The role of small RNAs in the control of amino sugar metabolism in Escherichia coli

近年来，小的非编码的Frans编码的RNA作为一类新的基因调控因子出现在所有生物体内。在许多细菌中，如大肠杆菌中，已经发现了大量这样的sRNA，但它们的生理功能仅在少数情况下被揭示。在已知的情况下，这些sRNA最常与其靶mRNAs碱基配对，从而影响其靶mRNAs的翻译和/或稳定性。我们最近发现，大肠杆菌氨基葡萄糖-6-磷酸合成酶(GLMS)的表达是由其产物氨基葡萄糖-6-磷酸(GlcN-6-P)反馈调节的，这个过程依赖于小RNA GlmZ(Kalamorz等人，2007，Mol。微生物。65,1518-1533)。GLMS启动氨基己糖途径，导致UDP-N-乙酰氨基葡萄糖的形成，UDP-N-乙酰氨基葡萄糖是细菌细胞壁生物合成肽聚糖的主要中间体。Sna GlmZ是经过加工的，我们发现细胞内GlcN-6-P浓度的下降会导致其未加工形式的积累。这种全长的GlmZ反过来可能通过直接碱化来稳定GLMS mRNA，从而导致更高的GLMS合成水平。此外，我们还鉴定了新的蛋白质YhbJ作为GlmZ加工的调节因子。此外，我们还发现了第二个SRNA，GlmY，它也受到yhbJ突变和GlcN-6-P浓度的影响，当过表达时，也会诱导GLMS的表达。我们的数据表明，GlmY影响GlmZ的加工，从而间接调节GLMS的表达。在这里，我们打算详细研究这一新的基因控制机制。我们的目标是阐明GlmZ和GlmY两个sRNAs在GLMS表达调控中的干扰作用，并鉴定GlcN-6-P传感器。此外，我们希望使用体内和体外技术来表征GlmZ/GLMS的基本特性。另一项重要任务是找出YhbJ调控这些sRNA加工的分子机制。为此，我们将对YhbJ的体外性质进行研究。最后，利用整体蛋白质组和转录组方法，我们想要找出参与氨基糖或氮代谢的额外基因是否受YhbJ、GlmZ和/或GlmY控制，即这些因素是否控制一个调节子。

In recent years, small non-coding frans-encoded RNAs emerged as a new class of gene regulators in all living organisms. In many bacteria, such as Escherichia coli, a plethora of such sRNAs has been identified but their physiological functions have been revealed in just a few cases. Where known, these sRNAs most often base-pair to and thereby affect translation and/or stability of their target mRNAs. We recently discovered that expression of the E. coli glucosamine-6-phosphate synthase (GlmS) is feedback-regulated by its product glucosamine-6-phosphate (GlcN-6-P) in a process that depends on the small RNA GlmZ (Kalamorz et al., 2007, Mol. Microbiol. 65, 1518-1533). GlmS initiates the hexosamine pathway leading to the formation of UDP-N-acetylglucosamine, the major intermediate in the biosynthesis of peptidoglycan of bacterial cell walls. The sRNA GlmZ is subject to processing and we found that a decrease in cellular GlcN-6-P concentrations leads to accumulation of its unprocessed form. This full-length GlmZ in turn stabilizes the glmS mRNA presumably by direct basepairing which results in higher GlmS synthesis levels. Furthermore, we identified the novel protein YhbJ as a regulator of GlmZ processing. In addition, we identified a second sRNA, GlmY, that is also affected by a yhbJ mutation and by GlcN-6-P concentrations and which when overexpressed also induces glmS expression. Our data suggest that GlmY affects processing of GlmZ and thereby indirectly regulates glmS expression. Here we propose to investigate this novel genetic control mechanism in detail. Our goals are to clarify the interference of the two sRNAs GlmZ and GlmY in the control of glmS expression and to identify the GlcN-6-P sensor. Furthermore, we would like to characterize the GlmZ/glmS basepairing using in vivo as well as in vitro techniques. Another important task is to find out the molecular mechanism by which YhbJ governs processing of these sRNAs. For this purpose, we will investigate the in vitro properties of YhbJ. Finally, using global proteome and transcriptome approaches we want to find out whether additional genes involved in amino sugar or nitrogen metabolism are controlled by YhbJ, GlmZ and/or GlmY i.e. whether these factors govern a regulon.