Expression data from sequential P. aeruginosa cystic fibrosis (CF) isolates. Pseudomonas aeruginosa

To provide a more detailed survey of adaptive changes in the physiology of P. aeruginosa (PA) during long-term infection of the cystic fibrosis (CF) lung, we performed a comparative proteome and transcriptome analysis of a set of isogenic sequential non-mutator and mutator isolates from three selected CF patients. Recently, we showed that during CF lung persistence PA mutators converge to a virulence-attenuated phenotype. In this study, we demonstrate that besides virulence-associated traits (VATs) the adaptation process of PA predominantly comprises metabolic pathways. In end-stage mutator strains, transcripts of genes encoding VATs, chemotaxis, degradation of aromatic compounds and several two-component regulatory systems were decreased. In contrast, several transcripts of genes or proteins involved in metabolism of fatty acids, nucleotides, amino acids and the generation of energy were increased. Of particular interest is the increased expression level of genes involved in (i) the anaerobic arginine-deiminase pathway, (ii) the anaerobic respiration such as nitrate-uptake protein OprF, redox-active azurin and cytchrome c551 peroxidase, (iii) the micro-aerobic respiration such as high oxygen-affinity cytochrome oxidase cbb3 (iv) the tricarboxylic acid cycle (TCA), glyoxylate shunt and anaplerotic carboxylation reactions to oxaloacetate. Strikingly, an increased transcription of the anaerobic regulator gene anr correlates with the up-regulation of ANR-dependent genes. In conclusion, these changes in transcriptome and proteome indicate an adaptive shift towards constitutive expression of genes of metabolic pathways obviously required for growth under micro-aerobic and nutritional conditions of suppurative CF lung tissue. Finally, these results provide us with new targets for antimicrobial agents and biomarkers reflecting adaptation of PA towards progressive CF lung disease. Keywords: in vitro study/interstrain comparison/clinical isolates/early nonmutator vs. late mutator; variable time point of isolation from cf respiratory secretions Overall design: P. aeruginosa isolates recovered from different time points of chronic cystic fibrosis lung disease were cultered in vitro, harvested for RNA extraction and hybridization on Affymetrix microarrays. We compared the transcriptome (triplicate microarrays) of early non-mutator P. aeruginosa isolates with late mutator isolates with high mutation frequency probably the driving force of an efficient adaptation to changing environements to conclude from differences in gene expression to the requirements of CF lung environment. Second publication of array data to be added later

为更细致地探究铜绿假单胞菌（Pseudomonas aeruginosa, PA）在囊性纤维化（cystic fibrosis, CF）肺部长期感染过程中的生理适应性变化，我们对3名选定CF患者来源的一系列同基因连续非突变型与突变型分离株开展了比较蛋白质组学与转录组学分析。
此前我们的研究已证实，在CF肺部定殖过程中，PA突变株会逐渐趋同至毒力减弱的表型。
本研究证实，除毒力相关性状（virulence-associated traits, VATs）外，PA的适应性演化过程主要涉及代谢通路。在终末期突变株中，编码VATs、趋化系统、芳香族化合物降解通路以及多种双组分调控系统的基因转录产物水平显著下调。与之相反，参与脂肪酸、核苷酸、氨基酸代谢及能量生成的部分基因/蛋白的转录产物水平显著上调。
尤为值得关注的是，参与以下通路的基因表达水平显著上调：(i) 厌氧精氨酸脱亚胺酶途径；(ii) 厌氧呼吸通路，包括硝酸盐摄取蛋白OprF、氧化还原活性天青蛋白（azurin）及细胞色素c551过氧化物酶；(iii) 微需氧呼吸通路，如高氧亲和力细胞色素氧化酶cbb3；(iv) 三羧酸循环（tricarboxylic acid cycle, TCA）、乙醛酸分流及生成草酰乙酸的回补性羧化反应。
值得注意的是，厌氧调控基因anr的转录水平上调，与ANR依赖型基因的表达上调呈显著相关性。
综上，转录组与蛋白质组的这些变化表明，PA发生了适应性表型转变，其代谢通路基因呈现组成型表达模式——这显然符合化脓性CF肺组织的微需氧与营养环境下的生长需求。
本研究结果为抗菌药物研发提供了全新靶点，同时也找到了可反映PA向进行性CF肺部疾病适应性演化的生物标志物。
关键词：体外研究/菌株间比较/临床分离株/早期非突变株与晚期突变株；从CF呼吸道分泌物分离的可变时间点样本
实验设计：从慢性囊性纤维化肺部疾病不同时间点分离得到的PA分离株经体外培养后，收集样本用于RNA提取，并在Affymetrix微阵列上进行杂交。我们将早期非突变型PA分离株与高突变频率的晚期突变型PA分离株的转录组（每组设置3次重复微阵列实验）进行比较——高突变频率可能是高效适应多变环境的驱动因素，旨在通过基因表达差异推导CF肺部环境的生存需求。本阵列数据的第二篇发表内容将于后续补充。