
      • Mei Q.
      • Lu L.
      • Yu X.
      • Li S.
      Histone acetyltransferase Gcn5 regulates gene expression by promoting the transcription of histone methyltransferase SET1.
      ). At the protein level, Hmt1p-mediated asymmetric dimethylation at H3R2 negatively regulates adjacent trimethylation of H3K4 by Set1p, thus highlighting functional cross talk between arginine and lysine methylation on histone proteins (Fig. 2B). H3R2me2a spatially occludes the COMPASS subunit, Spp1p, which is essential for H3K4 trimethylation (
      • Kirmizis A.
      • Santos-Rosa H.
      • Penkett C.J.
      • Singer M.A.
      • Vermeulen M.
      • Mann M.
      • Bähler J.
      • Green R.D.
      • Kouzarides T.
      Arginine methylation at histone H3R2 controls deposition of H3K4 trimethylation.
      ). Other histone residues are known to regulate Set1p activity, namely, H3K14, which may positively cross talk with H2B ubiquitination and/or directly interact with COMPASS through electrostatic attraction to promote H3K4 methylation (
      • Nakanishi S.
      • Sanderson B.W.
      • Delventhal K.M.
      • Bradford W.D.
      • Staehling-Hampton K.
      • Shilatifard A.
      A comprehensive library of histone mutants identifies nucleosomal residues required for H3K4 methylation.
      ).

      Set2p

      SET domain-containing 2 (Set2p), also known as KMT3 or EZL1, is the sole H3K36-specific methyltransferase in the budding yeast proteome and is central to the regulation of transcriptional initiation and elongation. In 2002, Strahl et al. first purified and biochemically characterized Set2p from S. cerevisiae and demonstrated that it catalyzes the processive mono-, di-, and trimethylation of H3K36 through its catalytic SET domain in vivo (Table 1) (
      • Venkatesh S.
      • Smolle M.
      • Li H.
      • Gogol M.M.
      • Saint M.
      • Kumar S.
      • Natarajan K.
      • Workman J.L.
      Set2 methylation of histone H3 lysine 36 suppresses histone exchange on transcribed genes.
      ,
      • Strahl B.D.
      • Grant P.A.
      • Briggs S.D.
      • Sun Z.-W.
      • Bone J.R.
      • Caldwell J.A.
      • Mollah S.
      • Cook R.G.
      • Shabanowitz J.
      • Hunt D.F.
      Set2 is a nucleosomal histone H3-selective methyltransferase that mediates transcriptional repression.
      ). This domain is comprised of AWS (associated with SET; residues 63–118), SET (residues 120–237), and post-SET (residues 244–260) motifs (Fig. 3) (
      • McDaniel S.L.
      • Strahl B.D.
      Shaping the cellular landscape with Set2/SETD2 methylation.
      ). Curiously, the isolated SET domain is capable of methylating free histones, whereas full-length Set2p preferentially acts upon nucleosomal substrates (
      • Wang Y.
      • Niu Y.
      • Li B.
      Balancing acts of SRI and an auto-inhibitory domain specify Set2 function at transcribed chromatin.
      ), suggesting that sequences distal to the methyltransferase domain regulate substrate specificity. Indeed, an N-terminal acidic patch (residues 31–39) and a C-terminal Set2-Rbp1 interacting (SRI; residues 619–718, PDB ID: 2C5Z (
      • Vojnic E.
      • Simon B.
      • Strahl B.D.
      • Sattler M.
      • Cramer P.
      Structure and carboxyl-terminal domain (CTD) binding of the Set2 SRI domain that couples histone H3 Lys36 methylation to transcription.
      )) domain control the association of Set2p with histone H4 (
      • Du H.-N.
      • Fingerman I.M.
      • Briggs S.D.
      Histone H3 K36 methylation is mediated by a trans-histone methylation pathway involving an interaction between Set2 and histone H4.
      ,
      • Du H.-N.
      • Briggs S.D.
      A nucleosome surface formed by histone H4, H2A, and H3 residues is needed for proper histone H3 Lys36 methylation, histone acetylation, and repression of cryptic transcription.
      ) and RNAPII (
      • Kizer K.O.
      • Phatnani H.P.
      • Shibata Y.
      • Hall H.
      • Greenleaf A.L.
      • Strahl B.D.
      A novel domain in Set2 mediates RNA polymerase II interaction and couples histone H3 K36 methylation with transcript elongation.
      ), respectively. The tryptophan-tryptophan (WW) domain (residues 475–507, PDB ID: 1E0N (
      • Macias M.J.
      • Gervais V.
      • Civera C.
      • Oschkinat H.
      Structural analysis of WW domains and design of a WW prototype.
      )) is currently of no known function, although its deletion does not modulate H3K36 methylation levels or Set2p RNAPII binding (
      • Xiao T.
      • Hall H.
      • Kizer K.O.
      • Shibata Y.
      • Hall M.C.
      • Borchers C.H.
      • Strahl B.D.
      Phosphorylation of RNA polymerase II CTD regulates H3 methylation in yeast.
      ). It has been speculated that it may control nonhistone methylation given that the WW domain of SETD2, the mammalian ortholog of Set2p, mediates its interaction with the Huntingtin protein (
      • McDaniel S.L.
      • Strahl B.D.
      Shaping the cellular landscape with Set2/SETD2 methylation.
      ,
      • Faber P.W.
