|
|
|
|||||||
|
|||||||||
J Biol Chem, Vol. 275, Issue 13, 9773-9781, March 31, 2000
From the The protein product of the Toll-like receptor
(TLR) 4 gene has been implicated in the signal transduction events
induced by lipopolysaccharide (LPS). In mice, destructive mutations of
Tlr4 impede the normal response to LPS and cause a high
susceptibility to Gram-negative infection. Expression of TLR4 mRNA
in humans is restricted to a small number of cell types, including
LPS-responsive myeloid cells, B-cells, and endothelial cells. To
investigate the molecular basis for TLR4 expression in cells of myeloid
origin, we cloned the human TLR4 gene and analyzed its putative
5'-proximal promoter. In transient transfections a region of only 75 base pairs upstream of the major transcription initiation site was sufficient to induce maximal luciferase activity in THP-1 cells. The
sequence of this region is similar in human and mouse TLR4 genes and
lacks a TATA box, typical Sp1-sites or CCAAT box sequences. Instead, it
contains consensus-binding sites for Ets family transcription factors,
octamer-binding factors, and a composite interferon response factor/Ets
motif. The activity of the promoter in macrophages was strictly
dependent on the integrity of both half sites of the composite
interferon response factor/Ets motif, which was constitutively bound by
the myeloid and B-cell-specific transcription factor PU.1 and
interferon consensus sequence-binding protein. These results indicate
that the two tissue-restricted transcription factors PU.1 and
interferon consensus sequence-binding protein participate in the basal
regulation of human TLR4 in myeloid cells. Cloning of the human TLR4
gene provides a basis for further investigation of the possible impact
of genetic variations on the susceptibility to infection and sepsis.
Recognition of bacterial lipopolysaccharide
(LPS),1 the abundant
glycolipid of the outer membrane of Gram-negative bacteria, is an
important function of innate immune cells. Detection of LPS is
exquisitely sensitive. Minute quantities can activate mammalian immune
cells (in particular, monocytes and tissue macrophages) that leads to
rapid secretion of cytokines (e.g. tumor necrosis factor,
IL-1, IL-6, and IL-10) and enhanced microbicidal capacities. Failure to
contain Gram-negative infection and to control pro-inflammatory cytokine secretion can result in septic shock and death (1-3).
The interaction of LPS with monocytes and neutrophils is known to occur
via CD14 (4) and is strongly enhanced by LPS-binding protein (5, 6).
Multiple signal transduction pathways are activated in macrophages upon
LPS stimulation, including the activation of transcription factor
NF- The prototypic Toll protein of Drosophila controls
dorsal-ventral patterning in the embryo and is required for anti-fungal responses in adult flies (8). A plasma membrane receptor, Toll has a
leucine-rich ectodomain (9, 10) and a cytoplasmic domain with sequence
homology to several mammalian proteins, including both chains of the
IL-1 receptor (11), the IL-18 receptor (12), SIGRR (13), MyD88 (14),
and all members of the newly described Toll-like receptor (TLR) family
(15-17). At least two of these proteins (IL-1R and TLR4) are known to
signal via activation of MyD88, IRAK, TRAF6, and NF- Positional cloning analysis recently revealed that mice of the C3H/HeJ
and C57BL/10ScCr strains, which are profoundly unresponsive to LPS (23,
24), have separate genetic defects of Tlr4 (25). This
finding suggests that TLR4 is, in fact, the transducing subunit of the
LPS receptor. Direct measurements of TLR4 activity in macrophages sustain the conclusion, in that modest overexpression of a mutant form
of Tlr4, corresponding to that encoded by the codominant Tlr4Lps-d allele present in C3H/HeJ mice, causes a dramatic
increase in the LPS EC50 for tumor necrosis factor
production. Overexpression of the native protein depresses the LPS
EC50.2
Interestingly, TLR4 appears to exist in association with MD-2, an
exteriorized protein expressed by monocytic cells (27). It may be that
co-expression is required for the LPS transducing activity of TLR4.
Historically, a number of mysteries have beset the field of LPS signal
transduction. Among these, there has been no molecular explanation for
the large interspecific differences in LPS response (28), suppression
of LPS response by corticosteroids (29), augmentation of the response
by various bacterial infections (30-33) or by interferons (34-39),
and tolerance to LPS elicited by pretreatment with LPS itself (40).
TLR4 has thus become a focus of inquiry aimed at understanding each of
these phenomena.
The transcriptional regulation of TLR4 is a starting point for such an
inquiry. Expression of human TLR4 is restricted to a small number of
cell types, including endothelial cells, B-cells, and predominantly
myeloid cells (monocytes, macrophages, dendritic cells, and
granulocytes) (15, 16, 21, 41). Both the basal level of TLR4 expression
and its regulation in myeloid cells may influence responses to LPS and,
hence, Gram-negative infection. Accordingly, we have sought to define
the TLR4 promoter and to analyze those factors that govern TLR4 gene
expression in human macrophages, which are the main LPS-responsive cell
type. This analysis has entailed determination of the complete sequence
of TLR4 and flanking genomic DNA.
Chemicals--
All chemical reagents used were purchased from
Sigma-Aldrich unless otherwise noted. Protease inhibitors are from
Roche Molecular Biochemicals. Oligonucleotides were synthesized by TIB
Molbiol (Berlin, Germany). Antisera for supershift analyses were
purchased from Santa Cruz.
Cells--
Peripheral blood mononuclear cells were separated by
leukapheresis of healthy donors, followed by density gradient
centrifugation over Ficoll/Hypaque. Monocytes were isolated from
mononuclear cells by countercurrent centrifugal elutriation in a J6M-E
centrifuge (Beckman, München, Germany) as described previously
(42). Monocytes were >90% pure as determined by morphology and
expression of CD14 antigen. Isolated monocytes were cultured in RPMI
1640 medium (Biochrom KG, Berlin, Germany) supplemented with vitamins,
antibiotics, pyruvate, nonessential amino acids (all from Life
Technologies, Inc.), 5 × 10 Gene Cloning--
The human TLR4 gene was identified by
hybridization screening in BAC 110p15 (Genome Systems human BAC
library). The mouse Tlr4 gene, as previously reported (43),
resides in BAC 152c16 (Research Genetics mouse BAC library, 129 strain). Both BACs were fragmented by ultrasonic disruption, repaired
using the Klenow fragment of DNA polymerase I, and shotgun cloned into
the vector pBluescript(KS). Bidirectional sequencing was carried out on
clones picked at random from the library. To increase the density of reads over the gene itself, both BACs were amplified using long range
PCR to yield a 12-kb fragment containing all of the human exons and a
16-kb fragment containing all of the mouse exons. These fragments were
also shotgun cloned and sequenced.
Assembly was accomplished using the programs phred and Phrap, and
contiguous sequences were visualized using the program
consed_alpha. All of these programs, produced by the University of
Washington Genome Center, were run under a UNIX operating system, on a
DEC-alpha computer.
A 4.3-kb genomic fragment of the hTLR promoter was amplified from the
BAC clone containing the human TLR4 gene using the Expand High Fidelity
PCR system (Roche Biochemicals) and the primers 5'-CTC CAT GGC ACA TTC
TGC AGT AAA CTT GGA GGC-3' (sense) and 5'-CAC GCA GGA GAG GAA GGC CAT
GGC TG-3' (antisense).
RNA Preparation and RT-PCR Analysis--
Total RNA was isolated
from different cell types by the guanidine thiocyanate/acid phenol
method (44). 2.0 µg of total RNA from either cell type was reverse
transcribed using oligo(dT) primer and Superscript II (Life
Technologies, Inc.). Primer positions and sizes for the amplified
fragments of TLR4 and Primer Extension Analysis--
Poly(A) RNA was isolated from
THP-1 cells using poly(A)Tract System 1000 (Promega). Determination of
the transcription initiation site of human TLR4 was performed as
described by Calzone et al. (45). Briefly, 1 and 4 µg of
poly(A) RNA were annealed to the 32P end-labeled
oligonucleotide 5'-GGT GTC TTC TCT TCC TCG AGC-3' at 58 °C for 20 min and then cooled for 10 min at room temperature. The reverse
transcription was performed with AMV RT (Promega) at 41 °C for 30 min and then stopped by addition of 8 µl of loading buffer. Extended
products were separated on a 6% polyacrylamide/8 M urea
sequencing gel along with sequencing products of the corresponding region of the TLR4 gene obtained with the above oligonucleotide
Plasmid Construction and Purification--
PCR fragments were
inserted into the plasmid vector pCR2.1-TOPO (TOPO Cloning Kit,
Invitrogen) for sequencing and subcloning purposes. The 4.3-kb genomic
PCR fragment of the hTLR4 proximal promoter was subcloned into the
NcoI restriction site of pGL3-B (Promega) and sequenced.
Deletions of this constructs were generated by digestion with either
KpnI, ApaI, HindIII, EcoRI,
PstI, or XhoI and subsequent religation of the
remaining plasmid. Mutations of the PU.1, OCT, and IRF-binding sites
were done by PCR-mediated mutagenesis. PCR fragments with correctly
introduced mutations were subcloned back into the TLR4 DNA Sequence Analysis--
The cDNA sequencing was done by
Dye Deoxy Terminator Cycle Sequencing (Applied Biosystem) according to
the manufacturer's instructions, and sequences were analyzed on the
Applied Biosystems DNA Sequencing System (model 373A).
