Subunit interactions and protein-DNA interactions of the Drosophila melanogaster small nuclear RNA activating protein complex

Small nuclear RNAs (snRNAs) are important in eukaryotic organisms because they are an integral part of the spliceosome, an RNA-protein complex that removes introns from the primary transcript of messenger RNAs. The focus of the work in our lab is to understand the transcriptional regulation of the genes that code for the snRNAs. In particular, we are interested in the molecular mechanisms and underlying structural differences that contribute to RNA polymerase specificity at the promoters of small nuclear RNA genes transcribed by RNA polymerase II (Pol II) or RNA polymerase III (Pol III) in Drosophila melanogaster. Transcription of genes coding for the small nuclear RNAs depends upon a unique transcription factor known as the small nuclear RNA activating protein complex SNAPc. In D. melanogaster, DmSNAPc consists of three distinct subunits (DmSNAP190, DmSNAP50, and DmSNAP43) so named to correspond with the three homologous human SNAPc subunits. Chapter 1 of this dissertation describes the identification of domains within each subunit that are 1) involved in complex formation with the other two subunits or 2) required for the DNA binding activity of the complex. Chapter 2 describes work I performed in an attempt to mutate live D. melanogaster flies to induce altered RNA polymerase specificity in vivo at a hybrid U1 snRNA promoter transgene inserted into the fly genome. In Chapter 3, I describe an attempt to alter RNA polymerase specificity in vitro using a small, synthetic polyamide that binds in a sequence-specific manner to various versions of the U1 promoter that contain a recognition site for the polyamid