To establish the potential for the nuclear factor, Restrictive Element Silencing Transcription factor (REST), to negatively regulate CRH gene expression via the Restrictive Element 1 (RE-1 element) located in the first intron of the CRH gene. A) Demonstrate specific REST binding to the CRH RE-1 sequence in vitro, by Electrophoretic Mobility Shift Assay (EMSA) to establish the possibility for RE-1 mediated CRH transcriptional regulation by REST. B) Show by in vitro transcriptional assays that RE-1 can mediate CRH transcriptional repression by REST. II. To test the hypothesis that the RE-1 element mediates important in vivo transcriptional regulation of the CRH gene by mouse knock-in of a version of the gene lacking the RE-1.
