Neuronal ER–plasma membrane junctions organized by Kv2–VAP pairing recruit Nir proteins and affect phosphoinositide homeostasisThe association of plasma membrane (PM)-localized voltage-gated potassium (Kv2) channels with endoplasmic reticulum (ER)-localized vesicle-associated membrane protein-associated proteins VAPA and VAPB defines ER–PM junctions in mammalian brain neurons. Here, we used proteomics to identify proteins associated with Kv2/VAP-containing ER–PM junctions. We found that the VAP-interacting membrane-associated phosphatidylinositol (PtdIns) transfer proteins PYK2 N-terminal domain-interacting receptor 2 (Nir2) and Nir3 specifically associate with Kv2.1 complexes.
A catalytic domain variant of mitofusin requiring a wildtype paralog for function uncouples mitochondrial outer-membrane tethering and fusionMitofusins (Mfns) are dynamin-related GTPases that mediate mitochondrial outer-membrane fusion, a process that is required for mitochondrial and cellular health. In Mfn1 and Mfn2 paralogs, a conserved phenylalanine (Phe-202 (Mfn1) and Phe-223 (Mfn2)) located in the GTPase domain on a conserved β strand is part of an aromatic network in the core of this domain. To gain insight into the poorly understood mechanism of Mfn-mediated membrane fusion, here we characterize a Mitofusin mutant variant etiologically linked to Charcot–Marie–Tooth syndrome.
The intramembrane protease SPP impacts morphology of the endoplasmic reticulum by triggering degradation of morphogenic proteinsThe endoplasmic reticulum (ER), as a multifunctional organelle, plays crucial roles in lipid biosynthesis and calcium homeostasis as well as the synthesis and folding of secretory and membrane proteins. Therefore, it is of high importance to maintain ER homeostasis and to adapt ER function and morphology to cellular needs. Here, we show that signal peptide peptidase (SPP) modulates the ER shape through degradation of morphogenic proteins. Elevating SPP activity induces rapid rearrangement of the ER and formation of dynamic ER clusters.
Phospholipid subcellular localization and dynamicsMembrane biology seeks to understand how lipids and proteins within bilayers assemble into large structures such as organelles and the plasma membranes. Historically, lipids were thought to merely provide structural support for bilayer formation and membrane protein function. Research has now revealed that phospholipid metabolism regulates nearly all cellular processes. Sophisticated techniques helped identify >10,000 lipid species suggesting that lipids support many biological processes. Here, we highlight the synthesis of the most abundant glycerophospholipid classes and their distribution in organelles.
Purification and functional characterization of the vacuolar malate transporter tDT from ArabidopsisThe exact transport characteristics of the vacuolar dicarboxylate transporter tDT from Arabidopsis are elusive. To overcome this limitation, we combined a range of experimental approaches comprising generation/analysis of tDT overexpressors, 13CO2 feeding and quantification of 13C enrichment, functional characterization of tDT in proteoliposomes, and electrophysiological studies on vacuoles. tdt knockout plants showed decreased malate and increased citrate concentrations in leaves during the diurnal light-dark rhythm and after onset of drought, when compared with wildtypes.
Functional regions of the peroxin Pex19 necessary for peroxisome biogenesisThe peroxins Pex19 and Pex3 play an indispensable role in peroxisomal membrane protein (PMP) biogenesis, peroxisome division, and inheritance. Pex19 plays multiple roles in these processes, but how these functions relate to the structural organization of the Pex19 domains is unresolved. To this end, using deletion mutants, we mapped the Pex19 regions required for peroxisome biogenesis in the yeast Pichia pastoris. Surprisingly, import-competent peroxisomes still formed when Pex19 domains previously believed to be required for biogenesis were deleted, although the peroxisome size was larger than that in wild-type cells.
RUTBC1 Functions as a GTPase-activating Protein for Rab32/38 and Regulates Melanogenic Enzyme Trafficking in MelanocytesTwo cell type-specific Rab proteins, Rab32 and Rab38 (Rab32/38), have been proposed as regulating the trafficking of melanogenic enzymes, including tyrosinase and tyrosinase-related protein 1 (Tyrp1), to melanosomes in melanocytes. Like other GTPases, Rab32/38 function as switch molecules that cycle between a GDP-bound inactive form and a GTP-bound active form; the cycle is thought to be regulated by an activating enzyme, guanine nucleotide exchange factor (GEF), and an inactivating enzyme, GTPase-activating protein (GAP), which stimulates the GTPase activity of Rab32/38.
Role of Pex21p for Piggyback Import of Gpd1p and Pnc1p into Peroxisomes of Saccharomyces cerevisiaeBackground: PTS2 proteins Gpd1p and Pnc1p are imported into peroxisomes in a PTS2 receptor-dependent manner.Results: PTS2co-receptor Pex21p is required for peroxisomal piggyback import of Gpd1p and Pnc1p.Conclusion: PTS2 co-receptors Pex18p and Pex21p enable targeting of distinct cargo proteins under variable stress conditions.Significance: The life span-regulating Pnc1p is transported into peroxisomes via a novel import route.
Small GTPase Rab2B and Its Specific Binding Protein Golgi-associated Rab2B Interactor-like 4 (GARI-L4) Regulate Golgi MorphologyBackground: Rab small GTPases are membrane trafficking proteins in eukaryotes.Results: Comprehensive knockdown screening identified six Rab isoforms that are involved in regulating Golgi morphology in HeLa-S3 cells.Conclusion: Five of the six Rabs, including Rab2A and Rab2B, non-redundantly regulate Golgi morphology. A Rab2B-specific effector, GARI-L4, also regulates it.Significance: This is the first study to systematically analyze all human Rabs in the Golgi.
Mechanistic Aspects of Folded Protein Transport by the Twin Arginine Translocase (Tat)The twin arginine translocase (Tat) transports folded proteins of widely varying size across ionically tight membranes with only 2–3 components of machinery and the proton motive force. Tat operates by a cycle in which the receptor complex combines with the pore-forming component to assemble a new translocase for each substrate. Recent data on component and substrate organization in the receptor complex and on the structure of the pore complex inform models for translocase assembly and translocation.
Cox17 Protein Is an Auxiliary Factor Involved in the Control of the Mitochondrial Contact Site and Cristae Organizing SystemBackground: MICOS is a recently identified complex that is important for maintaining the architecture of the mitochondrial inner membrane.Results: Cox17 interacts with the MICOS complex, and this interaction is promoted by copper ions.Conclusion: Cox17 regulates MICOS integrity.Significance: Learning the way in which MICOS is regulated provides important insights into processes that are responsible for mitochondrial form and function.