GENETIC ANALYSIS OF RNA POLYMERASE II TRANSCRIPTIONAL REGULATION IN DROSOPHILA

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We are interested in understanding how transcription of protein-coding genes is regulated in vivo. Current models for transcriptional regulation are based largely on in vitro studies conducted under non-physiological conditions using highly purified general transcription factors, synthetic activator proteins, and non-chromatin DNA templates. By studying transcription of endogenous genes that are required for proper Drosophila melanogaster development, we hope to identify cis- and trans-acting factors that play regulatory roles in vivo and determine how the coordinated action of different sets of factors establish the broad transcriptional repertoire of eukaryotic cells. To this end, we are studying transcription of the Drosophila sevenless (sev) gene. The sev gene was chosen because minor alterations in the level or pattern of sev transcription cause easily observable phenotypes in the developing fly, making sev transcription amenable to genetic dissection. Knowledge gained from studies of sev will be tested on a genome wide basis to assess the generality of the Sev model.Sev-based genetic screens conducted in our laboratory have identified mutations in components of the RNA polymerase (pol) II preinitiation complex [TBP-associated factor (TAF)60 and TAF110 and RNA pol II subunits] as well as factors that modulate transcription through their affects on chromatin structure [Trithorax group (TRX-G) genes, RPD3, and SIN3]. Additional mutations have been identified that may also encode transcriptional regulators. Phenotypic and molecular characterization of mutant flies is being used to identify genes that are transcriptionally regulated by the encoded factors, and analysis of genetic interactions between mutant flies is being used to assess functional interactions between the encoded factors.