Jakob Kinga, Satorhelyi Peter, Lange Christian, Wendisch Volker F, Silakowski Barbara, Scherer Siegfried, Neuhaus Klaus
Lehrstuhl für Mikrobielle Okologie, Technische Universität München, D-85354 Freising, Germany.
J Bacteriol. 2007 Aug;189(15):5582-90. doi: 10.1128/JB.00082-07. Epub 2007 May 25.
Corynebacteria form an important part of the red smear cheese microbial surface consortium. To gain a better understanding of molecular adaptation due to low pH induced by lactose fermentation, the global gene expression profile of Corynebacterium glutamicum adapted to pH 5.7 with lactic acid under continuous growth in a chemostat was characterized by DNA microarray analysis. Expression of a total of 116 genes was increased and that of 90 genes was decreased compared to pH 7.5 without lactic acid, representing 7% of the genes in the genome. The up-regulated genes encode mainly transcriptional regulators, proteins responsible for export, import, and metabolism, and several proteins of unknown function. As much as 45% of the up-regulated open reading frames code for hypothetical proteins. These results were validated using real-time reverse transcription-PCR. To characterize the functions of 38 up-regulated genes, 36 single-crossover disruption mutants were generated and analyzed for their lactic acid sensitivities. However, only a sigB knockout mutant showed a highly significant negative effect on growth at low pH, suggesting a function in organic-acid adaptation. A sigE mutant already displayed growth retardation at neutral pH but grew better at acidic pH than the sigB mutant. The lack of acid-sensitive phenotypes in 34 out of 36 disrupted genes suggests either a considerable redundancy in acid adaptation response or coincidental effects. Other up-regulated genes included genes for ion transporters and metabolic pathways, including carbohydrate and respiratory metabolism. The enhanced expression of the nrd (ribonucleotide reductase) operon and a DNA ATPase repair protein implies a cellular response to combat acid-induced DNA damage. Surprisingly, multiple iron uptake systems (totaling 15% of the genes induced >or=2-fold) were induced at low pH. This induction was shown to be coincidental and could be attributed to iron-sequestering effects in complex media at low pH.
棒状杆菌是红涂抹奶酪微生物表面菌群的重要组成部分。为了更好地理解乳糖发酵诱导的低pH条件下的分子适应性，通过DNA微阵列分析对在恒化器中持续生长时适应pH 5.7乳酸环境的谷氨酸棒状杆菌的全局基因表达谱进行了表征。与无乳酸的pH 7.5相比，共有116个基因的表达增加，90个基因的表达减少，占基因组中基因的7%。上调的基因主要编码转录调节因子、负责输出、输入和代谢的蛋白质以及几种功能未知的蛋白质。多达45%的上调开放阅读框编码假设蛋白质。这些结果通过实时逆转录PCR得到验证。为了表征38个上调基因的功能，构建了36个单交叉破坏突变体并分析了它们对乳酸的敏感性。然而，只有一个sigB基因敲除突变体在低pH条件下对生长表现出高度显著的负面影响，表明其在有机酸适应中发挥作用。一个sigE突变体在中性pH条件下已经表现出生长迟缓，但在酸性pH条件下比sigB突变体生长得更好。36个被破坏基因中的34个缺乏酸敏感表型，这表明酸适应反应中存在相当大的冗余或巧合效应。其他上调基因包括离子转运蛋白和代谢途径的基因，包括碳水化合物和呼吸代谢。nrd（核糖核苷酸还原酶）操纵子和一种DNA ATP酶修复蛋白的表达增强意味着细胞对酸诱导的DNA损伤的反应。令人惊讶的是，在低pH条件下诱导了多个铁摄取系统（总共占诱导倍数≥2倍的基因的15%）。这种诱导被证明是巧合的，可能归因于低pH条件下复杂培养基中铁螯合效应。
