大肠埃希菌中PNPase调控持留菌形成的机制研究

细菌持留感染导致疾病反复发作，久治不愈，但其确切机制尚不清楚。mRNA的降解途径可能在持留细菌形成中发挥重要作用，课题组前期研究发现抗结核药物吡嗪酰胺可抑制结核分枝杆菌mRNA通过反式翻译的特殊降解途径发挥作用，本课题我们将在大肠埃希菌中，对mRNA一般降解途径中的关键基因调控细菌持留形成作深入研究。多核苷酸磷酸化酶（PNPase）是RNA降解体的主要成分，在mRNA一般降解过程中发挥关键性的作用。我们拟通过研究pnp基因敲除株在不同压力条件处理后的存活能力，及在转录组、蛋白组水平上基因表达的变化；比较正常细菌和持留菌中pnp基因在各种应激条件下的表达差异；研究动物模型中PNPase对大肠埃希菌持留、毒力等方面的功能，来明确PNPase在持留菌存活和毒力中发挥的作用。通过本研究能够明确PNPase在持留菌存活的重要作用，阐明其调控持留菌形成的机制，为研发持留菌新型抗生素提供靶标。

In clinical setting, persister bacteria frequently cause post-antibiotic relapse and are underlying lengthy therapy. The exact mechanisms of persister formation are still unclear. Through the study of persister-specific front-line tuberculosis drug pyrazinamide (PZA), we have found that the special mRNA degradation pathway (trans-translation) is critical for persister survival. But so far, no systematic research has been carried out on common mRNA degradation pathway. Polynucleotide phosphorylase (PNPase) is a major component of RNA Degradosome , which plays a key role in mRNA degrading process. In this proposal, we propose to investigate the role of PNPase in the persister survival and virulence , and clarify the exact mechanisms involved in persister formation. Specifically, we will study the viability of pnp mutant after the exposure of various stresses or antibiotics; compare the pnp gene expression levels between persister cells and normal cells; assess the effects of PNPase on persistence, virulence and antibiotic clearance in animal models; compare changes in genes expression of PNPase mutant in transcriptome and proteome level. This research will clarify the important influence of PNPase in persister survival, and shed new light on mechanisms by which PNPase contributes to persister formation, and provide validated new targets for developing a new generation of persister antibiotics for more effective treatment of persistent bacterial infections.