Abstract 216: Bcl11b-Mediated Regulation of Vascular Smooth Muscle Specific Gene Expression

Background: A recent genome wide association study has shown a link between increased risk of arterial stiffness and SNPs in an enhancer approximately 850 kb downstream of Bcl11b, a zinc finger transcription factor known to regulate neuronal and T cell development. We were the first to show that Bcl11b is present in vascular smooth muscle (VSM), VSM specific Bcl11b knockouts mice (BSMKO) have increased arterial stiffness, measured in vivo by pulse wave velocity, and BSMKO aortas have decreased expression of smooth muscle myosin and α-smooth muscle actin. Very little is known about the molecular mechanisms through which Bcl11b can affect gene expression in VSM. Aims: CArG cassettes at VSM gene promoters are known to be a major site of gene activation through both genetic and epigenetic means. As Bcl11b has been shown to interact with CG rich sequences, we investigated whether Bcl11b binds to CArG sites and the methylation/acetylation state of surrounding histones of VSM specific genes ( Myh11, Acta2, Sm22 ) in aortas from wild type (WT), and BSMKO mice. Methods: Aortas and thymuses from WT (n=4) and BSMKO (n=4) were collected. ChIP analysis on DNA fragments was then used to target residues on histones 3 and 4 for epigenetic marks associated with active gene expression (H3K4me2, H3k9Ac and H4K16ac) and Bcl11b binding at CArG sites of Myh11, Acta2 and Sm22, as well as the promoter of Interleukin-2 ( Il2 ), a known Bcl11b target. Ongoing studies are aimed at performing ChIP analysis in aortas from aged and high fat, high sucrose-fed mice, two models of arterial stiffness to study the role of Bcl11b-dependent VSM gene expression regulation in pathological conditions. Results: Preliminary results indicate that Bcl11b binds to the CArG sites in the aorta, a 2.5 fold enrichment compared to IgG control. Methylation of H3K4 was approximately 3 fold more enriched in aortic tissue compared to the thymus while acetylation was not significantly different between the two tissues. Conslusions: Our data indicate that Bcl11b regulates the expression of specific VSM genes by binding to the CArG regions and promoting methylation at the H3K4 residue. By understanding the roles and targets of Bcl11b we can further deduce target pathways for therapeutics to preventatively treat arterial stiffness.