The Transcriptional Corepressor, PELP1, Recruits HDAC2 and Masks Histones Using Two Separate Domains

PELP1 (proline-, glutamic acid-, and leucine-rich protein 1) has been recognized as a coactivator of estrogen receptor (ER)-recruiting p300/CREB-binding protein histone acetyltransferase to the target chromosome. The present study shows that PELP1 does indeed coactivate ER-mediated transcription but also serves as a corepressor of other nuclear hormone receptors (NR)- and non-NR sequence-specific transcription factors tested, including GR, Nur77, AP1, NF-κB, and TCF/SRF. PELP1 expression also retarded the proliferation of mouse fibroblast cell lines in which there was no detectable ER. This was due, at least in part, to the suppressed activation of serum-response genes such as c-fos that in turn resulted from the blocked histone hyperacetylation of nucleosomes containing the c-fos promoter region. The N-terminal leucine-abundant region of PELP1 was observed to interact with HDAC2 and exhibited repressive activity when tethered to the chromatin. In addition, the C-terminal glutamic acid-abundant region bound to the hypoacetylated histones H3 and H4 and prevented them from becoming substrates of histone acetyltransferase. Thus PELP1 promotes and maintains the hypoacetylated state of histones at the target genomic site, and ER binding reverses its role to hyperacetylate histones through an as yet unidentified mechanism. PELP1 (proline-, glutamic acid-, and leucine-rich protein 1) has been recognized as a coactivator of estrogen receptor (ER)-recruiting p300/CREB-binding protein histone acetyltransferase to the target chromosome. The present study shows that PELP1 does indeed coactivate ER-mediated transcription but also serves as a corepressor of other nuclear hormone receptors (NR)- and non-NR sequence-specific transcription factors tested, including GR, Nur77, AP1, NF-κB, and TCF/SRF. PELP1 expression also retarded the proliferation of mouse fibroblast cell lines in which there was no detectable ER. This was due, at least in part, to the suppressed activation of serum-response genes such as c-fos that in turn resulted from the blocked histone hyperacetylation of nucleosomes containing the c-fos promoter region. The N-terminal leucine-abundant region of PELP1 was observed to interact with HDAC2 and exhibited repressive activity when tethered to the chromatin. In addition, the C-terminal glutamic acid-abundant region bound to the hypoacetylated histones H3 and H4 and prevented them from becoming substrates of histone acetyltransferase. Thus PELP1 promotes and maintains the hypoacetylated state of histones at the target genomic site, and ER binding reverses its role to hyperacetylate histones through an as yet unidentified mechanism. Eukaryotic chromatin is a highly compact DNA-protein complex composed of an array of nucleosomes. The histone octamer, containing two copies each of histones H2A, H2B, H3, and H4, provides the protein core of a nucleosome into which the DNA is packaged (1Luger K. Mader A. Richmond R. Sargent D. Richmond T. Nature. 1997; 389: 251-260Crossref PubMed Scopus (7141) Google Scholar). The modulation of chromatin compactness, i.e. chromatin remodeling, is important for rendering transcription factors and the basic transcription machinery accessible to their target gene promoters. Chromatin remodeling can be achieved by at least two enzyme complexes, the histone-modifying enzyme complexes and the ATP-dependent remodeling complexes (2Cress W.D. Seto E. J. Cell. Physiol. 2000; 184: 1-16Crossref PubMed Scopus (582) Google Scholar, 3Gray S.G. Ekstrom T.J. Exp. Cell Res. 2001; 262: 75-83Crossref PubMed Scopus (504) Google Scholar, 4Horn P.J. Peterson C.L. Science. 2002; 297: 1824-1827Crossref PubMed Scopus (334) Google Scholar, 5Marmorstein R. Roth S.Y. Curr. Opin. Genet. Dev. 2001; 11: 155-161Crossref PubMed Scopus (326) Google Scholar, 6Tyler J.K. Eur. J. Biochem. 2002; 269: 2268-2274Crossref PubMed Scopus (124) Google Scholar). Hyperacetylation of the lysine residues contained in histones is believed to be a mechanism for the destabilization of nucleosomes and the facilitation of transcription factor binding (7Hebbes T.R. Thorne A.W. Crane-Robinson C. EMBO J. 1988; 7: 1395-1402Crossref PubMed Scopus (744) Google Scholar, 8Narlikar G.J. Fan H.