Efficient production of recombinant human transcription factor IIE.

Transcription factor IIE (TFIIE) is a general initiation and promoter escape factor for RNA polymerase II composed of p56 (TFIIE-alpha) and p34 (TFIIE-beta) subunits. Our laboratories experienced difficulty producing adequate quantities of recombinant human TFIIE-alpha for in vitro studies using available clones. We therefore re-engineered the TFIIE subunit production vectors and tested various Escherichia coli host strains to optimize expression.

Promoter escape by RNA polymerase II. Downstream promoter DNA is required during multiple steps of early transcription.

Recent evidence, obtained in a reconstituted RNA polymerase II transcription system, indicated that the promoter escape stage of transcription requires template DNA located downstream of the elongating polymerase. In the absence of downstream DNA, very early elongation complexes are unable to synthesize transcripts longer than approximately 10-14 nucleotides. In contrast, once transcripts longer than approximately 15 nucleotides have been synthesized, an extended region of downstream DNA is no longer required (Dvir, A.

Promoter escape by RNA polymerase II.

Transcription of protein-coding genes is one of the most fundamental processes that underlies all life and is a primary mechanism of biological regulation. In eukaryotic cells, transcription depends on the formation of a complex at the promoter region of the gene that minimally includes RNA polymerase II and several auxiliary proteins known as the general transcription factors. Transcription initiation follows at the promoter site given the availability of nucleoside triphosphates and ATP.