A Small Non-Coding RNA Mediates Transcript Stability and Expression of Cytochrome bd Ubiquinol Oxidase Subunit I in Rickettsia conorii.
A Small Non-Coding RNA Mediates Transcript Stability and Expression of Cytochrome bd Ubiquinol Oxidase Subunit I in Rickettsia conorii.
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DOI：
10.3390/ijms24044008
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发表时间：
2023-02-16
影响因子：
5.6
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中科院分区：
文献类型：
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小调节 RNA (sRNA) 现在因其在细菌毒力和生长的转录后调节中的作用而得到广泛认可。我们之前已经证明了康氏立克次体在与人类宿主和节肢动物载体相互作用过程中的生物发生和差异表达，以及康氏立克次体 sRNA Rc_sR42 与双顺反子细胞色素 bd 泛醇氧化酶亚基 I 和 II (cydAB) mRNA 的体外结合。然而，sRNA结合对cydAB双顺反子转录物稳定性以及cydA和cydB基因表达的调节机制和影响仍不清楚。在本研究中，我们确定了 R. conorii 体内感染期间小鼠肺和脑组织中 Rc_sR42 及其同源靶基因 cydA 和 cydB 的表达动态，并采用荧光和报告基因检测来解码 sRNA 在调节同源基因转录本中的作用。定量 RT-PCR 显示 R. conorii 体内感染期间 sRNA 及其同源靶基因转录本的表达发生显着变化，并且与脑组织相比，在肺部观察到这些转录本的丰度更高。有趣的是，虽然 Rc_sR42 和 cydA 的表达变化模式相似，表明 sRNA 对 mRNA 靶标的影响，但 cydB 的表达独立于 sRNA 表达。此外，我们构建了 sRNA 和 cydAB 双顺反子 mRNA 的报告质粒，以破译 sRNA 对 CydA 和 CydB 表达的作用。我们观察到在 sRNA 存在的情况下 CydA 的表达增加，但在 sRNA 存在或不存在的情况下检测到 CydB 表达没有变化。总之，我们的结果表明，Rc_sR42 的结合是 cydA 的调节所必需的，但 cydB 的调节则不然。目前正在进行进一步研究，以了解这种相互作用在 R. conorii 感染过程中对哺乳动物宿主和蜱虫媒介的影响。
Small regulatory RNAs (sRNAs) are now widely recognized for their role in the post-transcriptional regulation of bacterial virulence and growth. We have previously demonstrated the biogenesis and differential expression of several sRNAs in Rickettsia conorii during interactions with the human host and arthropod vector, as well as the in vitro binding of Rickettsia conorii sRNA Rc_sR42 to bicistronic cytochrome bd ubiquinol oxidase subunits I and II (cydAB) mRNA. However, the mechanism of regulation and the effect of sRNA binding on the stability of the cydAB bicistronic transcript and the expression of the cydA and cydB genes are still unknown. In this study, we determined the expression dynamics of Rc_sR42 and its cognate target genes, cydA and cydB, in mouse lung and brain tissues during R. conorii infection in vivo and employed fluorescent and reporter assays to decode the role of sRNA in regulating cognate gene transcripts. Quantitative RT-PCR revealed significant changes in the expression of sRNA and its cognate target gene transcripts during R. conorii infection in vivo, and a greater abundance of these transcripts was observed in the lungs compared to brain tissue. Interestingly, while Rc_sR42 and cydA exhibited similar patterns of change in their expression, indicating the influence of sRNA on the mRNA target, the expression of cydB was independent of sRNA expression. Further, we constructed reporter plasmids of sRNA and cydAB bicistronic mRNA to decipher the role of sRNA on CydA and CydB expression. We observed increased expression of CydA in the presence of sRNA but detected no change in CydB expression in the presence or absence of sRNA. In sum, our results demonstrate that the binding of Rc_sR42 is required for the regulation of cydA but not cydB. Further studies on understanding the influence of this interaction on the mammalian host and tick vector during R. conorii infection are in progress.
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