| Friend of GATA (FOG)-2 is a transcriptional co-repressor critical for cardiac development. Mice deficient in FOG-2 die at embryonic day (E)13.5 of congestive heart failure due to severe cardiac malformations. FOG-2 expression is first observed in the developing heart and the pro-epicardium at E8.5. By E16.5, FOG-2 is predominantly expressed in the brain, heart, and gonads. Despite FOG-2's critical role in cardiac development, little is known about the early cardiac-restricted regulation of the FOG-2 gene.; In this thesis, I first describe an alternative transcript produced from the FOG-2 gene, FOG-2S. The FOG-2S message encodes a 1019 amino acid protein, which is identical to amino acids 133 thru 1151 of FOG-2. This protein can bind to GATA4 as demonstrated by in vitro binding assays, but is unable to repress GATA4 activity because it lacks the N-terminal FOG repression motif. RT-PCR experiments show that FOG-2S is expressed predominantly in embryonic and adult heart. Using a combination of 5' RACE and RT-PCR, I was able to identify the transcriptional start site for the FOG-2 and FOG-2S transcripts and identify two alterative promoters, P1 and P2, respectively. Lastly, I analyzed the P1 minimal cardiac promoter by promoter deletion analysis. I found that this promoter consists of the first 550bps 5' of the transcriptional start site and an enhancer element that is located between +250 and +270. The minimal cardiac promoter contains an NFAT site that when mutated causes a reduction of FOG-2 promoter activity in neonatal rat cardiocytes. The enhancer element located in the 5' UTR region of the FOG-2 gene contains two CACCC-boxes that have been known to bind the SP1/KLF family of transcription factors. These sites when mutated abolish FOG-2 promoter activity in neonatal rat cardiocytes and also attenuate GATA4-mediated transactivation of the FOG-2 minimal promoter in NIH 3T3 fibroblasts. Interestingly, Ets-1 and GATA4 are able to synergistically activate the FOG-2 P1 minimal cardiac promoter. This work identifies some of the upstream regulators of the FOG-2 gene and suggests a role of FOG-2 as a transcriptional modulator of the genetic network of cardiac development. |