-Y. Kingston R.E. Cell. 2002; 108: 475-487Abstract Full Text Full Text PDF PubMed Scopus (1260) Google Scholar). The steady state acetylation level of the core histones is maintained by a balance between the opposing activities of HATs 1The abbreviations used are: HAT, histone acetyltransferase; PELP1, proline-, glutamic acid-, and leucine-rich protein 1; LAR, leucine-abundant region; PAR, proline-abundant region; GAR, glutamic acid-abundant region; HDAC, histone deacetylase; NR, nuclear hormone receptor; ER, estrogen receptor; AP1, activating protein 1; TCF/SRF, ternary complex factor/serum response factor; NF-κB, nuclear factor-kappa B; ChIP, chromatin immunoprecipitation; INHAT, inhibitor of acetyltransferase; nt, nucleotide; CBP, CREB-binding protein; SRE, serum-response element; GST, glutathione S-transferase; Pan, pantothenate; TSA, trichostatin A; MMTV, mouse mammary tumor virus; SRF, serum-response factor; E2, 17β-estradiol; GR, glucocorticoid receptor; pRb, retinoblastoma protein; CaMK IV, calmodulin-dependent kinase IV; ERE, ER-responsive element; RT, reverse transcription; DB, DNA binding. and HDACs (8Narlikar G.J. Fan H.-Y. Kingston R.E. Cell. 2002; 108: 475-487Abstract Full Text Full Text PDF PubMed Scopus (1260) Google Scholar). Many transcriptional coactivators, including GCN5/PCAF, CBP/p300, and NcoA, exhibit intrinsic HAT activity (5Marmorstein R. Roth S.Y. Curr. Opin. Genet. Dev. 2001; 11: 155-161Crossref PubMed Scopus (326) Google Scholar), whereas most transcriptional corepressors studied thus far recruit specific HDACs to the chromatin of target genes (5Marmorstein R. Roth S.Y. Curr. Opin. Genet. Dev. 2001; 11: 155-161Crossref PubMed Scopus (326) Google Scholar, 9Xu L. Glass C.K. Rosenfeld M.G. Curr. Opin. Genet. Dev. 1999; 9: 140-147Crossref PubMed Scopus (820) Google Scholar). For example, Sin3, Mi-2/NuRD, and retinoblastoma protein (pRb) associate with HDAC1 and -2, CtBP with class II HDACs, and NcoR and SMRT with HDAC3 (10Brehm A. Miska E.A. Mccance D.J. Reid J.L. Bannister A.J. Kouzarides T. Nature. 1998; 391: 597-601Crossref PubMed Scopus (1085) Google Scholar, 11Chen G. 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Some transcriptional coregulators for NR-mediated transcription, such as ZAC1, NSD1, and RIP14, have been demonstrated to possess dual functions, acting as both coactivators and corepressors (17Cavailles S. G. S. M. EMBO J. PubMed Scopus Google Scholar, E. T. R. EMBO J. 1998; PubMed Scopus Google Scholar, Mol. Cell. 2000; PubMed Scopus Google Scholar, C. Mol. Cell. 1998; PubMed Scopus Google Scholar). the nuclear protein PELP1 (proline-, glutamic acid-, and leucine-rich protein 1) has been to be for NR-mediated transcription; i.e. a coactivator for ER and a corepressor for glucocorticoid receptor Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). PELP1 is in most but in and such as the and the mammary The of expression of PELP1 in the of the coactivator of PELP1 in the of mammary PELP1 in its N-terminal by a proline-abundant region and an region containing an of glutamic at the other of the protein Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). PELP1 does to have intrinsic HAT but does to a activity for ER binding to both and through its N-terminal which PELP1 also a role in cell with retinoblastoma protein through its N-terminal S. J. Full Text Full Text PDF PubMed Scopus Google Scholar). In to the transcriptional for PELP1, has also been that a ER and kinase through its and C. E. S. A. 2002; 99: PubMed Scopus Google Scholar). In the present that PELP1 as a corepressor of sequence-specific transcription including and non-NR transcription factors such as AP1, TCF/SRF, and transcriptional both the N-terminal leucine-abundant region and the C-terminal glutamic acid-abundant which with HDAC2 and hypoacetylated core PELP1 can be to to a class of in that histone acetylation and core histones from and of PELP1 was by of from the cell its was by DNA and was into the and of of PELP1 by and into the in the the of PELP1 for the was into the of and in to copies of The of was by and into the at the of in into the of that contained the the was into the and of The calmodulin-dependent kinase Nur77, and from J. and was from K. C. from with the and from J. was from and and from J. the kinase promoter was from and was the C-terminal PELP1 and CBP, histone H3, and histones and H3 and H4 and histones H2A, H2B, H3, and H4 from Cell and and Cell maintained in with and maintained in with and cell lines PELP1 and PELP1 the with and PELP1 by and in in the presence of with expression PELP1 and for with for and nuclear of the as J. D. Scholar). nuclear of in the presence of of in binding and of For the the nuclear with