Human SWI/SNF-Associated PRMT5 Methylates Histone H3 Arginine 8 and Negatively Regulates Expression of ST7 and NM23 Tumor Suppressor Genes

Abstract

Protein arginine methyltransferases (PRMTs) have been implicated in transcriptional activation and repression, but their role in controlling cell growth and proliferation remains obscure. We have recently shown that PRMT5 can interact with flag-tagged BRG1-and hBRM-based hSWI/SNF chromatin remodelers and that both complexes can specifically methylate histones H3 and H4. Here we report that PRMT5 can be found in association with endogenous hSWI/SNF complexes, which can methylate H3 and H4 N-terminal tails, and show that H3 arginine 8 and H4 arginine 3 are preferred sites of methylation by recombinant and hSWI/SNFassociated PRMT5. To elucidate the role played by PRMT5 in gene regulation, we have established a PRMT5 antisense cell line and determined by microarray analysis that more genes are derepressed when PRMT5 levels are reduced. Among the affected genes, we show that suppressor of tumorigenicity 7 (ST7) and nonmetastatic 23 (NM23) are direct targets of PRMT5-containing BRG1 and hBRM complexes. Furthermore, we demonstrate that expression of ST7 and NM23 is reduced in a cell line that overexpresses PRMT5 and that this decrease in expression correlates with H3R8 methylation, H3K9 deacetylation, and increased transformation of NIH 3T3 cells. These findings suggest that the BRG1-and hBRM-associated PRMT5 regulates cell growth and proliferation by controlling expression of genes involved in tumor suppression.During cell growth and proliferation several genes become either repressed or activated. These variations in expression often correlate with changes in chromatin structure, which can be induced by a variety of enzymes that can disrupt nucleosomes in an ATP-dependent manner and/or covalently modify nucleosomal histones (17,29,41,58). Biochemical characterization of different members of the SWI2/SNF2 family of chromatin remodeling complexes revealed that there are complexes that can catalyze both ATP-dependent nucleosome disruption and histone deacetylation (48,49,55,59). Unlike the nucleosome remodeling and deacetylase complex, human SWI/SNF (hSWI/SNF) complexes can be purified either alone or in combination with mSin3A/histone deacetylase, indicating that there are different pools of BRG1-and hBRM-based hSWI/ SNF complexes (19,27,42). Recent work has also shown that flag-tagged BRG1 and hBRM complexes include the type II protein arginine methyltransferase 5 (PRMT5) and that these complexes are involved in transcriptional repression of the MYC/MAX/MAD target gene CAD (32). These studies and work by various groups suggest that ATP-dependent chromatin remodeling complexes can act in concert with various histone-modifying enzymes to modulate chromatin structure (10, 29). Although PRMT5 has been implicated in transcriptional repression of CYCLIN E and CAD, it is not clear whether it is involved in regulating a broader spectrum of genes and whether it has any effects on cell growth and proliferation.Histone methylation has been identified as an important modification for both transcriptional activation and transcript...

Application of machine learning methods to histone methylation ChIP-Seq data reveals H4R3me2 globally represses gene expression

“…A number of experimental studies have shown that PRMT5 and H4R3me2 s repress gene expression [15-28]. In an experiment that is a direct analogue of our knockout analysis, silencing of PRMT5 in mouse cell lines resulted in more de-repressed than repressed genes in a microarray analysis [22], supporting our result that H4R3me2 s is globally repressive. PRMT5 is a member of the multi-subunit mSin3A and NuRD histone deacetylase complexes [26], suggesting H4R3me2 is associated with deacetylation and hence gene inactivation [22,23,26].…”

ST7-mediated suppression of tumorigenicity of prostate cancer cells is characterized by remodeling of the extracellular matrix

“…Previously we have shown that ST7 mutations are absent from primary breast cancers (Thomas et al, 2001) pointing to the downregulation of ST7 via epigenetic mechanisms. In pilot experiments, we have not detected significant hypermethylation of the ST7 gene promoter in primary breast cancers (unpublished observations) and it is likely that histone acetylation is the predominant mode of regulation of ST7 (Pal et al, 2004).…”

“…Interestingly, recent evidence suggests that histone methylation may be an important epigenetic modifier of ST7 transcription. The protein arginine methyltransferase 5 (PRMT5) can associate with the BRG1 chromatin remodeler to directly repress transcriptional activation of ST7 through regulating chromatin accessibility (Pal et al, 2004). Furthermore, exogenous expression of PRMT5 in NIH 3T3 cells induced cell transformation that was associated with a decrease in ST7 expression.…”

“…However, the rarity of somatic mutations in ST7 among primary cancers or cancer cell lines suggests that epigenetic alterations of ST7 mediated by histone acetylation (Pal et al, 2004) may be the primary mechanism of inactivation. Microarray studies imply that ST7 induces remodeling of the extracellular matrix indicating that ST7 mediates tumor suppression by changing the tumor microenvironment, which is now recognized as an important factor in determining the malignant potential of cancer cells.…”

Mechanisms of ATP dependent chromatin remodeling

“…In Drosophila the two forms of SWI/SNF called BAP (Brahma associated proteins) and PBAP (Polybromo-associated BAP) both contain the same catalytic subunit (Brahma), but are distinguished by BAP containing the OSA subunit and PBAF containing the Polybromo and BAP170 subunits [10]. Although human SWI/SNF can be characterized as being of two forms, namely BAF (BRG1/hBRM-Associated Factors) and PBAF (Polybromo-associated BAF), there are many forms of human SWI/SNF that acquire tissue-specific subunits [11] or additional sub-complexes in which the SWI/SNF-type remodelers are associated with other factors such as BRCA1 [12,13], components of the histone deacetylase Sin3 complex [14] and histone methylases [15,16]. Recently, Rtt102p was identified as the newest subunit of both SWI/SNF and RSC complexes by MudPIT or mass spectrometry analysis [17].…”