      • Barnes G.T.
      • Srinidhi J.
      • Chen J.
      • Gusella J.F.
      • MacDonald M.E.
      Huntingtin interacts with a family of WW domain proteins.
      ,
      • Gao Y.-G.
      • Yang H.
      • Zhao J.
      • Jiang Y.-J.
      • Hu H.-Y.
      Autoinhibitory structure of the WW domain of HYPB/SETD2 regulates its interaction with the proline-rich region of huntingtin.
      ). Yeast Set2p, however, has no known nonhistone substrates identified to date. Finally, an AID (residues 262–476) attenuates Set2p-mediated H3K36 trimethylation by antagonizing its catalytic activity and fine-tuning several functions of SRI (Fig. 3) (
      • Wang Y.
      • Niu Y.
      • Li B.
      Balancing acts of SRI and an auto-inhibitory domain specify Set2 function at transcribed chromatin.
      ).
      In the context of transcription, Set2p recruits several key complexes that cooperate to re-establish a compact chromatin landscape in the wake of elongating RNAPII (
      • Jha D.K.
      • Pfister S.X.
      • Humphrey T.C.
      • Strahl B.D.
      SET-ting the stage for DNA repair.
      ). Set2p co-transcriptionally modifies H3K36, which is in turn recognized by the chromodomain of Eaf3p, a subunit of the Rpd3S HDAC complex (Fig. 1) (
      • Fuchs S.M.
      • Kizer K.O.
      • Braberg H.
      • Krogan N.J.
      • Strahl B.D.
      RNA polymerase II carboxyl-terminal domain phosphorylation regulates protein stability of the Set2 methyltransferase and histone H3 di-and trimethylation at lysine 36.
      ). Rpd3S functions to keep gene bodies deacetylated and thus restores chromatin structure between multiple rounds of transcription (
      • Fuchs S.M.
      • Kizer K.O.
      • Braberg H.
      • Krogan N.J.
      • Strahl B.D.
      RNA polymerase II carboxyl-terminal domain phosphorylation regulates protein stability of the Set2 methyltransferase and histone H3 di-and trimethylation at lysine 36.
      ,
      • Li B.
      • Gogol M.
      • Carey M.
      • Pattenden S.G.
      • Seidel C.
      • Workman J.L.
      Infrequently transcribed long genes depend on the Set2/Rpd3S pathway for accurate transcription.
      ,
      • Lickwar C.R.
      • Rao B.
      • Shabalin A.A.
      • Nobel A.B.
      • Strahl B.D.
      • Lieb J.D.
      The Set2/Rpd3S pathway suppresses cryptic transcription without regard to gene length or transcription frequency.
      ). This epigenetic resetting protects genes from inappropriate and bidirectional transcription, from cryptic initiation sites within open reading frames (
      • Carrozza M.J.
      • Li B.
      • Florens L.
      • Suganuma T.
      • Swanson S.K.
      • Lee K.K.
      • Shia W.-J.
      • Anderson S.
      • Yates J.
      • Washburn M.P.
      Histone H3 methylation by Set2 directs deacetylation of coding regions by Rpd3S to suppress spurious intragenic transcription.
      ,
      • Lickwar C.R.
      • Rao B.
      • Shabalin A.A.
      • Nobel A.B.
      • Strahl B.D.
      • Lieb J.D.
      The Set2/Rpd3S pathway suppresses cryptic transcription without regard to gene length or transcription frequency.
      ,
      • Hacker K.E.
      • Fahey C.C.
      • Shinsky S.A.
      • Chiang Y.-C.J.
      • DiFiore J.V.
      • Jha D.K.
      • Vo A.H.
      • Shavit J.A.
      • Davis I.J.
      • Strahl B.D.
      Structure/function analysis of recurrent mutations in SETD2 protein reveals a critical and conserved role for a SET domain residue in maintaining protein stability and histone H3 Lys-36 trimethylation.
      ). Prevention of intragenic transcription by H3K36 methylation is conserved in human as deletion of mammalian SETD2 causes upregulation of spurious mRNA transcripts (
      • Carvalho S.
      • Raposo A.C.
      • Martins F.B.
      • Grosso A.R.
      • Sridhara S.C.
      • Rino J.
      • Carmo-Fonseca M.
      • de Almeida S.F.
      Histone methyltransferase SETD2 coordinates FACT recruitment with nucleosome dynamics during transcription.
      ,
      • Gopalakrishnan R.
      • Marr S.K.
      • Kingston R.E.
      • Winston F.
      A conserved genetic interaction between Spt6 and Set2 regulates H3K36 methylation.
      ), a phenotype shared by set2Δ yeast cells (
      • Li B.
      • Gogol M.
      • Carey M.
      • Pattenden S.G.
      • Seidel C.
      • Workman J.L.
      Infrequently transcribed long genes depend on the Set2/Rpd3S pathway for accurate transcription.
      ,
      • Lickwar C.R.
      • Rao B.
      • Shabalin A.A.
      • Nobel A.B.
      • Strahl B.D.
      • Lieb J.D.
      The Set2/Rpd3S pathway suppresses cryptic transcription without regard to gene length or transcription frequency.
      ). This repressive transcriptional environment is reinforced by the recruitment of Isw1b, a chromatin remodeling complex that binds H3K36me and reorganizes nucleosomes to allow Rpd3S-mediated deacetylation of neighboring nucleosomes (Fig. 1) (