Transient DNA Transfections--
THP-1 cells were transfected
using DEAE-dextran. 5 × 105 THP-1 cells/ml were
seeded into tissue culture flasks the day before transfection. On the
next day, 6 ml of cell suspension were washed twice with STBS (46) and
pelleted. 0.2 µg of reporter plasmid and 0.02 µg of
Renilla luciferase control vector were mixed with DEAE-dextran (400 µg/ml) in 140 µl of STBS buffer and immediately added to the pelleted THP-1 cells. The cells were incubated at 37 °C
for 20 min, washed twice with STBS, resuspended, and cultured in
complete RPMI medium. The two cell lines HeLa and Mel Im were transfected using LipofectAMINE reagent (Life Technologies, Inc.) according to the manufacturers instructions. Briefly, 4 × 105 HeLa cells were transfected using 7 µl of
LipofectAMINE and 1 µg of total DNA (including 0.05 µg of
Renilla luciferase control vector) and incubated for 5 h before the addition of complete medium. 2.7 × 105
Mel Im cells were transfected using 5 µl of LipofectAMINE and 1 µg
of total DNA (including 0.05 µg of Renilla control vector) and incubated for 6 h before the addition of complete medium. The
transfection mixture was removed after 24 h and substituted with
complete medium. The transfected cell lines were cultivated for 48 h and harvested, and cell lysates were assayed for firefly and
Renilla luciferase activity using the Dual Luciferase
Reporter Assay System (Promega) on a Lumat LB9501 (Berthold). Firefly
luciferase activity of individual transfections was normalized against
Renilla luciferase activity.
Nuclear Extracts and Electrophoretic Mobility Shift
Assay--
Nuclear extracts were prepared with a variation of the
method of Osborn et al. (47). All buffers used contained 1 mM Na3VO4 and a mixture of protease
inhibitors (2 µg/ml aprotinin, 0.5 µg/ml leupeptin, 1 µg/ml
pepstatin, 20 µg/ml bestatin, 5 µg/ml E46, 50 µg/ml antipain, 100 µg/ml chymostatin). Oligonucleotides were end-labeled with
[ Cloning and Characterization of the Human TLR4 Gene--
A BAC
containing the human TLR4 gene was isolated by hybridization screening,
shotgun cloned and sequenced as described under "Experimental
Procedures." The mouse Tlr4 gene was similarly sequenced from a previously described BAC (43). A sequence 19 kb in length containing the human gene and a sequence 91.7 kb in length containing the mouse gene have been deposited in GenBankTM (accession
numbers AF177765 and AF177767, respectively). These sequences are
considered to be 99.99% accurate.
Comparison of the two published cDNA sequences (accession numbers
NM_003266 and U93091) for human TLR4 revealed the presence of an
additional 120-bp region in the sequence reported by Rock et
al. (16). A corresponding additional exon, which is not present in
the mouse gene, was identified in the human TLR4 genomic sequence (Fig.
1). The presence of this exon in TLR4
transcripts introduces an in-frame stop codon, which would
theoretically terminate TLR4 translation after 34 amino acids. To
clarify whether this transcript is present in macrophages, we performed
RT-PCR with specific primers flanking the region between exons I-IV of
the human TLR4 gene. As shown in Fig. 1B, three products
were amplified from macrophage and THP-1 cDNA that represent three
different splicing variants of human TLR4. Sequencing of the purified
fragments confirmed the specificity and nature of the three splicing
forms, which is indicated in Fig. 1B. As outlined in Fig.
1C, only the transcript containing exon I, III, and IV
yields the proper TLR4 protein including a putative signal peptide. The
amino acid sequence as proposed by Rock et al. (16) is
proceeded by an in-frame stop codon and does not contain a typical
leader sequence (Fig. 1C). It is therefore unlikely that the
combination of exons I, II, III, and IV would yield a functional TLR
protein.
To determine the transcriptional start site of the TLR4 gene we
performed primer extension analysis. Two specific extension products
were obtained from THP-1 RNA (Fig. 2).
The major start site resides 190 bp upstream of the adenine residue of
the first start codon. A second, minor initiation site was detected 74 bp upstream of the major start site.
Myeloid-specific Activity of the Proximal Human TLR4
Promoter--
Published tissue Northern analyses suggest a predominant
expression of hTLR4 in spleen and peripheral blood leukocytes (15, 16).
RT-PCR with freshly separated and in vitro cultivated human blood cells indicated that human TLR4 is mainly expressed in myeloid cells, including monocytes, in vitro differentiated
macrophages, monocyte-derived dendritic cells, granulocytes, and, to a
much lesser extent, mixed lymphocytes (data not shown).
To further analyze the myeloid expression of TLR4, we cloned fragments
of the 5'-proximal promoter region of the human TLR4 gene, ranging from
4.3 kb to 100 bp upstream of the ATG start codon, into a luciferase
reporter plasmid (Fig. 3). Transient transfection analysis was performed in the monocytic cell line THP-1
and two nonmyeloid cell lines, Mel Im (melanoma) and HeLa (cervical
carcinoma). Luciferase activities were normalized for transfection
efficiency by co-transfection with a Renilla luciferase construct, and results for individual cell lines were compared relative
to the activity of the promoter-less pGL3-basic construct. As shown in
Fig. 3, the presence of both positive and negative elements can be
observed. A negative regulatory region seems to reside between
nucleotides The Proximal Promoter Regions of Human and Mouse TLR4 Are Similar
and Contain Several Purine-rich Motifs--
Sequence comparison of the
proximal promoter regions of both mouse and human TLR4 revealed a high
degree of conservation. The 5' proximal regions of both TLR4 genes are
characterized by the absence of TATA boxes, consensus initiator
sequences, or GC-rich regions found in "housekeeping" genes that
normally determine transcriptional initiation (Fig.
4). They also lack Sp1 or CCAAT box
sequences and instead contain several purine-rich elements with a
5'-GGAA-3' core on either strand, a characteristic feature of many
myeloid-specific genes (49-56). The smallest promoter fragment directing maximal myeloid activity (hTLR4 PU.1 Binds to Both Elements within the Proximal hTLR4 Identification of the PU.1 Interacting Factor at the Inner
Purine-rich Motif--
In gel shift assays nuclear extracts of various
cell types, the slower migrating, PU.1 containing complex was only
observed in THP.1 cells and human macrophages. In contrast to THP-1
cells, PU.1 was detectable as both slower and faster migrating band in human macrophages (Fig. 6A).
To further investigate the nature of the slower migrating, PU.1
containing complex, we prepared THP-1 nuclear extracts either without
protease inhibitors or without the phosphatase inhibitor
Na3VO4 and performed EMSA with the PU-12 oligonucleotide. Extract preparation without protease inhibitors yielded a decrease in complex formation. Without the phosphatase inhibitor Na3VO4, the slower migrating complex
disintegrated and a faster migrating complex containing PU.1 alone
appeared, suggesting that phosphorylation of either factor or both is
necessary for complex formation.
Further sequence analysis revealed a possible binding site for members
of the family of IRF located next to the inner purine-rich motif PU-12.
Interactions between PU.1 and members of the IRF family have been
observed before (57-60). Fig.
7A compares the putative
PU.1/IRF-binding sites of the human and mouse TLR4 promoter with so far
identified sequences of similar binding sites in the human
gp91phox (58), CD20 (59), murine IL-18 promoters (60), and
immunoglobulin light chain enhancers E A Conserved Element Adjacent to the Composite ICSBP·PU.1 Motif Is
a Weak Binding Site for Octamer Transcription Factors--
A highly
conserved octamer element is present in both human and mouse TLR4
promoters, which prompted us to investigate its interaction with POU
domain transcription factors. Direct binding of nuclear proteins to the
site was almost undetectable in THP-1 cells and macrophages (data not
shown); however, competition studies with macrophage nuclear proteins
and the consensus octamer motif (OCT consensus) identified the TLR4
octamer element as a weak binding site for Oct-1 and Oct-2 (Fig.
8). Cold oligonucleotide with a 3-bp
mutation of the putative binding site (OCT-M) did not affect the
binding of octamer factors to the consensus octamer sequence.
Mutational Analysis of the Identified Binding Sites and Effect of
IFN-
To investigate whether the activity of the TLR4 promoter could be
induced in a nonmyeloid cell line that does not express either PU.1 or
ICSBP, we co-transfected HeLa (cervical carcinoma) cells with the hTLR4
IFN- In this study we investigated the transcriptional regulation of
TLR4 in human macrophages, the main LPS-responsive cell type. We cloned
and sequenced 19 kb of the human TLR4 gene, sequenced 91.7 kb of the
mouse Tlr4 gene, determined the transcriptional start sites, and
identified three alternative splicing forms of human TLR4 in human
myeloid cells. Furthermore, we performed an initial characterization of
regulatory elements controlling the expression of human TLR4 and
defined a minimal proximal promoter that confers full reporter activity
in human monocytic THP-1 cells.
Gene cloning and sequence comparison revealed the existence of an
additional exon (exon II) in the human TLR4 gene, which is not present
in the mouse gene. Its integration into TLR4 mRNA introduces an
in-frame stop codon, which prompted us to investigate the presence and
ration of alternative splicing forms of TLR4. By RT-PCR, we were able
to detect three alternative splicing variants of hTLR4; however, only
one of them encodes the proper TLR4 protein. Although we did not
observe major changes in the relative abundance of the three
transcripts in myeloid cells, alternative splicing could represent an
additional level of TLR4 regulation in humans. Disruptions of
alternative splicing have been correlated with some human genetic
diseases (67); mutations that affect the splicing of TLR4 mRNA
could favor the expression of noncoding transcripts. It is also
possible that alternative splicing events are regulated in response to
developmental or physiological cues (67). As a consequence, the
expression of TLR4 protein could, for example, be switched off, if the
second exon is included in all transcripts. However, further
investigations will be needed to clarify these issues.
