Schramm Franziska, Borst Andreas, Linne Uwe, Soppa Jörg
Institute for Molecular Biosciences, Biocentre, Goethe-University, Frankfurt, Germany.
Mass Spectrometry Facility, Department of Chemistry, Phillipps University Marburg, Marburg, Germany.
Front Microbiol. 2021 Oct 26;12:742806. doi: 10.3389/fmicb.2021.742806. eCollection 2021.
Translation is an important step in gene expression. Initiation of translation is rate-limiting, and it is phylogenetically more diverse than elongation or termination. Bacteria contain only three initiation factors. In stark contrast, eukaryotes contain more than 10 (subunits of) initiation factors (eIFs). The genomes of archaea contain many genes that are annotated to encode archaeal homologs of eukaryotic initiation factors (aIFs). However, experimental characterization of aIFs is scarce and mostly restricted to very few species. To broaden the view, the protein-protein interaction network of aIFs in the halophilic archaeon has been characterized. To this end, tagged versions of 14 aIFs were overproduced, affinity isolated, and the co-isolated binding partners were identified by peptide mass fingerprinting and MS/MS analyses. The aIF-aIF interaction network was resolved, and it was found to contain two interaction hubs, (1) the universally conserved factor aIF5B, and (2) a protein that has been annotated as the enzyme ribose-1,5-bisphosphate isomerase, which we propose to rename to aIF2Bα. Affinity isolation of aIFs also led to the co-isolation of many ribosomal proteins, but also transcription factors and subunits of the RNA polymerase (Rpo). To analyze a possible coupling of transcription and translation, seven tagged Rpo subunits were overproduced, affinity isolated, and co-isolated proteins were identified. The Rpo interaction network contained many transcription factors, but also many ribosomal proteins as well as the initiation factors aIF5B and aIF2Bα. These results showed that transcription and translation are coupled in haloarchaea, like in . It seems that aIF5B and aIF2Bα are not only interaction hubs in the translation initiation network, but also key players in the transcription-translation coupling.
翻译是基因表达中的一个重要步骤。翻译起始是限速步骤，并且在系统发育上比延伸或终止更为多样。细菌仅含有三种起始因子。与之形成鲜明对比的是，真核生物含有超过10种（亚基的）起始因子（eIFs）。古菌的基因组包含许多被注释为编码真核生物起始因子（aIFs）的古菌同源物的基因。然而，对aIFs的实验表征很少，并且大多限于极少数物种。为了拓宽视野，已经对嗜盐古菌中aIFs的蛋白质 - 蛋白质相互作用网络进行了表征。为此，过量表达了14种aIFs的标签版本，进行亲和分离，并通过肽质量指纹图谱和串联质谱分析鉴定共分离的结合伴侣。解析了aIF - aIF相互作用网络，发现它包含两个相互作用中心，（1）普遍保守的因子aIF5B，以及（2）一种被注释为核糖 - 1,5 - 二磷酸异构酶的蛋白质，我们建议将其重新命名为aIF2Bα。aIFs的亲和分离还导致许多核糖体蛋白以及转录因子和RNA聚合酶（Rpo）的亚基的共分离。为了分析转录和翻译可能的偶联，过量表达了7种带标签的Rpo亚基，进行亲和分离，并鉴定共分离的蛋白质。Rpo相互作用网络包含许多转录因子，但也有许多核糖体蛋白以及起始因子aIF5B和aIF2Bα。这些结果表明，嗜盐古菌中的转录和翻译是偶联的，就像在……中一样。似乎aIF5B和aIF2Bα不仅是翻译起始网络中的相互作用中心，也是转录 - 翻译偶联中的关键参与者。
