Poster abstracts
Poster number 132 submitted by Courtney Szyjka
Suppression screen for the identification of protein interactors of the trp RNA-binding attenuation protein (TRAP)
Courtney E. Szyjka (Department of Biological Sciences, University at Buffalo), Natalie McAdams (Department of Microbiology and Immunology, University at Buffalo), Paul Gollnick (Department of Biological Sciences, University at Buffalo)
Abstract:
Transcriptional regulation of the tryptophan (trp) biosynthetic operon in Bacillus subtilis is controlled by the trp RNA-binding attenuator protein (TRAP). Regulation of this operon was initially described as involving two competing RNA structures present in the leader region upstream of the first gene in the operon. Formation of these structures, designated as the anti-terminator and terminator, is mutually exclusive due to shared bases. In the presence of excess tryptophan, TRAP binds to 11 (G/U)AG repeats in the trp leader region RNA and prevents anti-terminator formation, allowing formation of the terminator and thus the trp genes are not transcribed. In limiting tryptophan conditions, TRAP does not bind and the trp genes are transcribed and translated to produce the tryptophan biosynthesis enzymes. Recent work has shown that, in vivo, the terminator structure isn’t required for transcription attenuation in the presence of tryptophan, indicating that TRAP may have a more direct role in transcription attenuation. Through a genetic selection, a TRAP mutant (E60K) was isolated that is capable of binding RNA and tryptophan at wild-type (WT) levels, but is deficient in transcription attenuation. We have designed a suppression screen to find protein interactors of TRAP that are necessary for transcription attenuation at the trp operon. By controlling the expression of a toxic protein (MazF) with the trp leader region, we seek to identify Bacillus subtilis proteins that will enhance transcription attenuation of the E60K TRAP mutant in vivo.
Keywords: Transcription, Gene regulation, Termination