Expression of human TLR4 transcripts has primarily been detected in
myeloid cells (monocytes, macrophages, and dendritic cells) and some
B-cell lines (15, 16, 27). In accordance with the observed cell type
restricted expression, the TLR4 gene is controlled by a typical myeloid
type promoter in macrophages. The 5'-proximal region lacks a TATA box,
consensus initiator sequences, or GC-rich regions found in
"housekeeping" genes and instead contains multiple purine-rich
sequence motifs that are recognized by transcription factors of the Ets
family, including PU.1. The transcription factor PU.1 was shown to be
required for the optimal expression of a growing number of
myeloid-specific genes (49-56, 68-70). Our data suggest that PU.1
also plays an important role in the myeloid expression of TLR4. In
human macrophages, PU.1 binds an essential motif in the proximal
promoter and recruits ICSBP to an adjacent binding site that is also
required for the optimal activity of the promoter.
ICSBP is a member of the IRF family of transcription factors and is
expressed mainly in cells of hematopoietic origin (71). Its mouse
homologue was originally cloned as interferon- We identified a similar site in the human TLR4 promoter and showed that
both PU.1 and ICSBP bind to adjacent elements that are indispensable
for the full activity of the TLR4 promoter. As expected, formation of
the ternary complex is dependent on the presence of PU.1 and its state
of phosphorylation. These data suggest that the interaction between the
two tissue-specific transcription factors PU.1 and ICSBP is important
for the basal activity of the TLR4 promoter in human myeloid cells.
Preliminary gel shift analyses using B-cell extracts indicate the
presence of a similar complex.3 The composite
IRF/Ets motif may therefore also contribute to the expression of TLR4
in human B-cells. Using in vitro translated proteins, we
observed that the complex between PU.1 and ICSBP migrates slightly
faster than the native THP-1 complex. This difference could be due to
altered post-translational modifications or an effect of the
(5-10-fold) higher total protein concentrations in the preparations of
in vitro translated proteins. However, it is also possible
that the native THP-1 complex contains another, as yet unidentified protein.
We identified constitutive nuclear ICSBP protein in the human monocytic
cell line THP-1 and in vitro differentiated human macrophages, which is consistent with the previously described constitutive expression of ICSBP in the human premonocytic cell line
U937. This indicates a more constitutive expression in human macrophages rather than a strictly inducible expression pattern, which
is described for mouse macrophages, where constitutive expression of
ICSBP is undetectable, and both mRNA and protein levels are strongly induced after stimulation with IFN- ICSBP-deficient mice show altered antiviral and antibacterial responses
and develop a chronic myelogenous leukemia-like syndrome (76). An
observed failure to develop Th-1-driven immune responses has been
correlated with a defect in IL-12 p40 production by cells of myeloid
origin (26, 77). The response of ICSBP-deficient mice to LPS is
described as normal (77). However, it is not clear whether the murine
Tlr4 gene is similarly dependent on the IRF/Ets motif, which
seems to form a less stable complex with ICSBP and PU.1. Analysis of
the mouse promoter will clarify whether ICSBP plays a role in the
transcriptional regulation of Tlr4 in mice.
In conclusion, our observations suggest that a functional cooperation
between PU.1 and ICSBP regulates the myeloid expression of TLR4 in
humans. Additional elements and mechanisms are likely to contribute to
the regulation of TLR4 in health and disease and are the subject to
further studies. Transcriptional regulation, alternative splicing
events, or function of TLR4 protein could be altered because of
mutations or polymorphisms of the TLR4 gene. This study provides the
basis for further investigating a possible correlation of yet unknown
genetic variations with a higher susceptibility for infection and
septic complications.
We are grateful to B.-Z. Levi and R. Maki for
providing ICSBP and PU.1 expression vectors and to E. K. L. Chan, who participated in the sequencing of the mouse Tlr4
locus. We thank D. Hume for suggestions and members of our laboratories
for helpful discussions.
*
This work was supported by a grant from the University of
Regensburg (to M. R.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
§
To whom correspondence may be addressed: Dept. of Hematology and
Oncology, University of Regensburg, 93042 Regensburg, Germany. E-mail:
Michael.Rehli@klinik.uni-regensburg.de.
**
To whom correspondence may be addressed: Howard Hughes Medical
Inst., UT Southwestern Medical Center, Dallas, TX 75235-9050. E-mail:
beutler@howie.swmed.edu.
2
X. Du, A. Poltorak, M. Silva, and B. Beutler,
submitted for publication.
3
M. Rehli, unpublished observation.
The abbreviations used are:
LPS, lipopolysaccharide;
EMSA, electrophoretic mobility shift assay;
ICSBP, interferon consensus sequence-binding protein;
IRF, interferon response
factor;
PIP, PU.1 interaction partner;
TLR, toll-like receptor;
IL, interleukin;
bp, base pair(s);
IFN, interferon;
PCR, polymerase chain
reaction;
RT, reverse transcriptase;
kb, kilobase(s).
PU.1 and Interferon Consensus Sequence-binding Protein Regulate
the Myeloid Expression of the Human Toll-like Receptor 4 Gene*
§,,,, and
**
Department of Hematology and Oncology,
University of Regensburg, 93042 Regensburg, Germany, and Howard
Hughes Medical Institute and the ¶ Department of Internal
Medicine, University of Texas Southwestern Medical Center,
Dallas, Texas 75235-9050
ABSTRACT
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
B, nonreceptor tyrosine kinases, protein kinase C, and several
members of the mitogen-activated protein kinase family (7). CD14,
however, is a glycosyl-phosphatidylinositol-anchored membrane protein
that is unable to transduce the LPS signal through the membrane.
B (18-22).
EXPERIMENTAL PROCEDURES
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
8 M
-mercaptoethanol, 2% human pooled AB-group serum on Teflon foils
for a period of 7 days. The human monocytic cell line THP-1 was grown
in RPMI 1640 medium supplemented with 10% fetal calf serum (Life
Technologies, Inc.). The human cervical carcinoma cell line HeLa and
the human melanoma cell line Mel Im (a gift from A. Bosserhof) were
maintained in Dulbecco's modified Eagle's medium plus 10% fetal calf
serum. Where indicated, cells were treated with human recombinant
IFN-
(200 units/ml; Roche Molecular Biochemicals).
-actin are indicated in Fig. 1. PCR conditions
were optimized to assure that the amplification was still exponential
at the indicated cycle numbers. The amplified products were sequenced
to confirm their identity.
75
plasmid. The mouse PU.1 expression plasmid pECE-PU.1 was a gift from
Dr. R. Maki. The expression plasmid for human ICSBP (ICSBP-pTargeT) was
a gift from Dr. B. Levi. The open reading frame of PU.1 was subcloned into pGEM3 (Promega) for in vitro translation with the TNT
Coupled Reticulocyte Lysate System (Promega). For transient
transfections, plasmids were isolated and purified using the QiaFilter
Plasmid Midi Kit from Qiagen.
-32P]ATP using T4 polynucleotide kinase. The binding
reaction contained 2.5 µg of nuclear extract protein or 0.5-2 µl
of the in vitro translation reaction, 0.5 µg of
poly(dI-dC), 20 mM HEPES, pH 7.9, 60 mM KCl, 1 mM dithiothreitol, 1 mM EDTA, pH 8.0, 5%
glycerol, and 20 nmol of probe DNA in a final volume of 10 µl.
Antisera used in supershift analyses were added after 15 min, and
samples were loaded onto polyacrylamide gels after standing at room
temperature for a total of 30 min. Buffers and running conditions used
have been described (48). Gels were fixed in 5% acetic acid, dried, and autoradiographed.
RESULTS
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
View larger version (30K):
[in a new window]
Fig. 1.
The human TLR4 gene. A,
physical map of 19 kb of the human TLR4 gene. Boxes mark the
position of exons I-IV. In frame start and stop codons are indicated.
B, RT-PCR with total RNA from in vitro
differentiated macrophages and THP-1 cells using primers located in
exon I (86S, 5'-CAT GGC CTT CCT CTC CTG CGT G-3') and exon IV (769AS,
5'-GGC ATT TGA TGT AGA ACC CGC AAG TC-3'). The amplified fragments were
separated by agarose gel electrophoresis along with molecular weight
markers and stained with ethidium bromide. The size and composition of
each detected fragment is indicated. C, predicted N-terminal
amino acid sequences resulting from each alternative splicing product.
The N-terminal amino acid sequence resulting from transcripts
containing exons I, III, and IV is in bold lettering. Exon
sequences are given in capital letters, and intronic
sequences are in lowercase letters. All splice junctions
contain the expected GT splice donor and AG splice acceptor.
View larger version (48K):
[in a new window]
Fig. 2.
Determination of the transcriptional start
sites by primer extension. An oligonucleotide complementary
to the 5'-end of hTLR4 cDNA was 32P end-labeled and
hybridized to THP-1 mRNA. Extension products were separated on a
6% sequencing gel along with sequencing reactions primed with the
identical product. The positions of the two products are marked.
3228 and 743. Within this region elements may exist
that repress the activity of luciferase constructs specifically in
THP-1 cells. Reporter constructs including residues above 385 showed
low but significant reporter activity in transfected HeLa cells.
However, further deletion localizes a region directing macrophage-specific reporter gene expression to approximately 75 bp
proximal to the major transcriptional start site.
View larger version (22K):
[in a new window]
Fig. 3.
Deletion analysis of the human TLR4
promoter. Each deletion construct was transiently transfected into
myeloid THP-1 and nonmyeloid cell lines HeLa and Mel Im as described
under "Experimental Procedures." Luciferase activity is relative to
the empty control vector pGL3-B, and values are the means + S.D.
obtained from at least three independent experiments.
75) contains two
purine-rich elements that might be bound by members of the Ets family,
which includes the myeloid and B-cell-specific transcription factor PU.1. In addition, a highly conserved consensus-binding site for octamer transcription factors was detected in both human and mouse proximal TLR4 promoters.
View larger version (43K):
[in a new window]
Fig. 4.
Sequence alignment of murine and human TLR4
5'-flanking regions. Potential binding sites for transcription
factors are overlined (human TLR4) or underlined
(mouse TLR4). Transcriptional start sites are marked with
asterisks. Identical nucleotides in both sequences are in
bold lettering, and the three nucleotides of the start codon
(+1) are in bold italics and underlined.
