Mechanisms of gene regulation by the conserved small RNA rnTrpL (formerly RcsR1) and its new interaction partners in Sinorhizobium meliloti

在所有被研究的生物中，反式作用小RNA(SRNAs)通过与目标mRNAs不完全的碱基配对来调节基因的表达。通常，预测的目标比验证的目标多，这表明在特定条件下可以解决一些目标。然而，对sRNA重新编程以响应刺激的例子仍然缺乏。我们正在研究SRNA rnTrpL(以前的RcsR1)，它是由于色氨酸生物合成基因trpE(G)的转录减弱而产生的。该SRNA含有编码前导肽peTrpL的TRPL ORF，并具有许多预测的靶点。在第一个资助期，我们验证了其中的两个，trpDC和rplUrpmA mRNAs，后者编码核糖体蛋白L21和L27。令人惊讶的是，与trpDC相反，rplUrpmA与peTrpL多肽一起被rnTrpL靶向，并且只有在接触翻译抑制抗生素时才被靶向。RnTrpL SRNA与rplU在抗生素依赖核糖核蛋白(ARNP)复合体中配对，该复合体还包括peTrpL肽、抗生素和抗生素诱导的反义RNA。在下一个资金阶段，我们的目标是分析抗生素和peTrpL重新编程rnTrpL以支持rplUrpmA的机制。我们将研究rnTrpL-ARNP复合体的组装和结构，rplUrpmA失稳的机制和作用，以及反义RNA在这一过程中的作用。另一个目标是在几种条件下鉴定rnTrpL相互作用组。这一结果将揭示一个保守的sRNA的网络和机制，该网络和机制具有对抗生素反应的特异性变化。

In all investigated organisms, trans-acting small RNAs (sRNAs) regulate gene expression by imperfect base pairing with target mRNAs. Usually more targets are predicted than validated, suggesting that some targets could be addressed under specific conditions. However, examples for sRNA reprogramming in response to stimuli are still lacking. We are investigating the sRNA rnTrpL (formerly RcsR1), which arises due to transcription attenuation of the tryptophan biosynthesis gene trpE(G). This sRNA contains a trpL ORF encoding the leader peptide peTrpL and has many predicted targets. In the first funding period, we validated two of them, the trpDC and rplUrpmA mRNAs, the latter encoding ribosomal proteins L21 and L27. Surprisingly, in contrast to trpDC, rplUrpmA was targeted by rnTrpL in conjunction with the peTrpL peptide and only upon exposure to translation inhibiting antibiotics. The rnTrpL sRNA base pairs with rplU in an antibiotic-dependent ribonucleoprotein (ARNP) complex, which in addition comprises the peTrpL peptide, an antibiotic, and an antibiotic-induced antisense RNA. In the next funding period, we aim to analyze the mechanism by which antibiotics and peTrpL reprogram rnTrpL in favor of rplUrpmA. We will study the assembly and structure of rnTrpL-ARNP complexes, the mechanism and role of rplUrpmA destabilization, and the role of antisense RNAs in this process. Another goal is to identify the rnTrpL interactome under several conditions. The results will uncover the network and mechanisms of a conserved sRNA with specificity change in response to antibiotics.