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首页> 外文期刊>Journal of bacteriology >RNase III-Dependent Expression of the rpsO-pnpOperon of Streptomyces coelicolor
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RNase III-Dependent Expression of the rpsO-pnpOperon of Streptomyces coelicolor

机译：腔链链霉菌rpsO-pnpOperon的RNase III依赖性表达

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We have examined the expression of the rpsO-pnpoperon in an RNase III (rnc) mutant of Streptomyces coelicolor. Western blotting demonstrated that polynucleotide phosphorylase (PNPase) levels increased in the rncmutant, JSE1880, compared with the parental strain, M145, and this observation was confirmed by polymerization assays. It was observed that rpsO-pnpmRNA levels increased in the rncmutant by 1.6- to 4-fold compared with M145. This increase was observed in exponential, transition, and stationary phases, and the levels of the readthrough transcript, initiated upstream of rpsOin the rpsO-pnpoperon; the pnptranscript, initiated in the rpsO-pnpintergenic region; and the rpsOtranscript all increased. The increased levels of these transcripts in JSE1880 reflected increased chemical half-lives for each of the three. We demonstrated further that overexpression of the rpsO-pnpoperon led to significantly higher levels of PNPase activity in JSE1880 compared to M145, reflecting the likelihood that PNPase expression is autoregulated in an RNase III-dependent manner in S. coelicolor. To explore further the increase in the level of the pnptranscript initiated in the intergenic region in JSE1880, we utilized that transcript as a substrate in assays employing purified S. coelicolorRNase III. These assays revealed the presence of hitherto-undiscovered sites of RNase III cleavage of the pnptranscript. The position of those sites was determined by primer extension, and they were shown to be situated in the loops of a stem-loop structure.
机译：我们检查了天蓝色链霉菌RNase III（ rnc ）突变体中 rpsO-pnp 操纵子的表达。 Western印迹表明，与亲本菌株M145相比， rnc 突变体JSE1880中的多核苷酸磷酸化酶（PNPase）水平增加，并且该观察结果得到了聚合分析的证实。观察到与m145相比， rnc 突变体中的 rpsO-pnp mRNA水平提高了1.6到4倍。在指数阶段，过渡阶段和固定阶段，以及在 rpsO-pnp 操纵子中 rpsO 上游启动的通读转录本水平，均观察到这种增加;在 rpsO-pnp 基因间区域启动的 pnp 转录本;并且 rpsO 脚本全部增加。在JSE1880中，这些转录本水平的提高反映了三个化合物的化学半衰期均增加。我们进一步证明， rpsO - pnp 操纵子的过表达导致JSE1880中的PNPase活性水平明显高于M145，这反映了RNase中PNPase表达被自动调节的可能性。 S. coelicolor中的III依赖方式。为了进一步探讨在JSE1880的基因间区域中启动的 pnp 转录水平的增加，我们在使用纯化的Co.coloreliaseRNase III的分析中以该转录为底物。这些测定揭示了迄今尚未发现的 pnp 转录本的RNase III切割位点。这些位点的位置由引物延伸确定，并显示它们位于茎-环结构的环中。

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