Location of subunit-subunit contact sites on RNA polymerase II subunit 3 from the fission yeast Schizosaccharomyces pombe

Biochemistry
K YasuiA Ishihama

Abstract

RNA polymerase II from the fission yeast Schizosaccharomyces pombe consists of 10 putative subunits. Subunit 3 (Rpb3) is a homologue of prokaryotic alpha subunit, which plays a key role in the assembly of core enzyme subunits. Previously we indicated that Rpb3 also plays an essential role in subunit assembly because it interacts with at least four subunits, two large subunits (Rpb1 and Rpb2) and two medium-sized subunits (Rpb3 and Rpb5) (1), and it constitutes a core subassembly consisting of Rpb2, Rpb3, and Rpb11 (2). Using a synthetic mixture of equimolar amounts of individual subunits, which were all purified from cDNA-expressed Escherichia coli, we found here that Rpb3 also interacts with Rpb11, another alpha homologue. By making a set of Rpb3 deletion derivatives, we carried out mapping of the Rpb5- and Rpb11-contact sites on Rpb3. By far-Western blot and GST pull-down assays, we found that the amino acid sequence between residues 105-263 of Rpb3 is involved in binding Rpb5, and the sequence between residues 105-297 is required for binding Rpb11. Although the Rpb5- and Rpb11-contact sites on Rpb3 overlap each other, both subunits are able to associate with Rpb3 simultaneously. The binding of Rpb5 stabilizes the Rpb3-Rpb11 ...Continue Reading

References

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Mar 1, 1996·The Journal of Biological Chemistry·T UlmasovT J Guilfoyle
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Citations

Sep 28, 2002·Trends in Genetics : TIG·Aled M EdwardsMark Gerstein
Mar 22, 2000·Proceedings of the National Academy of Sciences of the United States of America·B P SchlegelJ D Parvin
Jan 29, 2000·Nucleic Acids Research·M Kimura, A Ishihama
Jun 29, 1999·Cold Spring Harbor Symposia on Quantitative Biology·N A Woychik
Dec 19, 2001·BMC Molecular Biology·S GrandemangeM Vigneron
Sep 19, 2002·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Nicoletta CorbiClaudio Passananti

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