McPhee Joseph B, Bains Manjeet, Winsor Geoff, Lewenza Shawn, Kwasnicka Agnieszka, Brazas Michelle D, Brinkman Fiona S L, Hancock R E W
Department of Microbiology and Immunology and Centre for Microbial Diseases and Immunity Research, University of British Columbia, 232-2259 Lower Mall, Vancouver, BC, Canada V6T 1Z4.
J Bacteriol. 2006 Jun;188(11):3995-4006. doi: 10.1128/JB.00053-06.
When grown in divalent cation-limited medium, Pseudomonas aeruginosa becomes resistant to cationic antimicrobial peptides and polymyxin B. This resistance is regulated by the PhoP-PhoQ and PmrA-PmrB two-component regulatory systems. To further characterize Mg(2+) regulation in P. aeruginosa, microarray transcriptional profiling was conducted to compare wild-type P. aeruginosa grown under Mg(2+)-limited and Mg(2+)-replete conditions to isogenic phoP and pmrA mutants grown under Mg(2+)-limited conditions. Under Mg(2+)-limited conditions (0.02 mM Mg(2+)), approximately 3% of the P. aeruginosa genes were differentially expressed compared to the expression in bacteria grown under Mg(2+)-replete conditions (2 mM Mg(2+)). Only a modest subset of the Mg(2+)-regulated genes were regulated through either PhoP or PmrA. To determine which genes were directly regulated, a bioinformatic search for conserved binding motifs was combined with confirmatory reverse transcriptase PCR and gel shift promoter binding assays, and the results indicated that very few genes were directly regulated by these response regulators. It was found that in addition to the previously known oprH-phoP-phoQ operon and the pmrHFIJKLM-ugd operon, the PA0921 and PA1343 genes, encoding small basic proteins, were regulated by Mg(2+) in a PhoP-dependent manner. The number of known PmrA-regulated genes was expanded to include the PA1559-PA1560, PA4782-PA4781, and feoAB operons, in addition to the previously known PA4773-PA4775-pmrAB and pmrHFIJKLM-ugd operons.
当在二价阳离子受限的培养基中生长时，铜绿假单胞菌会对阳离子抗菌肽和多粘菌素B产生抗性。这种抗性由PhoP - PhoQ和PmrA - PmrB双组分调节系统调控。为了进一步表征铜绿假单胞菌中镁离子（Mg²⁺）的调节作用，进行了微阵列转录谱分析，以比较在Mg²⁺受限和Mg²⁺充足条件下生长的野生型铜绿假单胞菌与在Mg²⁺受限条件下生长的同基因phoP和pmrA突变体。在Mg²⁺受限条件（0.02 mM Mg²⁺）下，与在Mg²⁺充足条件（2 mM Mg²⁺）下生长的细菌相比，约3%的铜绿假单胞菌基因表达存在差异。只有一小部分受Mg²⁺调节的基因是通过PhoP或PmrA调节的。为了确定哪些基因是直接受调节的，将保守结合基序的生物信息学搜索与验证性逆转录酶PCR和凝胶迁移启动子结合试验相结合，结果表明很少有基因直接受这些反应调节因子的调节。研究发现，除了先前已知的oprH - phoP - phoQ操纵子和pmrHFIJKLM - ugd操纵子外，编码小碱性蛋白的PA0921和PA1343基因也以PhoP依赖的方式受Mg²⁺调节。已知受PmrA调节的基因数量有所增加，除了先前已知的PA4773 - PA4775 - pmrAB和pmrHFIJKLM - ugd操纵子外，还包括PA1559 - PA1560、PA4782 - PA4781和feoAB操纵子。
