Electronic Thesis and Dissertation Repository

Degree

Doctor of Philosophy

Program

Biochemistry

Supervisor

Dr. Joseph Torchia

Abstract

Arginine methylation is a prevalent post-translational modification that is found on many nuclear and cytoplasmic proteins, and has been implicated in the regulation of gene expression. CARM1 is a member of the protein arginine methyltransferase (PRMT) family of proteins, and is a key protein responsible for arginine methylation of a subset of proteins involved in transcription. In this thesis I examine some of the mechanisms through which CARM1 contributes to global transcriptional regulation.

Using a ChIP-DSL approach, we show that the p/CIP/CARM1 complex is recruited to 204 proximal promoters following 17β-estradiol (E2) treatment in MCF-7 cells. Many of the target genes have been previously implicated in signaling pathways related to oncogenesis. JAK2, a member of the Jak/Stat signaling cascade, is one of the direct E2-dependent targets of the p/CIP/CARM1 complex. Following E2-treatment, histone modifications at the JAK2 promoter are reflective of a transcriptionally permissive gene, and we observed modest increases in RNA and protein expression. Notably, E2-induced expression of Jak2 was diminished when p/CIP or CARM1 were depleted, suggesting that the p/CIP/CARM1 complex is required for the observed transcriptional response. Collectively, these results suggest that E2-dependent recruitment of the p/CIP/CARM1 complex causes JAK2 to become ‘poised’ for transcription, a finding that may be extendable to other target genes and signalling pathways. Furthermore, bioinformatic examination of p/CIP/CARM1 target promoters suggests that transcription factor crosstalk is the favored mechanism of E2-dependent p/CIP/CARM1 complex recruitment.

Using ChIP-Seq, we identified genomic regions to which CARM1 is recruited. Subsequent characterization of binding events suggest a role for CARM1 in transcriptional elongation, and implicate the transcription factor PAX1 as a novel mechanism through which CARM1 can be recruited to the genome. Identification of CARM1-dependent differentially expressed genes revealed that direct recruitment of CARM1 is not essential for the majority of its transcriptional effects in MEFs. However, CARM1 does play a critical role in cellular growth and proliferation, and in the absence of CARM1, the expression of many cell cycle regulators is dramatically affected.

Collectively, this work provides insight into some of the mechanisms through which CARM1 modulates transcription, and highlights its importance in diverse cellular processes.

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