Role of the lysine methyltransferase Set7 in smooth muscle gene expression and function

2017-03-14T04:33:46Z (GMT) by Natasha Kelly Tuano
Histone methylation is controlled by lysine methyltransferase enzymes to regulate the expression of genes implicated in cellular differentiation and lineage specificity. The lysine methyltransferase Set7 was originally described to monomethylate histone H3 on lysine 4 (H3K4me1) and more recently non-histone proteins. Set7 has been implicated in regulating key genes involved in the maintenance of pluripotency and skeletal and cardiac muscle development. While the importance of epigenetic modifiers in cardiac and skeletal muscle gene program has been described, epigenetic regulation of smooth muscle genes is poorly understood. The aims of this thesis are to identify (i) how the Set7 gene is regulated in embryonic stem cells; (ii) investigate the role of Set7 in smooth muscle gene regulation utilizing an in vitro stem cell differentiation system; and (iii) investigate the role of Set7 using a knockout mouse model. Promoter analysis of Set7 in mouse embryonic stem cells (mESCs) show bivalency with active H3K4me3 and repressive H3K27me3 modifications. The Set7 promoter is suppressed by the pluripotent transcription factors, Oct4 and Sox2. Deletion of the Oct4/Sox2 binding site activates Set7 transcription. Differentiation of mESCs is associated with reduced H3K27me3 and reduced Oct4 and Sox2 binding. Transcriptional network analysis identifies genes associated with differentiation are broadly regulated. More specifically, smooth muscle (SM)-associated gene expression is reduced in Set7 knockdown (KD) Sca1+ cells. Pharmacological inhibition of Set7 activity also reduces the expression of SM-associated genes corresponding with the results derived from Set7 KD cells. Furthermore, SM-associated gene regulation is subject to Set7 mediated methylation of H3K4me1 as well as the interaction with the Serum Response Factor (SRF) protein. Investigation of a Set7 knockout (KO) mouse model showed reduced expression of canonical SM-related genes in the aorta. Morphological abnormalities within SM layer of Set7-/- mice aorta were observed correlating with reduced vessel wall thickness and reduced SM cell number. Taken together, the results presented in this thesis suggest a novel role for Set7 in SM gene regulation, smooth muscle architecture and function.