Principal Investigator: PAUL KHAVARI
Abstract: The transcription factors HNF-1alpha and HNF-1beta are homeodomain proteins that regulate the expression of a large group of genes that have the common characteristic of being expressed in hepatocytes. These genes include those encoding coagulation factors such as fibrinogen, antithrombotic factors such as antithrombin III and protein C as well as those involved in cholesterol metabolism. The HNF-1 site plays a remarkably dominant role in regulation of many of these genes since single point-mutations in the HNF-1 site can give rise to clinical disease. Thus it seems quite likely that this protein and its binding site might serve as targets for both therapeutic approaches and pathogenic mechanism in cardiovascular disease. The control of HNF-1alpha and beta is largely developmental but is also controlled by an 11kDa protein that we have recently identified. This protein binds tightly to HNF-1alpha and acts as a transcriptional cofactor. We have named this protein transcription cofactor 1 (TCF-1), since it is to our knowledge the first transcriptional cofactor to be identified. We purified TCF-1 on the basis of its ability to bind tightly and specifically to HNF-1alpha. Peptide sequences were obtained from this purified protein and these were in turn used to isolate CDNA and genomic clones encoding TCF-1. Expression of TCF-1 in a cotransfection assay leads to coactivation of HNF-1alpha-dependent promoters. We will define the molecular basis of TCF-1 - HNF-1alpha binding by identifying the precise amino acid sequences involved from both proteins through generation of HNF- 1alpha binding by identifying the precise amino acid sequences involved from both proteins through generation of HNF-1alpha mutants and TCF-1 peptides. The functional importance of this binding will be determined in established in vitro transcription and co-transfection systems. We will also determine if TCF-1, either alone or when complexed to HNF-1alpha, binds bacterially expressed human TFIID. If there is evidence for such binding, we will define the peptide sequences involved as above and determine the functional importance of such binding by in vitro transcription assays using appropriate mutant molecules of either TFIID, HNF-1alpha or TCF-1. We will investigate the ability of TCF-1 to enhance transcriptional activation by transcription factors other than HNF-1alpha by testing a series of homeodomain and non-homeodomain containing proteins, such as Oct-1, Pit-1/GHF and others by co-transfection experiments. Finally, we will search for TCF-1homologues through genomic and cDNA screening. These studies of TCF-1 will provide insight into the molecular basis of transcriptional control in eukaryotes and may clearly define a new class of transcription factors enhancing the cooperative interaction between regulatory activating proteins and the basal transcription complex.
Funding Period: 1992-09-01 - 1997-08-31
more information: NIH RePORT