Widespread changes in gene expression accompany vascular smooth muscle cell (VSMC) phenotypic switching, a hallmark of vascular disease. Upon insult, VSMCs downregulate contractile proteins and upregulate genes linked to vascular remodelling, such as matrix metalloproteinases (MMPs) and pro-inflammatory cytokines. However, the epigenetic mechanisms which regulate VSMC phenotypic switching remain unclear. This thesis explores the role of histone 3 lysine 9 dimethylation (H3K9me2), a repressive epigenetic mark, in regulating the expression of disease-associated VSMC genes. Intriguingly, murine models of VSMC phenotypic switching revealed reduced levels of H3K9me2 upon loss of the contractile state while chromatin immunoprecipitation (ChIP) identified a subset of IL-1α/injury-responsive VSMC gene promoters enriched for H3K9me2. To test the functional importance of H3K9me2 for VSMC gene regulation the methyltransferase G9A/GLP was pharmacologically inhibited in vitro and in vivo. The resulting loss of H3K9me2 attenuated the expression of contractile VSMC markers and significantly potentiated IL-1α/injury-induced expression of MMP and pro-inflammatory genes. H3K9me2-mediated regulation of contractile and IL-1α-responsive VSMC gene expression was confirmed in cultured human VSMCs (hVSMCs). This prompted the use of hVSMCs to investigate the mechanism underlying H3K9me2-dependent regulation of IL-1α-mediated VSMC genes. Interestingly, G9A/GLP inhibition did not influence the level of IL-1α-induced nuclear localisation of the NFkB transcription factor p65 but significantly increased IL-1α-induced p65 binding to the IL6 promoter, correlating with reduced H3K9me2 levels. In contrast, enrichment of p65 was not observed at reported NFkB sites within the MMP3 promoter after IL-1α stimulation. Rather, IL-1α-induced MMP3 expression was dependent on JNK activity and G9A/GLP inhibition potentiated IL-1α-induced binding of the AP-1 transcription factor cJUN to the MMP3 promoter. Collectively, these findings suggest that H3K9me2 plays a role in maintaining the contractile VSMC state and prevents binding of both NFkB and AP-1 transcription factors at specific IL-1α-regulated genes to possibly block spurious induction of a pro-inflammatory state.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:767866 |
| Date | January 2019 |
| Creators | Harman, Jennifer |
| Contributors | Jorgensen, Helle |
| Publisher | University of Cambridge |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | https://www.repository.cam.ac.uk/handle/1810/289975 |
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