Chen Y, Chafin D, Price D H, Greenleaf A L
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
J Biol Chem. 1996 Mar 15;271(11):5993-9.
We have examined the properties of two Drosophila RNA polymerase II mutants, C4 and S1, during elongation, pyrophosphorolysis, and DmS-II-stimulated transcript cleavage. The C4 and S1 mutants contain a single amino acid substitution in the largest and second largest subunits, respectively. Compared with wild type, C4 had a lower elongation rate and was less efficient at reading through intrinsic elongation blocks. S1 had a higher elongation rate than wild type and was more efficient at reading through the same blocks. During elongation, C4 and wild type responded similarly to DmS-II and NH4+ whereas the S1 mutant was less responsive to both. Differences between the two mutants also appeared during DmS-II-mediated transcript cleavage and pyrophosphorolysis. During extended pyrophosphorolysis, S1 polymerase was fastest and C4 polymerase was slowest at generating the final pattern of shortened transcripts. S1 and wild type were equal in the rate of extended DmS-II mediated transcript cleavage, and C4 was slower. Our results suggest that the S1 mutation increases the time spent by the polymerase in elongation competent mode and that the C4 mutation may affect the movement of the polymerase.
我们研究了两种果蝇RNA聚合酶II突变体C4和S1在延伸、焦磷酸解以及DmS-II刺激的转录物切割过程中的特性。C4和S1突变体分别在最大亚基和第二大亚基中含有单个氨基酸取代。与野生型相比，C4的延伸速率较低，且在通读内在延伸障碍方面效率较低。S1的延伸速率高于野生型，并且在通读相同障碍方面效率更高。在延伸过程中，C4和野生型对DmS-II和NH4+的反应相似，而S1突变体对两者的反应较弱。在DmS-II介导的转录物切割和焦磷酸解过程中，两种突变体之间也出现了差异。在延长的焦磷酸解过程中，S1聚合酶在生成缩短转录物的最终模式方面最快，C4聚合酶最慢。S1和野生型在延长的DmS-II介导的转录物切割速率上相等，而C4较慢。我们的结果表明，S1突变增加了聚合酶处于延伸胜任模式所花费的时间，并且C4突变可能影响聚合酶的移动。