75
Promoter--
The importance of PU.1 recognition motifs for
tissue-restricted expression of myeloid genes has been demonstrated in
a growing number of cases (50-56). To test the ability of PU.1 to bind
elements within the human TLR4 promoter, we performed EMSA with
double-stranded oligonucleotides corresponding to the two proximal
purine-rich sequences. Fig. 5A
shows an EMSA with in vitro translated PU.1, which
specifically bound to both oligonucleotides. In both cases, complex
formation was disrupted by the addition of an excess amount of
unlabeled wild type oligonucleotide but not by an oligonucleotide with
a mutated 5'-GGAA-3' core sequence. Both complexes were supershifted by
the addition of a PU.1-specific antiserum. Similar experiments were
done using nuclear extracts of THP-1 cells. As shown in Fig. 5B, the outer purine-rich motif (PU-11) formed a specific
quickly migrating complex similar to the complex observed with in
vitro translated PU.1. The complex was disrupted by the addition
of a PU.1-specific antiserum but remained unchanged in the presence of
antisera against two other Ets family members Fli-1 and Elf-1. The
specific complex observed for the inner purine-rich motif (PU-12) was
migrating significantly more slowly compared with the complex observed
with in vitro translated PU.1. The complex was also
supershifted in the presence of PU.1-specific antiserum but not by
antisera against Fli-1 and Elf-1. The different mobility of this
complex suggested the presence of at least one additional factor that
interacts with PU.1 on this site.
View larger version (36K):
[in a new window]
Fig. 5.
PU.1 binding to GGAA motifs within the
75 hTLR4 promoter. Labeled PU-11 and PU-12
oligonucleotides were used in EMSA with in vitro translated
PU.1 protein (A) or THP-1 nuclear proteins (B).
Addition of unlabeled oligonucleotides for competition analysis or
antisera against Ets family transcription factors are indicated above
each lane. PU.1 containing complexes are marked with arrows,
and antibody supershifts are marked with with SS, and
unspecific complexes are marked with with asterisks.
View larger version (57K):
[in a new window]
Fig. 6.
Specificity and stability of the slower
migrating complex. A, EMSA using labeled PU-12
oligonucleotide and nuclear extract preparations of various cell types.
B, THP-1 nuclear extracts were prepared either without
protease inhibitors or without the phosphatase inhibitor
Na3VO4 as indicated above each
lane. EMSA is shown for the labeled PU-12 oligonucleotide.
PU.1 containing complexes are marked with arrows, and
unspecific complexes are marked with with asterisks.
2-4
B and
E
3' (57). We next performed EMSAs with specific antisera
against the three IRF family members (ICSBP, interferon-stimulated gene
factor 3, and PIP) to identify the interacting partner of PU.1 in
the context of the hTLR4 promoter. As shown in Fig. 7B, a
strong supershift was detected with THP-1 extracts and the ICSBP
antiserum. A supershift (albeit less intense) was also observed with
the PIP antiserum, suggesting the presence of PIP, which is generally
thought to be lymphocyte-specific (57), or a PIP cross-reactive protein in THP-1 nuclear proteins. No supershift was detectable with the interferon-stimulated gene factor 3 antiserum. In vitro
translated ICSBP was unable to bind the PU-12 element independently but
was able to interact with PU.1 to form a complex with slightly
decreased mobility to that observed in THP-1 nuclear extracts (Fig.
7C). The complex of in vitro translated PU.1 and
ICSBP was supershifted with the ICSBP and the PIP antiserum, indicating
that the latter is cross-reactive with human ICSBP protein in gel shift
assays. These results suggested that ICSBP and PU.1 are the main
components of the slower migrating PU-12 complex. A similar band
pattern of ICSBP·PU.1 and predominantly Pip·PU.1 containing
complexes was observed in gel shift experiments performed with nuclear
extracts from human B cells, which also express PU.1 and both PIP and
ICSBP (data not shown). Sequence comparison revealed a single base pair difference in the IRF-binding site between mouse and human promoter (Fig. 3). Binding of the PU.1/ICSBP complex was also detectable using
the corresponding mouse sequence (mPU-12; Table
I) and THP-1 nuclear extracts, although
the band corresponding to PU.1 binding alone increased relative to the
human sequence (data not shown).
View larger version (25K):
[in a new window]
Fig. 7.
PU.1 interacts with ICSBP on the inner
purine-rich motif. A, comparison of known adjacent
PU.1/IRF-binding sequences with the inner purine rich motif (PU-12) of
the human TLR4 promoter. Conserved nucleotides are indicated in
bold lettering. B, labeled PU-12 oligonucleotide
and THP-1 nuclear proteins were used in supershift experiments with
antisera against IRF family transcription factors or PU.1 as indicated
above each lane. C, labeled PU-12
oligonucleotide was used in EMSA with in vitro translated
PU.1 protein, in vitro translated ICSBP, or both. Complexes
observed using THP-1 nuclear proteins are shown in comparison. PU.1
containing complexes are marked with arrows, antibody
supershifts are marked with with SS, and unspecific
complexes are marked with with asterisks.
Sequences of oligonucleotides used in gel shift analysis and
site-directed mutagenesis
View larger version (71K):
[in a new window]
Fig. 8.
The conserved octamer sequence is a weak
binding site for octamer factors. Labeled OCT consensus
oligonucleotide was used in EMSA with nuclear proteins from in
vitro differentiated macrophages. Addition of unlabeled
oligonucleotides for competition analysis or antisera against Oct
family transcription factors are indicated above each
lane. Oct1/2-containing complexes are marked with
arrows, antibody supershifts are marked with with
SS, and unspecific complexes are marked with with
asterisks.
--
To analyze the functional significance of the four
identified binding sites in reporter assays, we used site-directed
mutagenesis to abolish each of the four putative binding sequences.
The oligonucleotides carrying the mutated binding sites (PU-11M,
PU-12M, IRF-M, and OCT-M; for sequences see Table I) did not compete
with complex formation of wild type oligonucleotides in gel shift
assays (data not shown). In monocytic THP-1 cells the reporter
constructs with mutations of either half-side of the PU.1/ICSBP motif
showed a markedly reduced activity compared with the wild type promoter (Fig. 9). The effect of the PU.1 mutation
was most distinct, and reporter activity was almost completely
abolished. In contrast, disruption of the octamer-binding site showed a
trend toward a slightly increased activity, although statistical
analysis revealed that the difference was not significant. Mutation of
the outer PU.1 motif also had no significant impact on the activity of
the hTLR4 75 promoter (Fig. 9).
View larger version (25K):
[in a new window]
Fig. 9.
Effect of site-directed mutations on the
hTLR4 75 promoter activity in myeloid cells.
Site-directed mutations, were introduced in the hTLR4 75
promoter construct using oligonucleotides listed in Table I. A
schematic representation of the reporter gene constructs is shown. Four
putative binding sites are indicated as boxes,
P1 and P2 (outer and inner Ets
motif), O (octamer motif), and I (IFR
factor-binding site). Mutations are indicated by crosses. Each
site-directed mutant was transiently transfected into the myeloid cell
line THP-1 as described under "Experimental Procedures." Luciferase
activity is relative to the wild type hTLR4 75 promoter
(100%), and values are the means + S.D. obtained from at least four
independent experiments.
75 reporter plasmid and expression plasmids for PU.1
(PU-pECE) and/or ICSBP (hICSBP-pTarget). We only observed a slight
up-regulation of relative luciferase activity (1.5-fold) when PU.1 was
co-transfected alone (data not shown). Co-transfection of PU.1 and
ICSBP expression plasmids had no cooperative effect on the promoter
activity in HeLa cells with or without the addition of IFN-. Similar
results have been reported for the functional ICSBP·PU.1-binding
motif of the CYBB gene, which encodes for gp91phox (58). This
might indicate that additional factors, which are not present in HeLa
cells, are required for the induction of the TLR4 promoter by
PU.1/ICSBP or that inhibitory proteins are present that prevent
PU.1/ICSBP binding and transactivation.
, a cytokine released by activated T lymphocytes, is a major
activator of macrophage microbicidal functions and enhances both
sensitivity and magnitude of their response to LPS (36, 37, 39, 40,
61). In mouse macrophages, priming with IFN- drastically
up-regulates expression of the transcription factor ICSBP (62), which
has been implicated in IFN--mediated gene regulation (63-65). These
observations prompted us to investigate a possible effect of IFN- on
PU.1·ICSBP complex formation in macrophages. Constitutive binding of
the ICSBP·PU.1 complex observed in untreated cells was only slightly
increased during IFN- stimulation for 3, 6, or 18 h. No
additional bands were observed upon IFN- treatment, and the
ICSBP·PU.1 complex was completely supershifted by the ICSBP antiserum
(data not shown). Whereas high levels of IRF-1 transcripts were
detectable after IFN- treatment, suggesting efficient IFN-
priming (66), only a slight induction (approximately 2-fold) of all
three TLR4 splicing forms by IFN- treatment was detectable by RT-PCR
using total RNA from in vitro differentiated macrophages or
THP-1 cells (data not shown). IFN- treatment of THP-1 cells did not
affect the intensity of the ICSBP·PU.1-DNA complex in gel shift
assays, and the relative activities of TLR4 promoter constructs in
transient transfections of THP-1 cells were not affected by INF-
treatment either 24 or 4 h before cell harvest (data not shown).
