Molecular factors whereby giant phage ΦKZ modulates host protein synthesis

巨噬菌体δKZ调节宿主蛋白质合成的分子因素

• 批准号: 465133664
• 负责人: Professor Dr. Jörg Vogel
• 金额: --
• 依托单位: Institut für Molekulare Infektionsbiologie (IMIB)
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/465133664?language=en
• 关键词: Molecular; factors; whereby; giant; phage

Phages with large genomes and complex lifestyles represent exciting opportunities to discover new molecular factors and principles of host manipulation during infection. The vast majority of the proteins encoded in the genome sequences of these phages have no matches in the current sequence databases. For example, the 280-kB genome of ΦKZ, a giant bacteriophage attacking pseudomonads, possesses 369 annotated open reading frames but only a small fraction of the encoded proteins are of known function. On the one hand, this promises a huge discovery space for what will likely be novel proteins with specialized functions in host manipulation. On the other hand, the sheer number of these uncharacterized proteins demands new approaches to accelerate functional analysis. In preliminary work for this proposal, we have used RNA-centric high-throughput techniques to map with high-resolution the transcriptomes of phage ΦKZ and infected Pseudomonas aeruginosa cells and to predict candidate phage proteins with new functions in usurping the host bacterium. For example, gradient fractionation of complexes by size and density (Grad-seq), allowed us to generate a catalogue of potential protein and RNA complexes in the early phase of ΦKZ infection of P. aeruginosa cells. This revealed a striking accumulation of phage transcripts in ribosomal fractions thereby indicating an efficient transfer of viral transcripts to the translation machinery. More importantly, we observed numerous phage proteins to occur in ribosomal fractions, raising the possibility that ΦKZ actively manipulates the protein synthesis machinery of its host to its own benefit. Thus, we hypothesize that ΦKZ expresses proteins that modify ribosomes in infected Pseudomonas cells to prioritize the synthesis of its own proteins upon infection. Therefore, in the initial phase of the SPP2330, we will identify phage protein complexes and characterise their impact on mRNA translation in host bacteria, with the long-term goal to better understand if and how host protein synthesis is controlled during phage infection. The identification of phage proteins that target host protein synthesis will provide important biological insight into the phage-host interface, has potential for exploitation in antimicrobial strategies, and may yield new tools for synthetic biology.

具有大基因组和复杂生活方式的噬菌体代表了发现感染期间宿主操纵的新分子因子和原则的令人兴奋的机会。这些基因组序列中编码的绝大多数蛋白质在当前的序列数据库中没有匹配。例如，攻击假单胞菌的巨大噬菌体ΦKZ的280 kB基因组具有369个注释的开放阅读框，但只有一小部分编码的蛋白质具有已知的功能。一方面，这为可能在宿主操纵中具有专门功能的新型蛋白质提供了巨大的发现空间。另一方面，这些未表征的蛋白质的绝对数量需要新的方法来加速功能分析。 在该提议的初步工作中，我们使用以RNA为中心的高通量技术以高分辨率绘制噬菌体ΦKZ和感染的铜绿假单胞菌细胞的转录组，并预测在篡夺宿主细菌中具有新功能的候选噬菌体蛋白。例如，复合物通过大小和密度的梯度分级分离（Grad-seq）允许我们在铜绿假单胞菌细胞的ΦKZ感染的早期阶段产生潜在蛋白质和RNA复合物的目录。这揭示了噬菌体转录物在核糖体部分中的显著积累，从而表明病毒转录物有效地转移到翻译机构。更重要的是，我们观察到许多噬菌体蛋白质出现在核糖体组分中，提高了ΦKZ主动操纵其宿主的蛋白质合成机制以实现自身利益的可能性。因此，我们假设ΦKZ在感染的假单胞菌细胞中表达修饰核糖体的蛋白质，以在感染时优先合成其自身的蛋白质。因此，在SPP 2330的初始阶段，我们将鉴定噬菌体蛋白复合物并研究其对宿主细菌中mRNA翻译的影响，长期目标是更好地了解噬菌体感染期间宿主蛋白合成是否以及如何控制。鉴定靶向宿主蛋白质合成的噬菌体蛋白质将为噬菌体-宿主界面提供重要的生物学见解，具有开发抗菌策略的潜力，并可能产生合成生物学的新工具。