The etiology of congenital heart disease (CHD), one of the most common congenital malformations, remains elusive. Prior to the work described in this proposal epidemiological studies and case reports pointed to a genetic etiology, but no specific human chromosomal locus had been linked to specific forms of CHD. Recent studies have demonstrated that patients with the DiGeorge and Velo-Cardio-Facial syndromes, who have a preponderance of conotruncal cardiac defects, have microdeletions of 22q11. Preliminary work completed by the candidate demonstrates that a significant proportion of non-syndromic patients with conotruncal defects have 22q11 microdeletions as well. We hypothesize that the deleted region contains a gene(s) relevant to cardiac morphogenesis.
The aims of this proposal build upon the candidate's initial work: definition of a cardiac critical region, isolation of genes from this region, and investigation of the genes' role in cardiac morphogenesis in a mammalian model, the developing mouse embryo. To define a cardiac critical region, additional patients will be screened for 22q11 microdeletions and the boundaries of their deletions determined using fluorescence in situ hybridization of metaphase spreads and Southern blot dosage analysis. Although the sponsor's laboratory is utilizing several techniques to identify genes from this region, this proposal will focus on exon prediction to identify possible coding regions in DNA sequence. Transcripts will be detected by reverse transcriptase polymerase chain reactions (RT-PCR). If authentic message is detected, libraries will be screened to isolate corresponding cDNA clones. These clones will then be used to study gene expression during early mouse morphogenesis to identify candidate genes relevant to cardiac development.