DISCUSSION
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
-regulated protein
that bound an interferon-inducible enhancer element of major
histocompatibility complex class I genes (71). Because of the
phosphorylation of tyrosine residues in vivo, ICSBP does not
bind to DNA by itself but has been shown to interact with two other IRF
family members (IRF-1 and IRF-2) (72, 73). Cooperative DNA binding of
ICSBP with one of the other IRF family members results in an increased
binding activity for IFN-stimulated response elements and
transcriptional repression of genes containing the IFN-stimulated
response elements (57, 63-65, 74). In addition to its interaction with
members of its own gene family, ICSBP and the related
lymphoid-restricted factor Pip (ICSAT/IRF-4) are able to form complexes
with the Ets family member PU.1 (57). ICSBP (or Pip) protein bind a
composite IRF/Ets motif only in the presence of PU.1, and complex
formation requires the phosphorylation of PU.1 at serine 148 (57, 75).
Composite IRF/Ets-binding sites have been implicated in cell
type-specific gene expression in myeloid and B-cells. The CD20 promoter
and immunoglobulin light chain enhancers E2-4B and
E3' contain IRF/Ets-binding sites that seem to be
important for their B-cell-specific regulation and are bound by PU.1
and Pip (57, 59). An IRF/Ets motif, which is bound by PU.1 and ICSBP,
seems important for the myeloid-restricted expression of
gp91phox (58). Kim et al. (60) recently described an
ICSBP-binding site that was critical for the activity of the murine
5'-flanking IL-18 promoter. Although ICSBP usually does not bind DNA
alone, the authors did not investigate the presence of additional
factors in the observed ICSBP-DNA complex. The binding sequence of the IL-18 promoter is similar to other IRF/Ets motifs (Fig. 7A);
it is therefore likely that Ets factors (most likely PU.1) also
participate in the regulation of the 5'-flanking IL-18 promoter.
(62). If at all, ICSBP
and the IRF/Ets motif in the TLR4 promoter only weakly confer IFN-
responsiveness. The ICSBP·PU.1 complex was slightly induced by
IFN- in human macrophages, and in both THP-1 cells and human macrophages, TLR4 mRNA expression was slightly enhanced by IFN- priming. However, IFN- stimulation did not significantly affect the
activity of TLR4 promoter constructs in THP-1 cells. It remains to be
clarified whether the observed increase in TLR4 message is regulated on
the level of gene transcription or whether other mechanisms
(e.g. mRNA stability) mediate the IFN- induced
increase in TLR4 mRNA.
ACKNOWLEDGEMENTS
FOOTNOTES
ABBREVIATIONS
REFERENCES
TOP
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS
DISCUSSION
REFERENCES
1.
Morrison, D. C.,
and Ryan, J. L.
(1987)
Annu. Rev. Med.
38,
417-432[Medline]
[Order article via Infotrieve]
2.
Gutierrez-Ramos, J. C.,
and Bluethmann, H.
(1997)
Immunol. Today
18,
329-334[CrossRef][Medline]
[Order article via Infotrieve]
3.
Ulevitch, R. J.,
and Tobias, P. S.
(1999)
Curr. Opin. Immunol.
11,
19-22[CrossRef][Medline]
[Order article via Infotrieve]
4.
Wright, S. D.,
Ramos, R. A.,
Tobias, P. S.,
Ulevitch, R. J.,
and Mathison, J. C.
(1990)
Science
249,
1431-1433 5.
Tobias, P. S.,
Soldau, K.,
and Ulevitch, R. J.
(1989)
J. Biol. Chem.
264,
10867-10871 6.
Schumann, R. R.,
Leong, S. R.,
Flaggs, G. W.,
Gray, P. W.,
Wright, S. D.,
Mathison, J. C.,
Tobias, P. S.,
and Ulevitch, R. J.
(1990)
Science
249,
1429-1431 7.
Sweet, M. J.,
and Hume, D. A.
(1996)
J. Leukocyte Biol.
60,
8-26[Abstract]
8.
Hoffmann, J. A.,
Reichhart, J. M.,
and Hetru, C.
(1996)
Curr. Opin. Immunol.
8,
8-13[CrossRef][Medline]
[Order article via Infotrieve]
9.
Kobe, B.,
and Deisenhofer, J.
(1995)
Nature
374,
183-186[CrossRef][Medline]
[Order article via Infotrieve]
10.
Kopp, E. B.,
and Medzhitov, R.
(1999)
Curr. Opin. Immunol.
11,
13-18[CrossRef][Medline]
[Order article via Infotrieve]
11.
Gay, N. J.,
and Keith, F. J.
(1991)
Nature
351,
355-356
12.
Torigoe, K.,
Ushio, S.,
Okura, T.,
Kobayashi, S.,
Taniai, M.,
Kunikata, T.,
Murakami, T.,
Sanou, O.,
Kojima, H.,
Fujii, M.,
Ohta, T.,
Ikeda, M.,
Ikegami, H.,
and Kurimoto, M.
(1997)
J. Biol. Chem.
272,
25737-25742 13.
Thomassen, E.,
Renshaw, B. R.,
and Sims, J. E.
(1999)
Cytokine
11,
389-399[CrossRef][Medline]
[Order article via Infotrieve]
14.
Mitcham, J. L.,
Parnet, P.,
Bonnert, T. P.,
Garka, K. E.,
Gerhart, M. J.,
Slack, J. L.,
Gayle, M. A.,
Dower, S. K.,
and Sims, J. E.
(1996)
J. Biol. Chem.
271,
5777-5783 15.
Medzhitov, R.,
Preston-Hurlburt, P.,
and Janeway, C. A.
(1997)
Nature
388,
394-397[CrossRef][Medline]
[Order article via Infotrieve]
16.
Rock, F. L.,
Hardiman, G.,
Timans, J. C.,
Kastelein, R. A.,
and Bazan, J. F.
(1998)
Proc. Natl. Acad. Sci. U. S. A.
95,
588-593 17.
Takeuchi, O.,
Kawai, T.,
Sanjo, H.,
Copeland, N. G.,
Gilbert, D. J.,
Jenkins, N. A.,
Takeda, K.,
and Akira, S.
(1999)
Gene (Amst.)
231,
59-65[CrossRef][Medline]
[Order article via Infotrieve]
18.
Cao, Z.,
Xiong, J.,
Takeuchi, M.,
Kurama, T.,
and Goeddel, D. V.
(1996)
Nature
383,
443-446[CrossRef][Medline]
[Order article via Infotrieve]
19.
Cao, Z.,
Henzel, W. J.,
and Gao, X.
(1996)
Science
271,
1128-1131[Abstract]
20.
Muzio, M.,
Ni, J.,
Feng, P.,
and Dixit, V. M.
(1997)
Science
278,
1612-1615 21.
Muzio, M.,
Natoli, G.,
Saccani, S.,
Levrero, M.,
and Mantovani, A.
(1998)
J. Exp. Med.
187,
2097-2101 22.
Wesche, H.,
Henzel, W. J.,
Shillinglaw, W.,
Li, S.,
and Cao, Z.
(1997)
Immunity
7,
837-847[CrossRef][Medline]
[Order article via Infotrieve]
23.
Coutinho, A.,
Forni, L.,
Melchers, F.,
and Watanabe, T.
(1977)
Eur. J. Immunol.
7,
325-328[Medline]
[Order article via Infotrieve]
24.
Sultzer, B. M.
(1968)
Nature
219,
1253-1254[CrossRef][Medline]
[Order article via Infotrieve]
25.
Poltorak, A.,
He, X.,
Smirnova, I.,
Liu, M. Y.,
Huffel, C. V.,
Du, X.,
Birdwell, D.,
Alejos, E.,
Silva, M.,
Galanos, C.,
Freudenberg, M.,
Ricciardi-Castagnoli, P.,
Layton, B.,
and Beutler, B.
(1998)
Science
282,
2085-2088 26.
Scharton-Kersten, T.,
Contursi, C.,
Masumi, A.,
Sher, A.,
and Ozato, K.
(1997)
J. Exp. Med.
186,
1523-1534 27.
Shimazu, R.,
Akashi, S.,
Ogata, H.,
Nagai, Y.,
Fukudome, K.,
Miyake, K.,
and Kimoto, M.
(1999)
J. Exp. Med.
189,
1777-1782 28.
Berczi, I.,
Bertok, L.,
and Bereznai, T.
(1966)
Can. J. Microbiol.
12,
1070-1071[Medline]
[Order article via Infotrieve]
29.
Beutler, B.,
Krochin, N.,
Milsark, I. W.,
Luedke, C.,
and Cerami, A.
(1986)
Science
232,
977-980 30.
Haranaka, K.,
Satomi, N.,
Sakurai, A.,
and Haranaka, R.
(1984)
Cancer Immunol. Immunother.
18,
87-90[Medline]
[Order article via Infotrieve]
31.
Vogel, S. N.,
Moore, R. N.,
Sipe, J. D.,
and Rosenstreich, D. L.
(1980)
J. Immunol.
124,
2004-2009[Abstract]
32.
Green, S.,
Dobrjansky, A.,
Chiasson, M. A.,
Carswell, E.,
Schwartz, M. K.,
and Old, L. J.
(1977)
J. Natl. Cancer Inst.
59,
1519-1522
33.
Matsuura, M.,
and Galanos, C.
(1990)
Infect. Immun.
58,
935-937 34.
Vogel, S. N.,
and Fertsch, D.
(1984)
Infect. Immun.
45,
417-423 35.
Vogel, S. N.,
Weedon, L. L.,
Moore, R. N.,
and Rosenstreich, D. L.
(1982)
J. Immunol.
128,
380-387[Abstract]
36.
Lau, A. S.,
and Livesey, J. F.
(1989)
J. Clin. Invest.
84,
738-743
37.
Beutler, B.,
Tkacenko, V.,
Milsark, I.,
Krochin, N.,
and Cerami, A.
(1986)
J. Exp. Med.
164,
1791-1796 38.
Pace, J. L.,
Russell, S. W.,
LeBlanc, P. A.,
and Murasko, D. M.
(1985)
J. Immunol.
134,
977-981[Abstract]
39.
