GENETIC MODIFIERS OF TGF BETA 1 ACTION IN VIVO

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Inactivation of the transforming growth factor beta1 (TGFbeta1) gene causes defects in angiogenesis and haematopoiesis that lead to prenatal death. The penetrance of this phenotype depends on genetic background. A major genetic modifier, tgfmod1, contributes over 50% of the genetic in control of the penetrance of TGFbeta1-/-embryo lethality. The overall goal of this project is to identify tgfmod1. A combined genetic and positional cloning approach will be taken. The identification of tgfmod1 will make a significant contribution to our understanding of embryonic angiogenesis and hematopoiesis in vivo, as well as that of TGFbeta1 regulation of these processes. Moreover, it will test the general approach of cloning genetic modifiers of embryo-lethal phenotypes, many of which have been found during generation of homozygous gene knock out mice. TGFbeta1 is implicated in many human diseases, including hereditary haemorrhagic telangiectasia (HHT), cancer, pathological angiogenesis, atherosclerosis, inflammation, osteoporosis, fibrosis, wound healing and glomerulonephritis. However, the severity of these disease phenotypes, including the single gene disorder, HHT, is very variable, suggesting the existence of modifying gene traits. Identification of genes that can modify the incidence and severity of diseases caused by mis-regulation of TGFbeta is therefore of value to general medicine. tgfmod1 will be definitively mapped to approximately 0.5cM, using our panel of reciprocal congenic mice, together with a proven functional genetic test cross. A BAC contig will be generated that spans this critical map region, and comparative genomic sequencing, together with a large selection of DNA sequence analysis software tools, will be used to identify every gene within the BAC contig. Candidate genes will be selected on the basis of their fine map location and gene expression pattern. Functional polymorphisms within candidate genes will be searched for by various molecular polymorphism detection techniques, DNA sequencing and RNA and protein analysis. Identity of tgfmod1 will be confirmed by genotype/phenotype correlation in several mouse strains, and by functional analysis, including transgenesis.