PRMT5 functionally associates with EZH2 to promote colorectal cancer progression through epigenetically repressing CDKN2B expression

Abstract

Background: Protein arginine methyltransferase 5 (PRMT5) is a type II arginine methyltransferase that symmetrically di-methylates arginine residues on both histone and non-histone protein substrates. Accumulating evidence suggests that PRMT5 exerts its oncogenic properties in a wide spectrum of human malignancies. However, the underlying mechanisms by which PRMT5 contributes to the progression of colorectal cancer (CRC) remain to be defined. Methods: Western blot and real-time PCR were used to analyze the expression of CDKN2B. Co-immunoprecipitation (Co-IP), immunofluorescence and GST pulldown assays were employed to investigate the interaction between PRMT5 and EZH2. Luciferase reporter and chromatin immunoprecipitation (ChIP) assays were performed to validate CDKN2B as a direct target of PRMT5/EZH2. DNA methylation status at the CpG islands of promoter region of CDKN2B gene was analyzed by bisulfite sequencing. The effect of PRMT5/EZH2 on malignant phenotypes was examined through in vitro and in vivo assays. PRMT5 and EZH2 protein expression levels in CRC tissues were analyzed by immunohistochemistry (IHC) staining. Results: We observed that PRMT5-deficient CRC cells exhibit proliferation defects in vitro . PRMT5 was identified as a major transcriptional repressor of CDKN2B (p15 INK4b ) for determining CRC progression. Mechanistically, PRMT5-mediated histone marks H4R3me2s and H3R8me2s were predominantly deposited at the promoter region of CDKN2B gene in CRC cells. Knockdown of PRMT5 in CRC cells decreased the accumulation of H4R3me2s and H3R8me2s marks and reduced the CpG methylation level of CDKN2B promoter, then re-activated CDKN2B expression. Strikingly, silencing of CDKN2B partially abrogated the proliferation defects caused by PRMT5 depletion in vitro and in vivo . Furthermore, we proved that PRMT5 interacted with Enhancer of zeste homolog 2 (EZH2), leading to enhanced EZH2 binding and H3K27me3 deposition together with decreased transcriptional output of CDKN2B gene. Importantly, we found that the combined interventions exerted a synergistic inhibitory effect of combined treatment with PRMT5i (GSK591) and EZH2i (GSK126) on the growth of CRC cells/xenografts in vitro and in vivo . Moreover, PRMT5 and EZH2 were found to be significantly elevated and associated with poor prognosis in CRC patients. Conclusion: PRMT5 functionally associates with EZH2 to promote CRC progression through epigenetically repressing CDKN2B expression. Thus, our findings raise the possibility that combinational intervention of PRMT5 and EZH2 may be a promising strategy for CRC therapy.

Research progress on PRMTs involved in epigenetic modification and tumour signalling pathway regulation (Review)

“…For instance, SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4, the ATPase subunit of the switch/sucrose nonfermentable (SWI/SNF) chromatin remodelling complex, acts as a binder for PRMT1 to methylate H4R3me2a and upregulate epidermal growth factor receptor (EGFR) in colorectal cancer (35). PRMT5-mediated H4R3me2s have been found on promoters of tumour suppressor and cyclin-dependent kinase (CDK) suppressor genes, which silence cancer-cell proliferation (36,37). Menin (MEN1) has an essential role in both repressing and activating gene expression.…”

The PRMT5-LSD1 axis confers Slug dual transcriptional activities and promotes breast cancer progression

“…It is conceivable that this selectively transcription initiation or repression on target genes by PRMT5 and LSD1 may depend on various molecular partners interacted with them. PRMT5 participates in transcriptional repression by mediating the H4R3me2s when it interacts with several transcriptional corepressors including NuRD, EZH2 or DNMT3A [ 23 , 35 , 36 ]. In contrast, when PRMT5 interacts with Sp1 or OXR1A, it stimulates transcription of target genes by mediating H3R2me2s [ 29 , 37 ].…”

Identification of hub genes and key pathways in pediatric Crohn’s disease using next generation sequencing and bioinformatics analysis

“…CDKN2B [772], hsa-mir-146a [768] and ESR1 [773] have been thought of as a specific and exclusive biomarkers for ulcerative colitis. CDKN2B [774], hsa-mir-25-5p [775], hsa-mir-146a [768], hsa-mir-148a-3p [776], NFYA (Nuclear Transcription Factor Y Subunit Alpha) [777], FOXA1 [778], ESR1 [779], ARID3A [780], PRDM1 [781], GATA2 [782], HOXA5 [783] and BRCA1 [784] were significantly associated in colorectal cancer. Theses colorectal cancer biomarkers might plays important regulatory roles in CD.…”