The universally conserved ATPase as mediator of cellular stress response

YchF是一种功能未知的普遍保守的ATP酶，虽然它优先水解ATP而不是GTP，但仍被分类为翻译因子样GTP酶的成员。核糖体结合显示E. coli和Trypanosoma cruzii YchF中的表达，但其对蛋白质合成的影响目前尚不清楚。人YchF同源物Ola 1似乎不与核糖体结合，而是参与氧化应激反应的翻译后调节和肿瘤发生。后一种建议与在许多肿瘤中观察到的Ola 1上调一致。 然而，Ola 1功能的分子机制和可能的相互作用伴侣是完全未知的。我们对该高度保守的蛋白在大肠杆菌中的功能进行了研究。大肠杆菌中表达的YchF抑制了氧化应激反应。抑制不是由主要氧化应激反应蛋白（OxyR，KatG）的转录或翻译下调引起的，而是由过氧化氢酶活性的直接抑制引起的（Wenk等人，2012年）。 通过位点特异性交联方法和质谱分析证实了YchF与过氧化氢酶/过氧化物酶KatG、KatE和AhpCF之间的直接相互作用。这些数据表明，YchF通过对主要应激蛋白的迄今未知的翻译后抑制来调节氧化应激反应。YchF的磷酸化/去磷酸化及其ATP酶活性似乎对于这种抑制功能是重要的。我们的数据表明E. coli YchF和人Ola 1具有明显非常相似的功能，并且作为普遍保守的氧化应激反应抑制剂。我们的目的是确定YchF介导的抑制作用的靶点并揭示其潜在的分子机制。模式生物E. coli和革兰氏阳性菌S.酿酒酵母将被用于确定YchF相互作用体对氧化应激的应答，并揭示靶蛋白的YchF依赖性抑制的生化和分子框架。这包括对YchF表达和氧化应激反应中的修饰的详细分析。考虑到Ola 1在肿瘤发展和转移中的假定参与，一旦我们建立了YchF在大肠杆菌中功能的坚实框架，我们计划将我们的分析扩展到人类细胞系。coli/S.啤酒。这一知识转移将是推广建议的主要重点。

YchF is a universally conserved ATPase of unknown function, which is classified as a member of the translation-factor like GTPases, although it preferentially hydrolyzes ATP over GTP. Ribosome binding was shown for E. coli and Trypanosoma cruzii YchF, but the possible impact on protein synthesis is unknown so far. The human YchF homologue Ola1 does not seem to bind to ribosomes and instead is implicated in the post-translational regulation of the oxidative stress respsonse and in tumour development. This latter proposal is in line with the observed up-regulation of Ola1 in many tumours. However, the molecular mechanisms of Ola1 function and possible interaction partners are entirely unknown. Our data on the function of this highly conserved protein in E. coli demonstrate that YchF inhibits the oxidative stress response. Inhibition is not caused by a transcriptional or translational down-regulation of major oxidative stress response proteins (OxyR, KatG), but rather by a direct inhibition of catalase activity (Wenk et al., 2012). A direct interaction between YchF and the catalases/peroxidases KatG, KatE and AhpCF was demonstrated by site-specific cross-linking approaches and by mass spectrometry. These data suggest that YchF regulates the oxidative stress response by a so far unknown post-translational inhibition of major stress proteins. Phosphorylation/ dephosphorylation of YchF and its ATPase activity appear to be important for this inhibitory function. Our data suggest that E. coli YchF and human Ola1 have apparently very similar function and act as universally conserved inhibitors of the oxidative stress response. Our aim is to identify the targets of YchF mediated inhibition and to disclose the underlying molecular mechanism. The easily amendable model organisms E. coli and S. cerevisiae will be employed to determine the YchF interactome in response to oxidative stress and to reveal the biochemical and molecular framework of YchF-dependent inhibition of target proteins. This includes the detailed analyses on YchF expression and modification in response to oxidative stress. Considering the postulated involvement of Ola1 in tumour-development and metastasis, we plan to extend our analyses to human cell lines, once we have established a solid framework of YchF function in E. coli/S. cerevisiae. This knowledge transfer will be the main focus of an extension proposal.