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Transational coupling via termination-reinitiation at overlapping genes in archaea and bacteria

通过古细菌和细菌中重叠基因的终止-重新启动进行转换耦合

基本信息

批准号：
387838868
负责人：
Professor Dr. Jörg Soppa
金额：
--
依托单位：
Arbeitskreis Biologie und Genetik von Prokaryonten
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
资助国家：
德国
起止时间：
项目状态：
未结题

来源：
https://gepris.dfg.de/gepris/projekt/387838868?language=en
关键词：
Transational coupling via termination reinitiation

项目摘要

In the first granting period, we have shown that translational coupling via termination-reinitiation (TeRe) is highly effective at overlapping gene pairs in the model archaeon Haloferax volcanii and the model bacterium Escherichia coli. The importance of the Shine Dalgarno motif for the efficiency of TeRe is highly variable and gene specific. The efficiency of TeRe critically depends on gene overlaps or extremely short intergenic regions of less than 30 nt. In the second granting period, we will perform in depth analyses of the sequence and structural motifs that lead to efficient TeRe, including saturation mutagenesis near gene overlaps in H. volcanii and E. coli. As we have found indications that one biological function of TeRe might be the efficient formation of heteromeric protein complexes, quantitative Western blot analyses will be used to determine the absolute TeRe efficiencies for several gene pairs of H. volcanii and E. coli. An in vitro translation system will be used to unravel whether translational coupling via TeRe depends on the 30S subunit or on the undissociated 70S ribosome. In addition, expression from gene pairs on bicistronic transcripts will be compared with expression of the same genes on two monocistronic transcripts to yield indications whether co-translational formation of heteromeric complexes occurs in H. volcanii. Taken together, the project will yield unprecedented insight into the mechanism and biological function of translational coupling via termination-reinitiation in archaea and bacteria.

在第一个授权期内，我们已经证明通过终止-再起始（TeRe）的翻译偶联在模型古菌火山盐铁菌和模型细菌大肠杆菌中非常有效地重叠基因对。Shine Dalgarno基序对TeRe效率的重要性是高度可变和基因特异性的。TeRe的效率主要取决于基因重叠或小于30 nt的极短基因间区域。在第二个授权期，我们将对导致高效TeRe的序列和结构基序进行深入分析，包括火山杆菌和大肠杆菌基因重叠附近的饱和诱变。由于我们已经发现有迹象表明，TeRe的一个生物学功能可能是有效地形成异质蛋白复合物，定量Western blot分析将用于确定火山嗜血杆菌和大肠杆菌的几个基因对的绝对TeRe效率。一个体外翻译系统将用于揭示通过TeRe的翻译偶联是取决于30S亚基还是取决于未解离的70S核糖体。此外，将双反子转录本上基因对的表达与同一基因在两个单反子转录本上的表达进行比较，以确定在火山螺中是否发生了异质复合物的共翻译形成。综上所述，该项目将对古细菌和细菌中通过终止-再起始的翻译偶联的机制和生物学功能产生前所未有的见解。