of for at and for with the and by in at a of and with of with the of expression The of was by with and of was to the DNA to the of the with with and and activities a For the and in containing with for at least and In with the for and by two of in and containing inhibitor and The cell by at for and with specific for at The complexes with by and to The with and by In with each of histone core histone complex in a binding containing and and for at and bound histones glutathione with binding containing and by in the by and by For H3 the and In of PELP1 and in in the presence of and with of H3 for with and histone by and by and was from the was with of a for an of The a of and for a of and for and a of and for of and by was to a for M. A. S. C. J. Genes Dev. 7: PubMed Scopus Google Scholar). with of for and with containing in and and by the of a and The in and and with H3, H4 and and at for to reverse the DNA in the complex by and and by two of specific for the c-fos gene as a of and for of the of SRE, and of and for of the of c-fos gene as a For the which been in containing and was in and to the as the In an for the of in of with PELP1 with to the The complexes by with H3 and by In HAT from of nuclear with of and of histone H3 and H4 in a and was with histones for the of the in HAT The was by the of and and by PELP1 a of ER but as a of has been as a coactivator of Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). The role of PELP1 by other sequence-specific transcription factors including GR, Nur77, NF-κB, AP1, and was in PELP1 at a level and exhibit ER of both ER and of PELP1 in the of ER-responsive in a as the other expression of PELP1 of the mouse mammary tumor promoter In addition, the CaMK activation of transcription was also by the expression of PELP1 PELP1 the of genes for hormone receptor transcription including NF-κB, AP1, and TCF/SRF, in a the transcription of the and genes also by PELP1, in the of expression of the gene the of the kinase promoter was in no in both the presence and the of The expression of PELP1 in the was by of with and that PELP1 as a transcriptional i.e. as a corepressor of sequence-specific transcription but as a coactivator of ER. the of ER has been in the of PELP1 as an ER coactivator and is at a in in the role of PELP1 in tumor also Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). For to be the highly PELP1 be to the activities of PELP1, in to the transcription of that is important in for in the mammary cell which both ER and PELP1, transcription by the of PELP1 the other in a to of PELP1 transcription in an cell a PELP1 level in the that ER with the role of PELP1 with to the transcriptional of in the was by be that the basic role of PELP1 is to the transcriptional activation of sequence-specific including SRE, and that the binding of ER to PELP1 PELP1 to a coactivator by an as yet mechanism. In the mechanism of PELP1 its transcriptional was as the target the and the for the of PELP1 in a that PELP1 is in the complex SRE, a nuclear of PELP1 and SRF, and and The used in the recognized PELP1 in the cell which was by with the PELP1 PELP1 does to DNA by Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar), of the DNA-protein complex by that PELP1 is to a complex with the and In addition, the of PELP1 to the the c-fos gene in was by a of used to the region of an in the region to of the c-fos and also a region in its region to as a The region to nt, but to nt, was from both and the that PELP1 to the promoter region of the c-fos gene In to the presence of PELP1 in the PELP1 to in was in PELP1 and PELP1 was in that PELP1 does indeed a complex with the is that the activity of PELP1 with to transcription can be achieved in a specific to the bound transcription The of PELP1 can be into as the N-terminal LAR, the PAR, and the C-terminal the PELP1 containing a of two of these and their nuclear expression For nuclear expression of the the nuclear of was to its The of that expression of PELP1, but the expression of of the of observed for other including the genes for GR, Nur77, NF-κB, and of the both the and of SRE, of its these that the corepressor of PELP1 can be achieved in a that of the and the transcriptional in a whereas the of the was to be at This that of the the a and that the and the of the activity of PELP1, these to be to as in and of The an to of PELP1 to DNA binding and into with a containing copies of the binding and the of the kinase promoter and a was The demonstrated that the transcription of by as of and the