Arnold G F, Tessman I
Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907.
J Bacteriol. 1988 Sep;170(9):4266-71. doi: 10.1128/jb.170.9.4266-4271.1988.
The highly defective rho-15 mutant of Escherichia coli produces plasmid DNA that is 22% less negatively supercoiled than DNA from an isogenic wild-type strain (J. S. Fassler, G. F. Arnold, and I. Tessman, Mol. Gen. Genet. 204:424-429, 1986). We extended our measurements of plasmid superhelicity to additional rho mutants and to strains containing mutations that suppress rho transcription termination defects; the suppressor mutations were in the rpoB and the rho genes. The superhelicity of plasmid DNA was reduced by 11 and 10%, respectively, in the rho-702 and rho-201 mutants, both of which are less defective in Rho-mediated transcription termination than rho-15. Plasmid superhelicity was restored in all the suppressed rho mutants; in one rpoB mutant, plasmid DNA was even more negatively supercoiled than in rpoB+ cells, whether in a rho+ or rho mutant background. Suppression of rho mutants enabled them to maintain plasmids that could not be maintained in the mutants in the absence of the suppressor mutations. The results indicate that in addition to DNA gyrase, topoisomerase I, and Rho, RNA polymerase is also a determinant of DNA superhelicity, and its effect is modified by the Rho protein. We propose that Rho may increase the degree of DNA unwinding by the transcription complex, possibly at transcription termination sites.
大肠杆菌高度缺陷型rho - 15突变体产生的质粒DNA，其负超螺旋程度比同基因野生型菌株的DNA低22%（J. S. Fassler、G. F. Arnold和I. Tessman，《分子与普通遗传学》204:424 - 429，1986年）。我们将质粒超螺旋的测量扩展到其他rho突变体以及含有抑制rho转录终止缺陷突变的菌株；抑制突变存在于rpoB和rho基因中。在rho - 702和rho - 201突变体中，质粒DNA的超螺旋程度分别降低了11%和10%，这两个突变体在Rho介导的转录终止方面的缺陷都比rho - 15小。在所有被抑制的rho突变体中，质粒超螺旋得以恢复；在一个rpoB突变体中，无论处于rho +还是rho突变背景下，质粒DNA的负超螺旋程度甚至比rpoB +细胞中的还要高。rho突变体的抑制使它们能够维持在没有抑制突变时无法在突变体中维持的质粒。结果表明，除了DNA促旋酶、拓扑异构酶I和Rho外，RNA聚合酶也是DNA超螺旋的一个决定因素，并且其作用会被Rho蛋白修饰。我们提出，Rho可能会增加转录复合物使DNA解旋的程度，可能是在转录终止位点。
