Proteomics analysis of the membrane vesicles in Pseudomonas aeruginosa under X-ray irradiation

Pseudomonas aeruginosa, X-ray irradiation, inner membrane blebbing, membrane vesicles Almost all bacteria produce membrane vesicles (MVs) that have been found to be generally composed of lipopolysaccharides (LPSs), phospholipids (PLs), outer membrane proteins, periplasmic proteins and peptidoglycan (PG), cytoplasmic proteins, as well as nucleic acids, metabolites and signaling molecules. We have recently demonstrated that X-ray irradiation strongly induced the MV production in P. aeruginosa, but the detailed biogenesis mechanisms and protein components of MVs were largely unclear. In this study, we performed a quantitative proteomics analysis of the MVs; a total of 2320 proteins were identified, among which 72.33% were cytoplasmic proteins, followed by periplasmic proteins (12.59%), IMPs (8.45%), OMPs (5.26%) and others (1.38%). Gene ontology (GO) enrichment analysis of biological processes showed that nucleoside phosphate metabolic process, ribose phosphate metabolic process, nucleoside metabolic process, and purine-containing compound biosynthetic process were significantly enriched. As expected, cellular component analysis confirmed the enrichment of cytoplasm. Notably, molecular functional enrichment analysis revealed that a large proportion of proteins that were related to RNA binding, tRNA binding, RNA methyltransferase activity, aminoacyl-tRNA ligase activity, and ligase activity were highly enriched. KEGG enrichment analysis showed that pathways involved in ribosome, purine metabolism, tRNA biogenesis and RNA degradation were enriched. These results suggest that under X-ray irradiation stress, P. aeruginosa PAO1 cells could transport the damaged nucleic acids by producing MVs, and the RNA binding and degradation proteins contained in MVs may help P. aeruginosa to identify and degrade damaged RNA.

铜绿假单胞菌（Pseudomonas aeruginosa）、X射线辐照、内膜出芽、膜囊泡。几乎所有细菌均可产生膜囊泡（membrane vesicles，MVs），现有研究表明其通常由脂多糖（lipopolysaccharides，LPSs）、磷脂（phospholipids，PLs）、外膜蛋白、周质蛋白、肽聚糖（peptidoglycan，PG）、细胞质蛋白，以及核酸、代谢物与信号分子构成。本研究团队近期证实，X射线辐照可强烈诱导铜绿假单胞菌产生膜囊泡，但膜囊泡的具体生物发生机制与蛋白质组成仍未完全明晰。本研究针对膜囊泡开展了定量蛋白质组学分析，共鉴定出2320种蛋白质，其中72.33%为细胞质蛋白，其次为周质蛋白（12.59%）、整合膜蛋白（integral membrane proteins，IMPs）、外膜蛋白（outer membrane proteins，OMPs）及其他蛋白（1.38%）。基因本体（Gene Ontology，GO）富集分析的生物学过程结果显示，核苷磷酸代谢过程、核糖磷酸代谢过程、核苷代谢过程及含嘌呤化合物生物合成过程显著富集。细胞组分分析验证了细胞质组分的富集，与预期相符。值得注意的是，分子功能富集分析揭示，大量与RNA结合、tRNA结合、RNA甲基转移酶活性、氨酰-tRNA连接酶活性及连接酶活性相关的蛋白质呈现高度富集。京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes，KEGG）富集分析显示，核糖体、嘌呤代谢、tRNA生物发生及RNA降解相关通路显著富集。上述结果表明，在X射线辐照胁迫下，铜绿假单胞菌PAO1可通过产生膜囊泡运输受损核酸，而膜囊泡所含的RNA结合与降解相关蛋白，或可帮助铜绿假单胞菌识别并降解受损RNA。