| Heart disease is the number one killer in developed countries, and of increasing importance in developing ones. Serum response factor is a critical regulator of many processes common to aging and diseased hearts. The expression of serum response factor is known to increase with advancing adult age. While a great deal is known about gene regulation by serum response factor (SRF), we are still unable to predict which genes will be up- or down-regulated by changes in SRF levels. Such understanding would be helpful in the design of therapies to mitigate the negative effects of adult cardiac aging or heart disease. The focus of this dissertation is to relate differential expression of genes in response to moderate SRF upregulation, to position-dependent colocalization of transcription factor binding sites. This dissertation met this aim through a detailed examination of the positions of CC-(A/T)6-GG (CArG), CArG-like, activator protein 1 (AP-1), and E-twenty-six (ETS) binding sites relative to one-another, and to the transcription start site. Sites in the -10Kbp promoter/enhancer regions and first two introns were mapped in genes that were up-regulated, down-regulated, and non-differentially expressed in response to moderate (∼50%) cardiac-specific over-expression of serum response factor. Classic CArG and CArG-like elements between -1.5Kbp and +2Kbp of the transcription start site were strongly associated with up-regulation. All classic CArG elements found in the first introns were in differentially expressed genes. AP-1 binding sites overlapping CArG or CArG-like elements were found only in non-differentially expressed genes. AP-1 binding sites near, but not overlapping, CArG or CArG-like elements, were strongly associated with up-regulation. ETS binding sites near CArG or CArG-like elements in up-regulated genes tended to be oriented away from the first position of the element. ETS binding sites beginning within 40bp CArG or CArG-like elements in down-regulated genes were nearly all in the reverse orientation. While it was previously shown that ETS binding sites near CArG and CArG-like elements were associated with differential expression, the novel finding in this study was the significance of the relative position and orientation of the binding sites. |
