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PRMT1-mediated methylation regulates MLL2 stability and gene expression

ABSTRACT: The interplay between multiple transcription factors precisely regulates eukaryotic transcription. Here, we report that the protein methyltransferases, MLL2 and PRMT1, interact directly and act collectively to regulate gene expression. PRMT1 binds to the N-terminal region of MLL2, considered an intrinsically disordered region, and methylates multiple arginine residues within its RGG/RG motifs. Notably, overexpression of PRMT1 decreased poly-ubiquitylation of MLL2, whereas mutations on methylation sites in MLL2 increased MLL2 poly-ubiquitylation, suggesting that PRMT1-mediated methylation stabilizes MLL2. MLL2 and PRMT1 cooperatively stimulated the expression of a chromosomal reporter gene in a PRMT1-mediated, MLL2-methylation–dependent manner. RNA-seq analysis found that MLL2 and PRMT1 jointly regulate the expression of genes involved in cell membrane and extracellular matrix functions, and depletion of either resulted in impaired cell migration and invasion. Our study provides evidence that PRMT1-mediated MLL2 methylation regulates MLL2 protein stability and the expression of their target genes.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Jaehoon Kim

PROVIDER: PXD055018 | JPOST Repository | Mon Oct 21 00:00:00 BST 2024

REPOSITORIES: jPOST

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Project description:Cyclin-dependent kinases (CDKs) are key regulators of the cell cycle or transcription, and are emerging as promising therapeutic targets in cancer cells. The transcriptional kinase, CDK12, is especially intriguing, as it modulates transcription elongation by phosphorylating the carboxy-terminal domain (CTD) of RNA polymerase (Pol II) with subsequent effects on expression of DNA damage response (DDR) genes. Yet the exact mechanism by which CDK12 regulates this vital process remains unclear. Using the selective inhibitor of CDK12/13, THZ5317 and transient transcriptome sequencing (TT-seq), we sought to capture the dynamic transcriptional changes linked to CDK12 by analyzing the immediate, early effects on nascent RNA production after CDK12 depletion in neuroblastoma (NB) cells. CDK12-induced inhibition resulted in gene length-dependent defects in transcription elongation, leading to a disproportionate loss of 3' reads in long genes. Short transcripts, by contrast, including those of replication-dependent histone genes, underwent 3' UTR extension. The effect on long genes was accompanied by premature cleavage and polyadenylation (PCPA), followed by early termination and loss of gene expression. DDR gene transcripts were the most prominent among those terminated by PCPA. Phosphoproteomic analysis indicated that pre-mRNA processing factors, including those involved in CPA , are direct phospho-targets of CDK12 and that CDK depletion phenocopied their effects on transcription. Importantly, sensitivity to CDK12 inhibition was predicted by a lower proportion of U1 snRNP binding sites compared to polyadenylation sites (PAS) and hence a higher propensity to intronic poly(A) site selection - a characteristic observed in most DDR genes. These results support a model in which CDK12 regulates expression of DDR genes not only by regulating transcription elongation through its effects on Pol II CTD phosphorylation, but also through direct phosphorylation of pre-mRNA processing factors. These mechanistic insights may enable the development of new anti-cancer strategies targeting multiple vulnerable steps in CDK12-dependent regulation of DNA damage repair.

			Action	DRS
	Control_IP-MS_rep1_1.raw	Raw
	Control_IP-MS_rep1_2.raw	Raw
	Control_IP-MS_rep1_3.raw	Raw
	Control_IP-MS_rep2_1.raw	Raw
	Control_IP-MS_rep2_2.raw	Raw

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PRMT1-mediated methylation regulates MLL2 stability and gene expression

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