The Rpb6 subunit of fission yeast RNA polymerase II is a contact target ofthe transcription elongation factor TFIIS

The Rpb6 subunit of RNA polymerase II is one of the five subunits common to three forms of eukaryotic RNA polymerase. Deletion and truncation analyses of the rpb6 gene in the fission yeast Schizosaccharomyces pombe indicated that Rpb6, consisting of 142 amino acid residues, is an essential protein f or cell viability, and the essential region is located in the C-terminal ha lf between residues 61 and 139. After random mutagenesis, a total of 14 tem perature-sensitive mutants were isolated, each carrying a single (or double in three cases and triple in one) mutation. Four mutants each carrying a s ingle mutation in the essential region mere sensitive to 6-azauracil (6AU), which inhibits transcription elongation by depleting the intracellular poo l of GTP and UTP. Both 6AU sensitivity and temperature-sensitive phenotypes of these rpb6 mutants were suppressed by overexpression of TFIIS, a transc ription elongation factor. In agreement with the genetic studies, the mutan t RNA polymerases containing the mutant Rpb6 subunits showed reduced affini ty for TFIIS, as measured by a pull-down assay of TFIIS-RNA polymerase II c omplexes using a fusion form of TFIIS with glutathione S-transferase. Moreo ver, the direct interaction between TFIIS and RNA polymerase II was compete d by the addition of Rpb6. Taken together, the results lead us to propose t hat Rpb6 plays a role in the interaction between RNA polymerase II and the transcription elongation factor TFIIS.