Hayes, M. P.,
and Zoon, K. C.
(1993)
Infect. Immun.
61,
3222-3227 40.
Mathison, J. C.,
Virca, G. D.,
Wolfson, E.,
Tobias, P. S.,
Glaser, K.,
and Ulevitch, R. J.
(1990)
J. Clin. Invest.
85,
1108-1118
41.
Zhang, F. X.,
Kirschning, C. J.,
Mancinelli, R.,
Xu, X. P.,
Jin, Y.,
Faure, E.,
Mantovani, A.,
Rothe, M.,
Muzio, M.,
and Arditi, M.
(1999)
J. Biol. Chem.
274,
7611-7614 42.
Andreesen, R.,
Brugger, W.,
Scheibenbogen, C.,
Kreutz, M.,
Leser, H. G.,
Rehm, A.,
and Lohr, G. W.
(1990)
J. Leukocyte Biol.
47,
490-497[Abstract]
43.
Poltorak, A.,
Smirnova, I.,
He, X.,
Liu, M. Y.,
Van Huffel, C.,
McNally, O.,
Birdwell, D.,
Alejos, E.,
Silva, M.,
Du, X.,
Thompson, P.,
Chan, E. K.,
Ledesma, J.,
Roe, B.,
Clifton, S.,
Vogel, S. N.,
and Beutler, B.
(1998)
Blood Cells Mol. Dis.
24,
340-355[CrossRef][Medline]
[Order article via Infotrieve]
44.
Chomczynski, P.,
and Sacchi, N.
(1987)
Anal. Biochem.
162,
156-159[Medline]
[Order article via Infotrieve]
45.
Calzone, F. J.,
Britten, R. J.,
and Davidson, E. H.
(1987)
Methods Enzymol.
152,
611-632[Medline]
[Order article via Infotrieve]
46.
Sambrock, J.,
Fritsch, E. F.,
and Maniatis, T.
(1989)
Molecular Cloning: A Laboratory Manual
, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY
47.
Osborn, L.,
Kunkel, S.,
and Nabel, G. J.
(1989)
Proc. Natl. Acad. Sci. U. S. A.
86,
2336-2340 48.
Ross, I. L.,
Dunn, T. L.,
Yue, X.,
Roy, S.,
Barnett, C. J.,
and Hume, D. A.
(1994)
Oncogene
9,
121-132[Medline]
[Order article via Infotrieve]
49.
Rehli, M.,
Lichanska, A.,
Cassady, A. I.,
Ostrowski, M. C.,
and Hume, D. A.
(1999)
J. Immunol.
162,
1559-1565 50.
Chen, H.,
Ray-Gallet, D.,
Zhang, P.,
Hetherington, C. J.,
Gonzalez, D. A.,
Zhang, D. E.,
Moreau-Gachelin, F.,
and Tenen, D. G.
(1995)
Oncogene
11,
1549-1560[Medline]
[Order article via Infotrieve]
51.
Eichbaum, Q. G.,
Iyer, R.,
Raveh, D. P.,
Mathieu, C.,
and Ezekowitz, R. A.
(1994)
J. Exp. Med.
179,
1985-1996 52.
Feinman, R.,
Qiu, W. Q.,
Pearse, R. N.,
Nikolajczyk, B. S.,
Sen, R.,
Sheffery, M.,
and Ravetch, J. V.
(1994)
EMBO J.
13,
3852-3860[Medline]
[Order article via Infotrieve]
53.
Heydemann, A.,
Juang, G.,
Hennessy, K.,
Parmacek, M. S.,
and Simon, M. C.
(1996)
Mol. Cell. Biol.
16,
1676-1686[Abstract]
54.
Pahl, H. L.,
Scheibe, R. J.,
Zhang, D. E.,
Chen, H. M.,
Galson, D. L.,
Maki, R. A.,
and Tenen, D. G.
(1993)
J. Biol. Chem.
268,
5014-5020 55.
Perez, C.,
Coeffier, E.,
Moreau-Gachelin, F.,
Wietzerbin, J.,
and Benech, P. D.
(1994)
Mol. Cell. Biol.
14,
5023-5031 56.
Zhang, D. E.,
Hetherington, C. J.,
Chen, H. M.,
and Tenen, D. G.
(1994)
Mol. Cell. Biol.
14,
373-381 57.
Eisenbeis, C. F.,
Singh, H.,
and Storb, U.
(1995)
Genes Dev.
9,
1377-1387 58.
Eklund, E. A.,
Jalava, A.,
and Kakar, R.
(1998)
J. Biol. Chem.
273,
13957-13965 59.
Himmelmann, A.,
Riva, A.,
Wilson, G. L.,
Lucas, B. P.,
Thevenin, C.,
and Kehrl, J. H.
(1997)
Blood
90,
3984-3995 60.
Kim, Y. M.,
Kang, H. S.,
Paik, S. G.,
Pyun, K. H.,
Anderson, K. L.,
Torbett, B. E.,
and Choi, I.
(1999)
J. Immunol.
163,
2000-2007 61.
Collart, M. A.,
Belin, D.,
Vassalli, J. D.,
de, K., S.,
and Vassalli, P.
(1986)
J. Exp. Med.
164,
2113-2118 62.
Politis, A. D.,
Ozato, K.,
Coligan, J. E.,
and Vogel, S. N.
(1994)
J. Immunol.
152,
2270-2278[Abstract]
63.
Nelson, N.,
Marks, M. S.,
Driggers, P. H.,
and Ozato, K.
(1993)
Mol. Cell. Biol.
13,
588-599 64.
Weisz, A.,
Marx, P.,
Sharf, R.,
Appella, E.,
Driggers, P. H.,
Ozato, K.,
and Levi, B. Z.
(1992)
J. Biol. Chem.
267,
25589-25596 65.
Weisz, A.,
Kirchhoff, S.,
and Levi, B. Z.
(1994)
Int. Immunol.
6,
1125-1131 66.
Lehtonen, A.,
Matikainen, S.,
and Julkunen, I.
(1997)
J. Immunol.
159,
794-803[Abstract]
67.
Lopez, A. J.
(1998)
Annu. Rev. Genet.
32,
279-305[CrossRef][Medline]
[Order article via Infotrieve], 279-305
68.
Eichbaum, Q.,
Heney, D.,
Raveh, D.,
Chung, M.,
Davidson, M.,
Epstein, J.,
and Ezekowitz, R. A.
(1997)
Blood
90,
4135-4143 69.
Ross, I. L.,
Yue, X.,
Ostrowski, M. C.,
and Hume, D. A.
(1998)
J. Biol. Chem.
273,
6662-6669 70.
Suzuki, S.,
Kumatori, A.,
Haagen, I. A.,
Fujii, Y.,
Sadat, M. A.,
Jun, H. L.,
Tsuji, Y.,
Roos, D.,
and Nakamura, M.
(1998)
Proc. Natl. Acad. Sci. U. S. A.
95,
6085-6090 71.
Driggers, P. H.,
Ennist, D. L.,
Gleason, S. L.,
Mak, W. H.,
Marks, M. S.,
Levi, B. Z.,
Flanagan, J. R.,
Appella, E.,
and Ozato, K.
(1990)
Proc. Natl. Acad. Sci. U. S. A.
87,
3743-3747 72.
Sharf, R.,
Meraro, D.,
Azriel, A.,
Thornton, A. M.,
Ozato, K.,
Petricoin, E. F.,
Larner, A. C.,
Schaper, F.,
Hauser, H.,
and Levi, B. Z.
(1997)
J. Biol. Chem.
272,
9785-9792 73.
Bovolenta, C.,
Driggers, P. H.,
Marks, M. S.,
Medin, J. A.,
Politis, A. D.,
Vogel, S. N.,
Levy, D. E.,
Sakaguchi, K.,
Appella, E.,
and Coligan, J. E.
(1994)
Proc. Natl. Acad. Sci. U. S. A.
91,
5046-5050 74.
Sharf, R.,
Azriel, A.,
Lejbkowicz, F.,
Winograd, S. S.,
Ehrlich, R.,
and Levi, B. Z.
(1995)
J. Biol. Chem.
270,
13063-13069 75.
Ortiz, M. A.,
Light, J.,
Maki, R. A.,
and Assa-Munt, N.
(1999)
Proc. Natl. Acad. Sci. U. S. A.
96,
2740-2745 76.
Holtschke, T.,
Lohler, J.,
Kanno, Y.,
Fehr, T.,
Giese, N.,
Rosenbauer, F.,
Lou, J.,
Knobeloch, K. P.,
Gabriele, L.,
Waring, J. F.,
Bachmann, M. F.,
Zinkernagel, R. M.,
Morse, H. C.,
Ozato, K.,
and Horak, I.
(1996)
Cell
87,
307-317[CrossRef][Medline]
[Order article via Infotrieve]
77.
Giese, N. A.,
Gabriele, L.,
Doherty, T. M.,
Klinman, D. M.,
Tadesse-Heath, L.,
Contursi, C.,
Epstein, S. L.,
and Morse, H. C.
