Level of the RNA polymerase II in the fission yeast stays constant but phosphorylation of its carboxyl terminal domain varies depending on the phase and rate of cell growth.
BACKGROUND
The RNA polymerase II of the fission yeast Schizosaccharomyces pombe consists of 12 Rpb subunits, of which four (Rpb1, Rpb2, Rpb3 and Rpb11) form the assembly and catalytic core and five (Rpb5, Rpb6, Rpb8, Rpb10 and Rpb12) are shared among RNA polymerases I, II and III. The intracellular levels of three RNA polymerase forms should be interrelated, but the control of RNA polymerase formation remains mostly unknown.


RESULTS
To reveal the physiological role and the synthesis control of each Rpb subunit, the intracellular levels of the Rpb proteins were examined in S. pombe growing at various phases under various conditions. Results indicate that the intracellular concentrations of the Rpb proteins stay constant at levels characteristic of the rate and phase of cell growth, and the relative level between the 12 subunits also remains constant, together implying that the intracellular concentration of RNA polymerase II stays constant, as in the case of prokaryotes. As an attempt to gain insights into the activity control of RNA polymerase II, we also analysed the phosphorylation level of the carboxyl-terminal domain (CTD) of the largest subunit Rpb1. Phosphorylated forms of Tyr1 and Thr4 within 29 repeats of the YSPTSPS heptapeptide were detected in both slow-migrating IIo and fast-migrating IIa forms of Rpb1 on SDS-PAGE (polyacrylamide gel electrophoresis). However, phosphorylated Ser2 and Ser5 were identified only in the IIo form, indicating that Ser phosphorylation contributes to the conformational change in CTD. The phosphorylation levels of Ser, Thr and Tyr all vary depending on the cell culture conditions.


CONCLUSIONS
The intracellular level of RNA polymerase II stays constant, but the amount engaged in transcription cycle varies depending on the culture conditions, as estimated from the sites and levels of phosphorylation of Rpb1 CTD.
Sakurai
Hitomi
H
Department of Molecular Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan.
Ishihama
Akira
A
D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D012319:RNA Polymerase II / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D013696:Temperature
D011494:Protein Kinases; C060137:carboxy-terminal domain kinase; D012319:RNA Polymerase II