transcription by to the other the of GAR, PAR, both to the transcription in the of The by the and is the presence of the the by the that the the important for the to the target chromatin site, as does in In addition, the to a role in the the also to the activity of The to an of glutamic Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). the has a to PELP1 to the the GAR, an to interact with basic such as histones with the histone and and glutathione and with histone in as H3 as a of the core the other with and H4 and with the containing whereas histone histones in of and H4 with was observed when and H4 with H3 in PELP1, but was with H3 that PELP1 with H3 through the of PELP1 with H3 in with PELP1 and that H3 was with PELP1 but with but H3 with PELP1 from cell the between PELP1 and The glutamic and of has been to interact with histones and to to the activity of the complex by histone acetylation S. A. D. Cell. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). the of the in the activity of PELP1, the of PELP1 was by the of This the of SRE, in PELP1 the of PELP1 has a to the in to with which be important in PELP1 to the chromatin. of PELP1 the of c-fos and and PELP1 activities for and is that PELP1 a role in cell mouse fibroblast cell and with PELP1, and and their and The expression of PELP1 retarded the proliferation of of both fibroblast the proliferation no of was observed the other the of was with that of the the expression level of was to that of PELP1 and the transcription of and and that serum-response genes such as c-fos and be target genes of the activity of in the of PELP1, the activation of both c-fos and genes was by of with and by for the expression of both c-fos and in the was to to in the and the other of PELP1 the activation of c-fos by and the activation the retarded proliferation of and as a of PELP1 expression is due, at least to the transcription of genes for cell a that c-fos and PELP1 by histone acetylation is at the of the transcriptional activation of such as c-fos and S. A. L. Mol. Cell. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, S. EMBO J. 2000; 19: PubMed Scopus Google Scholar, A. R. Cell. 1998; 475-487Abstract Full Text Full Text PDF PubMed Scopus Google Scholar). The that PELP1 the of nucleosomes the of the c-fos gene was by specific for H3 and H4 and The two of in used in The region to nt, but the region to nt, was from both and the that H3 and H4 in the promoter region of c-fos gene and the acetylation of both H3 and H4 at the c-fos promoter region in the and cell lines and and with their and of H3 in was but in in the cell and with and and the other acetylation of H4 in the was and with and and the transcription of target including by PELP1 is due, at least in part, to chromatin remodeling by histone The of PELP1 HDAC2 to the histone acetylation by PELP1 can be achieved the of HDACs to the chromatin by the of the of HDACs is in transcriptional of the of PELP1 the of with the trichostatin the transcription of in the of with also the transcription of the gene from an inhibitor of class HDAC, to have no the activities of PELP1 that PELP1 be with class II HDACs, through the PELP1 and but HDAC2 from the of the and in the in which PELP1 was PELP1 was with HDAC2 the of a between PELP1 and the of other HDACs be HDAC2 the most both in and in the -2, and used in a of and an of the PELP1, that HDAC2 was to the c-fos promoter region but in the presence of PELP1 the can be to to the activity of PELP1 by HDAC2 to the chromatin of the target genes. The of PELP1 to and was to interact with ER binding Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). the is important in acetylation is in binding to HDAC2 the that the HAT activity the acetylation of histones was The region the HAT of was to and with of PELP1 and the gene PELP1 and resulted in the of the transcription of the to be that the to the activity of PELP1 the of histone the histone acetylation is indeed of the of by its binding to histones in histones to the HAT the other the has an to the activity of the histones in the be and from cell nuclear H3 and H4 and of into the histone acetylation the other when of and histones with the CBP, histone acetylation a of histones in of resulted in the of H3 and H4 acetylation and of resulted in the of the transcriptional activity of the gene from the In the the HAT activity of was in two from the exhibited HAT activity The expression of and in response to HAT was as has been J. