Nolle Nicoletta, Felsl Angela, Heermann Ralf, Fuchs Thilo M
Lehrstuhl für Mikrobielle Ökologie, ZIEL-Institute for Food and Health, Wissenschaftszentrum Weihenstephan, Technische Universität München, Freising, Germany.
Biozentrum, Bereich Mikrobiologie, Ludwig-Maximilians-Universität München, Martinsried/Munich, Germany.
J Bacteriol. 2017 Jan 30;199(4). doi: 10.1128/JB.00595-16. Print 2017 Feb 15.
Galactitol degradation by salmonellae remains underinvestigated, although this metabolic capability contributes to growth in animals (R. R. Chaudhuri et al., PLoS Genet 9:e1003456, 2013, https://doi.org/10.1371/journal.pgen.1003456). The genes responsible for this metabolic capability are part of a 9.6-kb gene cluster that spans from gatY to gatR (STM3253 to STM3262) and encodes a phosphotransferase system, four enzymes, and a transporter of the major facilitator superfamily. Genome comparison revealed the presence of this genetic determinant in nearly all Salmonella strains. The generation time of Salmonella enterica serovar Typhimurium strain ST4/74 was higher in minimal medium with galactitol than with glucose. Knockout of STM3254 and gatC resulted in a growth-deficient phenotype of S Typhimurium, with galactitol as the sole carbon source. Partial deletion of gatR strongly reduced the lag phase of growth with galactitol, whereas strains overproducing GatR exhibited a near-zero growth phenotype. Luciferase reporter assays demonstrated strong induction of the gatY and gatZ promoters, which control all genes of this cluster except gatR, in the presence of galactitol but not glucose. Purified GatR bound to these two main gat gene cluster promoters as well as to its own promoter, demonstrating that this autoregulated repressor controls galactitol degradation. Surface plasmon resonance spectroscopy revealed distinct binding properties of GatR toward the three promoters, resulting in a model of differential gat gene expression. The cyclic AMP receptor protein (CRP) bound these promoters with similarly high affinities, and a mutant lacking crp showed severe growth attenuation, demonstrating that galactitol utilization is subject to catabolite repression. Here, we provide the first genetic characterization of galactitol degradation in Salmonella, revealing novel insights into the regulation of this dissimilatory pathway.
The knowledge of how pathogens adapt their metabolism to the compartments encountered in hosts is pivotal to our understanding of bacterial infections. Recent research revealed that enteropathogens have adapted specific metabolic pathways that contribute to their virulence properties, for example, by helping to overcome limitations in nutrient availability in the gut due to colonization resistance. The capability of Salmonella enterica serovar Typhimurium to degrade galactitol has already been demonstrated to play a role in vivo, but it has not been investigated so far on the genetic level. To our knowledge, this is the first molecular description of the galactitol degradation pathway of a pathogen.
尽管鼠伤寒沙门氏菌降解半乳糖醇的代谢能力有助于其在动物体内生长（R.R.乔杜里等人，《公共科学图书馆·遗传学》9：e1003456，2013年，https://doi.org/10.1371/journal.pgen.1003456），但对其研究仍不充分。负责这种代谢能力的基因是一个9.6 kb基因簇的一部分，该基因簇从gatY延伸至gatR（STM3253至STM3262），编码一个磷酸转移酶系统、四种酶和一个主要转运体超家族的转运蛋白。基因组比较显示，几乎所有沙门氏菌菌株中都存在这种遗传决定因素。在以半乳糖醇为唯一碳源的基本培养基中，肠炎沙门氏菌血清型鼠伤寒菌株ST4/74的代时比在以葡萄糖为碳源的培养基中更长。敲除STM3254和gatC导致鼠伤寒沙门氏菌在以半乳糖醇为唯一碳源时出现生长缺陷表型。gatR的部分缺失强烈缩短了以半乳糖醇为碳源时的生长延迟期，而过量表达GatR的菌株表现出近乎零生长的表型。荧光素酶报告基因检测表明，在存在半乳糖醇而非葡萄糖的情况下，gatY和gatZ启动子（控制该基因簇中除gatR外的所有基因）被强烈诱导。纯化的GatR与这两个主要的gat基因簇启动子以及其自身的启动子结合，表明这种自我调节的阻遏蛋白控制着半乳糖醇的降解。表面等离子体共振光谱揭示了GatR对这三个启动子具有不同的结合特性，从而得出了gat基因差异表达的模型。环腺苷酸受体蛋白（CRP）以相似的高亲和力结合这些启动子，缺乏crp的突变体表现出严重的生长衰减，表明半乳糖醇的利用受到分解代谢物阻遏的调控。在此，我们首次对沙门氏菌中半乳糖醇降解进行了遗传特征分析，并揭示了这条异化途径调控的新见解。
了解病原体如何使其代谢适应宿主内的微环境对于我们理解细菌感染至关重要。最近的研究表明，肠道病原体已经适应了特定的代谢途径，这些途径有助于其毒力特性，例如通过帮助克服由于定植抗性导致的肠道营养可用性限制。肠炎沙门氏菌血清型鼠伤寒菌株降解半乳糖醇的能力已被证明在体内发挥作用，但迄今为止尚未在基因水平上进行研究。据我们所知，这是对病原体半乳糖醇降解途径的首次分子描述。
