Poster Sessions

TRANS-7

Jordan Irvin

J. D. Irvin, J. N. Strathern

Fidelity of Transcriptional Elongation: Slippage on Homopolymeric Runs in vivo.

Transcriptional slippage is movement of mRNA relative to the DNA template. Slippage will create mRNA that is out of register with the DNA template, and could generate aberrant proteins with deleterious biological effects. We have constructed plasmid templates to assay slippage in E. coli and S. cerevisiae. The wild-type bacterial polymerase slips efficiently on homopolymeric runs of 10 nucleotides and favors slippage on tracts of As and Ts over Gs and Cs. Single nucleotide interruptions (i.e. A5GA5) abolish the slippage events. In S. cerevisiae, all protein-encoding genes are transcribed by RNA polymerase II (pol II), which is composed of 12 subunits. Rpb1 and Rpb2 form a channel for the template strand of DNA, the active site of NTP incorporation, and the nine nucleotide RNA:DNA hybrid. We randomly mutagenized RPB1 and isolated a class of mutants with defects in transcriptional fidelity. To our knowledge, these are the only reports of in-vivo slippage in yeast and the only RNA pol II mutations that increase slippage. With these mutants and at-risk reporter constructs, we will be screening for transcription factors that function to prevent transcriptional slippage and genes whose expression may be susceptible to transcriptional slippage.