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, T. G. M. C. J. C. S. S. M. B. Nature. 1997; 389: PubMed Scopus Google that histone acetylation by the is most to the of histones for the of the activities of the binding of PELP1 to H3 was by of H3 N-terminal but by H3 at lysine residues and and at a of that PELP1 to hypoacetylated H3 PELP1 in whereas H3 PELP1, but by acetylation by the other H3 to PELP1 at that the of as as its is important in the between PELP1 and histone PELP1 as a corepressor of and non-NR sequence-specific transcription the of histone acetylation in the target two the and the GAR, which recruit HDACs and histone acetylation The present study that PELP1 is a corepressor of non-NR sequence-specific transcription including NF-κB, AP1, and in to the with the of ER. coregulators of have been to including ZAC1, NSD1, and which to as coactivators as corepressors (17Cavailles S. G. S. M. EMBO J. PubMed Scopus Google Scholar, E. T. R. EMBO J. 1998; PubMed Scopus Google Scholar, Mol. Cell. 2000; PubMed Scopus Google Scholar, C. Mol. Cell. 1998; PubMed Scopus Google Scholar). the for the of these coregulators have yet to be mouse which to other coregulators including CBP, and has been to its activity to the activities of the Mol. Cell. 2000; PubMed Scopus Google Scholar). the other to its intrinsic activation and for its but is when and the other E. T. R. EMBO J. 1998; PubMed Scopus Google Scholar). PELP1 ER-mediated transcription but shows activity for of the other transcription factors thus far This was also maintained in a other cell including and and that the of PELP1 as a coactivator as a PELP1 HDAC2 through the and to and histones through the in to the activity of PELP1, ER binding both of histone and the to chromatin of PELP1 ER binding Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google the is can the of activity such the from to the transcriptional corepressors two to and the hypoacetylated state of HDACs for and binding to histones for of HAT for histones from SMRT be a two and each of which is in with HDAC3 and hypoacetylated J. T. M.G. EMBO J. PubMed Scopus Google Scholar). This to a of in that the complex histone in the of the SMRT for histones and promotes the of In addition, as in a study two the recruit corepressor in the of a Courey A.J. Res. 2000; PubMed Scopus Google Scholar). PELP1 to in a of in the of target gene of the resulted in in the activity of PELP1 whereas expression of the activity of PELP1 in a by the and the in PELP1 of HDAC2 and the histone activity of the presence of these two in as in PELP1, the target gene the does to possess HAT activity and is with to transcriptional be at least in part, by role of the GAR, i.e. of PELP1 to the target chromatin. This is by the that by when tethered to the chromatin by transcription of the gene an important as to PELP1 its target gene transcriptional corepressors thus far to the target chromatin by bound transcription including PELP1 in the LAR, which indeed has been to ER A. R. Cell. 1998; 475-487Abstract Full Text Full Text PDF PubMed Scopus Google Scholar), and as PELP1 both in and in complexes with the and the target genes of the role of PELP1 can be by bound transcription PELP1 can also be tethered to the chromatin by the GAR, through binding to hypoacetylated PELP1 also be to as a of the of the transcription of genes in a 1) be in a that PELP1 hypoacetylated histones through the GAR, in the of bound transcription and promotes the tethered complexes, in PELP1 with histones a for binding to histones of sequence-specific such as the in SMRT and other histone including and have also been demonstrated to interact with histones in a their M. K. 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Full Text Full Text PDF PubMed Scopus Google Scholar), the transcriptional of be to the of HDAC2 PELP1 the transcriptional activation of in the mouse the gene PELP1 does to in its activity for the transcriptional activation of the of PELP1 have been studied from the of ER as a coactivator of ER-mediated transcription, as a of the activity of ER Wang A. J. A. R. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, C. E. S. A. 2002; 99: PubMed Scopus Google Scholar). In the present that PELP1 is to the transcription by sequence-specific transcription including AP1, TCF/SRF, NF-κB, as as with the of ER. This activity was achieved the of and also by histone two as the present study the the activity of PELP1, to the role of be of and to PELP1 its activity and its ER coactivator J. J. and C. for genes and J. and C. for