(1997)
J. Exp. Med.
186,
1535-1546
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  M. Joo, M. Kwon, Y.-J. Cho, N. Hu, T. V. Pedchenko, R. T. Sadikot, T. S. Blackwell, and J. W. Christman Lipopolysaccharide-dependent interaction between PU.1 and cJun determines production of lipocalin-type prostaglandin D synthase and prostaglandin D2 in macrophages Am J Physiol Lung Cell Mol Physiol, May 1, 2009; 296(5): L771 - L779. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. L. Shaffer, N.C. T. Emre, P. B. Romesser, and L. M. Staudt IRF4: Immunity. Malignancy! Therapy? Clin. Cancer Res., May 1, 2009; 15(9): 2954 - 2961. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Shin, Y. Zhang, M. Jagannathan, H. Hasturk, A. Kantarci, H. Liu, T. E. Van Dyke, L. M. Ganley-Leal, and B. S. Nikolajczyk B cells from periodontal disease patients express surface Toll-like receptor 4 J. Leukoc. Biol., April 1, 2009; 85(4): 648 - 655. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. Regueiro, D. Moranta, M. A. Campos, J. Margareto, J. Garmendia, and J. A. Bengoechea Klebsiella pneumoniae Increases the Levels of Toll-Like Receptors 2 and 4 in Human Airway Epithelial Cells Infect. Immun., February 1, 2009; 77(2): 714 - 724. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Ghanim, P. Mohanty, R. Deopurkar, C. Ling Sia, K. Korzeniewski, S. Abuaysheh, A. Chaudhuri, and P. Dandona Acute Modulation of Toll-Like Receptors by Insulin Diabetes Care, September 1, 2008; 31(9): 1827 - 1831. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. Huang, C. Zhu, H. Wang, E. Horvath, and E. A. Eklund The Interferon Consensus Sequence-binding Protein (ICSBP/IRF8) Represses PTPN13 Gene Transcription in Differentiating Myeloid Cells J. Biol. Chem., March 21, 2008; 283(12): 7921 - 7935. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Zhu, S. Lindsey, I. Konieczna, and E. A. Eklund Constitutive activation of SHP2 protein tyrosine phosphatase inhibits ICSBP-induced transcription of the gene encoding gp91PHOX during myeloid differentiation J. Leukoc. Biol., March 1, 2008; 83(3): 680 - 691. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Alter-Koltunoff, S. Goren, J. Nousbeck, C. G. Feng, A. Sher, K. Ozato, A. Azriel, and B.-Z. Levi Innate Immunity to Intraphagosomal Pathogens Is Mediated by Interferon Regulatory Factor 8 (IRF-8) That Stimulates the Expression of Macrophage-specific Nramp1 through Antagonizing Repression by c-Myc J. Biol. Chem., February 1, 2008; 283(5): 2724 - 2733. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Weigelt, W. Ernst, Y. Walczak, S. Ebert, T. Loenhardt, M. Klug, M. Rehli, B. H. F. Weber, and T. Langmann Dap12 expression in activated microglia from retinoschisin-deficient retina and its PU.1-dependent promoter regulation J. Leukoc. Biol., December 1, 2007; 82(6): 1564 - 1574. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Lichtinger, R. Ingram, M. Hornef, C. Bonifer, and M. Rehli Transcription Factor PU.1 Controls Transcription Start Site Positioning and Alternative TLR4 Promoter Usage J. Biol. Chem., September 14, 2007; 282(37): 26874 - 26883. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Turcotte, S. Gauthier, D. Malo, M. Tam, M. M. Stevenson, and P. Gros Icsbp1/IRF-8 Is Required for Innate and Adaptive Immune Responses against Intracellular Pathogens J. Immunol., August 15, 2007; 179(4): 2467 - 2476. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T.-H. Pham, S. Langmann, L. Schwarzfischer, C. El Chartouni, M. Lichtinger, M. Klug, S. W. Krause, and M. Rehli CCAAT Enhancer-binding Protein beta Regulates Constitutive Gene Expression during Late Stages of Monocyte to Macrophage Differentiation J. Biol. Chem., July 27, 2007; 282(30): 21924 - 21933. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Schroder, M. Lichtinger, K. M. Irvine, K. Brion, A. Trieu, I. L. Ross, T. Ravasi, K. J. Stacey, M. Rehli, D. A. Hume, et al. PU.1 and ICSBP control constitutive and IFN-{gamma}-regulated Tlr9 gene expression in mouse macrophages J. Leukoc. Biol., June 1, 2007; 81(6): 1577 - 1590. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Foster, S. R. Lea, P. M. Preshaw, and J. J. Taylor Pivotal Advance: Vasoactive intestinal peptide inhibits up-regulation of human monocyte TLR2 and TLR4 by LPS and differentiation of monocytes to macrophages J. Leukoc. Biol., April 1, 2007; 81(4): 893 - 903. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. Huang, E. Horvath, and E. A. Eklund PU.1, Interferon Regulatory Factor (IRF) 2, and the Interferon Consensus Sequence-binding Protein (ICSBP/IRF8) Cooperate to Activate NF1 Transcription in Differentiating Myeloid Cells J. Biol. Chem., March 2, 2007; 282(9): 6629 - 6643. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Q. M. Nhu, N. Cuesta, and S. N. Vogel Transcriptional regulation of lipopolysaccharide (LPS)-induced Toll-like receptor (TLR) expression in murine macrophages: role of interferon regulatory factors 1 (IRF-1) and 2 (IRF-2) Innate Immunity, October 1, 2006; 12(5): 285 - 295. [Abstract] [PDF]
|
|
|
|
 |
|
 W. Huang, G. Saberwal, E. Horvath, C. Zhu, S. Lindsey, and E. A. Eklund Leukemia-Associated, Constitutively Active Mutants of SHP2 Protein Tyrosine Phosphatase Inhibit NF1 Transcriptional Activation by the Interferon Consensus Sequence Binding Protein. Mol. Cell. Biol., September 1, 2006; 26(17): 6311 - 6332. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. S. Sharma, I. Leyva, F. Schenkel, and N. A. Karrow Association of toll-like receptor 4 polymorphisms with somatic cell score and lactation persistency in holstein bulls. J Dairy Sci, September 1, 2006; 89(9): 3626 - 3635. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Datta, U. Sinha-Datta, N. K. Dhillon, S. Buch, and C. Nicot The HTLV-I p30 Interferes with TLR4 Signaling and Modulates the Release of Pro- and Anti-inflammatory Cytokines from Human Macrophages J. Biol. Chem., August 18, 2006; 281(33): 23414 - 23424. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Tsatsanis, A. Androulidaki, T. Alissafi, I. Charalampopoulos, E. Dermitzaki, T. Roger, A. Gravanis, and A. N. Margioris Corticotropin-Releasing Factor and the Urocortins Induce the Expression of TLR4 in Macrophages via Activation of the Transcription Factors PU.1 and AP-1 J. Immunol., February 1, 2006; 176(3): 1869 - 1877. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. Machida, K. T. H. Cheng, V. M.-H. Sung, A. M. Levine, S. Foung, and M. M. C. Lai Hepatitis C Virus Induces Toll-Like Receptor 4 Expression, Leading to Enhanced Production of Beta Interferon and Interleukin-6 J. Virol., January 15, 2006; 80(2): 866 - 874. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Grisaru, M. Pick, C. Perry, E. H. Sklan, R. Almog, I. Goldberg, E. Naparstek, J. B. Lessing, H. Soreq, and V. Deutsch Hydrolytic and Nonenzymatic Functions of Acetylcholinesterase Comodulate Hemopoietic Stress Responses J. Immunol., January 1, 2006; 176(1): 27 - 35. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. R. Blais, S. G. Vascotto, M. Griffith, and I. Altosaar LBP and CD14 Secreted in Tears by the Lacrimal Glands Modulate the LPS Response of Corneal Epithelial Cells Invest. Ophthalmol. Vis. Sci., November 1, 2005; 46(11): 4235 - 4244. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Dominguez-Soto, A. Puig-Kroger, M. A. Vega, and A. L. Corbi PU.1 Regulates the Tissue-specific Expression of Dendritic Cell-specific Intercellular Adhesion Molecule (ICAM)-3-grabbing Nonintegrin J. Biol. Chem., September 30, 2005; 280(39): 33123 - 33131. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Tamura, P. Thotakura, T. S. Tanaka, M. S. H. Ko, and K. Ozato Identification of target genes and a unique cis element regulated by IRF-8 in developing macrophages Blood, September 15, 2005; 106(6): 1938 - 1947. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. V. Pedchenko, G. Y. Park, M. Joo, T. S. Blackwell, and J. W. Christman Inducible binding of PU.1 and interacting proteins to the Toll-like receptor 4 promoter during endotoxemia Am J Physiol Lung Cell Mol Physiol, September 1, 2005; 289(3): L429 - L437. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. P. Gomariz, A. Arranz, C. Abad, M. Torroba, C. Martinez, F. Rosignoli, M. Garcia-Gomez, J. Leceta, and Y. Juarranz Time-course expression of Toll-like receptors 2 and 4 in inflammatory bowel disease and homeostatic effect of VIP J. Leukoc. Biol., August 1, 2005; 78(2): 491 - 502. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X. Zhou, X.-P. Gao, J. Fan, Q. Liu, K. N. Anwar, R. S. Frey, and A. B. Malik LPS activation of Toll-like receptor 4 signals CD11b/CD18 expression in neutrophils Am J Physiol Lung Cell Mol Physiol, April 1, 2005; 288(4): L655 - L662. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Chakravarty and M. Herkenham Toll-Like Receptor 4 on Nonhematopoietic Cells Sustains CNS Inflammation during Endotoxemia, Independent of Systemic Cytokines J. Neurosci., February 16, 2005; 25(7): 1788 - 1796. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Musikacharoen, A. Oguma, Y. Yoshikai, N. Chiba, A. Masuda, and T. Matsuguchi Interleukin-15 induces IL-12 receptor {beta}1 gene expression through PU.1 and IRF 3 by targeting chromatin remodeling Blood, January 15, 2005; 105(2): 711 - 720. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Ito, C. Nishiyama, M. Nishiyama, H. Matsuda, K. Maeda, Y. Akizawa, R. Tsuboi, K. Okumura, and H. Ogawa Mast Cells Acquire Monocyte-Specific Gene Expression and Monocyte-Like Morphology by Overproduction of PU.1 J. Immunol., January 1, 2005; 174(1): 376 - 383. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Zhu, G. Saberwal, Y. Lu, L. C. Platanias, and E. A. Eklund The Interferon Consensus Sequence-binding Protein Activates Transcription of the Gene Encoding Neurofibromin 1 J. Biol. Chem., December 3, 2004; 279(49): 50874 - 50885. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Takeshita, K. Suzuki, S. Sasaki, N. Ishii, D. M. Klinman, and K. J. Ishii Transcriptional Regulation of the Human TLR9 Gene J. Immunol., August 15, 2004; 173(4): 2552 - 2561. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Armstrong, A. R. L. Medford, K. M. Uppington, J. Robertson, I. R. Witherden, T. D. Tetley, and A. B. Millar Expression of Functional Toll-Like Receptor-2 and -4 on Alveolar Epithelial Cells Am. J. Respir. Cell Mol. Biol., August 1, 2004; 31(2): 241 - 245. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Rehli, H.-H. Niller, C. Ammon, S. Langmann, L. Schwarzfischer, R. Andreesen, and S. W. Krause Transcriptional Regulation of CHI3L1, a Marker Gene for Late Stages of Macrophage Differentiation J. Biol. Chem., November 7, 2003; 278(45): 44058 - 44067. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Zhu, K. Rao, H. Xiong, K. Gagnidze, F. Li, C. Horvath, and S. Plevy Activation of the Murine Interleukin-12 p40 Promoter by Functional Interactions between NFAT and ICSBP J. Biol. Chem., October 10, 2003; 278(41): 39372 - 39382. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. T. Abreu, L. S. Thomas, E. T. Arnold, K. Lukasek, K. S. Michelsen, and M. Arditi TLR signaling at the intestinal epithelial interface Innate Immunity, October 1, 2003; 9(5): 322 - 330. [Abstract] [PDF]
|
|
|
|
|
|
S. Heinz, V. Haehnel, M. Karaghiosoff, L. Schwarzfischer, M. Muller, S. W. Krause, and M. Rehli Species-specific Regulation of Toll-like Receptor 3 Genes in Men and Mice J. Biol. Chem., June 6, 2003; 278(24): 21502 - 21509. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. O'Reilly, C. M. Quinn, T. El-Shanawany, S. Gordon, and D. R. Greaves Multiple Ets Factors and Interferon Regulatory Factor-4 Modulate CD68 Expression in a Cell Type-specific Manner J. Biol. Chem., June 6, 2003; 278(24): 21909 - 21919. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Tamai, S. Sugawara, O. Takeuchi, S. Akira, and H. Takada Synergistic effects of lipopolysaccharide and interferon-{gamma} in inducing interleukin-8 production in human monocytic THP-1 cells is accompanied by up-regulation of CD14, Toll-like receptor 4, MD-2 and MyD88 expression Innate Immunity, June 1, 2003; 9(3): 145 - 153. [Abstract] [PDF]
|
|
|
|
|
|
M. D. Bjerregaard, J. Jurlander, P. Klausen, N. Borregaard, and J. B. Cowland The in vivo profile of transcription factors during neutrophil differentiation in human bone marrow Blood, June 1, 2003; 101(11): 4322 - 4332. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. K. Means, F. Hayashi, K. D. Smith, A. Aderem, and A. D. Luster The Toll-Like Receptor 5 Stimulus Bacterial Flagellin Induces Maturation and Chemokine Production in Human Dendritic Cells J. Immunol., May 15, 2003; 170(10): 5165 - 5175. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. J. Barnes, A. E. Field, and P. M. Pitha-Rowe Virus-induced Heterodimer Formation between IRF-5 and IRF-7 Modulates Assembly of the IFNA Enhanceosome in Vivo and Transcriptional Activity of IFNA Genes J. Biol. Chem., May 2, 2003; 278(19): 16630 - 16641. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Xiong, C. Zhu, H. Li, F. Chen, L. Mayer, K. Ozato, J. C. Unkeless, and S. E. Plevy Complex Formation of the Interferon (IFN) Consensus Sequence-binding Protein with IRF-1 Is Essential for Murine Macrophage IFN-gamma -induced iNOS Gene Expression J. Biol. Chem., January 17, 2003; 278(4): 2271 - 2277. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Kuwata, C. Gongora, Y. Kanno, K. Sakaguchi, T. Tamura, T. Kanno, V. Basrur, R. Martinez, E. Appella, T. Golub, et al. Gamma Interferon Triggers Interaction between ICSBP (IRF-8) and TEL, Recruiting the Histone Deacetylase HDAC3 to the Interferon-Responsive Element Mol. Cell. Biol., November 1, 2002; 22(21): 7439 - 7448. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. A. Eklund, I. Goldenberg, Y. Lu, J. Andrejic, and R. Kakar SHP1 Protein-tyrosine Phosphatase Regulates HoxA10 DNA Binding and Transcriptional Repression Activity in Undifferentiated Myeloid Cells J. Biol. Chem., September 20, 2002; 277(39): 36878 - 36888. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Ishida, H. Kubo, S. Suzuki, T. Suzuki, S. Akashi, K. Inoue, S. Maeda, H. Kikuchi, H. Sasaki, and T. Kondo Hypoxia Diminishes Toll-Like Receptor 4 Expression Through Reactive Oxygen Species Generated by Mitochondria in Endothelial Cells J. Immunol., August 15, 2002; 169(4): 2069 - 2075. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Haehnel, L. Schwarzfischer, M. J. Fenton, and M. Rehli Transcriptional Regulation of the Human Toll-Like Receptor 2 Gene in Monocytes and Macrophages J. Immunol., June 1, 2002; 168(11): 5629 - 5637. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. T. Abreu, E. T. Arnold, L. S. Thomas, R. Gonsky, Y. Zhou, B. Hu, and M. Arditi TLR4 and MD-2 Expression Is Regulated by Immune-mediated Signals in Human Intestinal Epithelial Cells J. Biol. Chem., May 31, 2002; 277(23): 20431 - 20437. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Bosisio, N. Polentarutti, M. Sironi, S. Bernasconi, K. Miyake, G. R. Webb, M. U. Martin, A. Mantovani, and M. Muzio Stimulation of toll-like receptor 4 expression in human mononuclear phagocytes by interferon-gamma : a molecular basis for priming and synergism with bacterial lipopolysaccharide Blood, May 1, 2002; 99(9): 3427 - 3431. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Kumatori, D. Yang, S. Suzuki, and M. Nakamura Cooperation of STAT-1 and IRF-1 in Interferon-gamma -induced Transcription of the gp91phox Gene J. Biol. Chem., March 8, 2002; 277(11): 9103 - 9111. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Tamai, T. Sakuta, K. Matsushita, M. Torii, O. Takeuchi, S. Akira, S. Akashi, T. Espevik, S. Sugawara, and H. Takada Human Gingival CD14+ Fibroblasts Primed with Gamma Interferon Increase Production of Interleukin-8 in Response to Lipopolysaccharide through Up-Regulation of Membrane CD14 and MyD88 mRNA Expression Infect. Immun., March 1, 2002; 70(3): 1272 - 1278. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. G. A. M. Wolfs, W. A. Buurman, A. van Schadewijk, B. de Vries, M. A. R. C. Daemen, P. S. Hiemstra, and C. van 't Veer In Vivo Expression of Toll-Like Receptor 2 and 4 by Renal Epithelial Cells: IFN-{gamma} and TNF-{alpha} Mediated Up-Regulation During Inflammation J. Immunol., February 1, 2002; 168(3): 1286 - 1293. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Thiele, M. Wasner, C. Muller, K. Engeland, and S. Hauschildt Regulation and Possible Function of {beta}-Catenin in Human Monocytes J. Immunol., December 15, 2001; 167(12): 6786 - 6793. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Wang, W. P. Lafuse, and B. S. Zwilling NF{kappa}B and Sp1 Elements Are Necessary for Maximal Transcription of Toll-like Receptor 2 Induced by Mycobacterium avium J. Immunol., December 15, 2001; 167(12): 6924 - 6932. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. N. Laribee and M. J. Klemsz Loss of PU.1 Expression Following Inhibition of Histone Deacetylases J. Immunol., November 1, 2001; 167(9): 5160 - 5166. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Alexander and E. Th. Rietschel Invited review: Bacterial lipopolysaccharides and innate immunity Innate Immunity, June 1, 2001; 7(3): 167 - 202. [Abstract] [PDF]
|
|
|
|
|
|
S. Marecki, C. J. Riendeau, M. D. Liang, and M. J. Fenton PU.1 and Multiple IFN Regulatory Factor Proteins Synergize to Mediate Transcriptional Activation of the Human IL-1{{beta}} Gene J. Immunol., June 1, 2001; 166(11): 6829 - 6838. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Deng and G. Q. Daley Expression of interferon consensus sequence binding protein induces potent immunity against BCR/ABL-induced leukemia Blood, June 1, 2001; 97(11): 3491 - 3497. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Liu, Y. Wang, M. Yamakuchi, S. Isowaki, E. Nagata, Y. Kanmura, I. Kitajima, and I. Maruyama Upregulation of Toll-Like Receptor 2 Gene Expression in Macrophage Response to Peptidoglycan and High Concentration of Lipopolysaccharide Is Involved in NF-{kappa}B Activation Infect. Immun., May 1, 2001; 69(5): 2788 - 2796. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Musikacharoen, T. Matsuguchi, T. Kikuchi, and Y. Yoshikai NF-{{kappa}}B and STAT5 Play Important Roles in the Regulation of Mouse Toll-Like Receptor 2 Gene Expression J. Immunol., April 1, 2001; 166(7): 4516 - 4524. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Faure, L. Thomas, H. Xu, A. E. Medvedev, O. Equils, and M. Arditi Bacterial Lipopolysaccharide and IFN-{{gamma}} Induce Toll-Like Receptor 2 and Toll-Like Receptor 4 Expression in Human Endothelial Cells: Role of NF-{{kappa}}B Activation J. Immunol., February 1, 2001; 166(3): 2018 - 2024. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |