Benson N R, Wong R M, McClelland M
The Sidney Kimmel Cancer Center, San Diego, California 92121, USA.
J Bacteriol. 2000 Jun;182(12):3490-7. doi: 10.1128/JB.182.12.3490-3497.2000.
We report an analysis of a sample of the SOS response of Salmonella enterica serovar Typhimurium using the differential display of RNA fingerprinting gels of arbitrarily primed PCR products. The SOS response was induced by the addition of mitomycin C to an exponentially growing culture of serovar Typhimurium, and the RNA population was sampled during the following 2 h. These experiments revealed 21 differentially expressed PCR fragments representing mRNA transcripts. These 21 fragments correspond to 20 distinct genes. All of these transcripts were positively regulated, with the observed induction starting 10 to 120 min after addition of mitomycin C. Fifteen of the 21 transcripts have no homologue in the public sequence data banks and are therefore classified as novel. The remaining six transcripts corresponded to the recE, stpA, sulA, and umuC genes, and to a gene encoding a hypothetical protein in the Escherichia coli lysU-cadA intergenic region; the recE gene was represented twice by nonoverlapping fragments. In order to determine if the induction of these 20 transcripts constitutes part of a classical SOS regulon, we assessed the induction of these genes in a recA mutant. With one exception, the increased expression of these genes in response to mitomycin C was dependent on the presence of a functional recA allele. The exception was fivefold induced in the absence of a functional RecA protein, suggesting another layer of regulation in response to mitomycin C, in addition to the RecA-LexA pathway of SOS induction. Our data reveal several genes belonging to operons known to be directly involved in pathogenesis. In addition, we have found several phage-like sequences, some of which may be landmarks of pathogenicity determinants. On the basis of these observations, we propose that the general use of DNA-damaging agents coupled with differential gene expression analysis may be a useful and easy method for identifying pathogenicity determinants in diverse organisms.
我们报告了一项对鼠伤寒沙门氏菌血清型鼠伤寒菌株SOS应答样本的分析，该分析使用了任意引物PCR产物的RNA指纹图谱凝胶的差异显示技术。通过向指数生长的鼠伤寒血清型培养物中添加丝裂霉素C来诱导SOS应答，并在接下来的2小时内对RNA群体进行采样。这些实验揭示了21个差异表达的PCR片段，代表mRNA转录本。这21个片段对应于20个不同的基因。所有这些转录本均受到正向调控，观察到的诱导在添加丝裂霉素C后10至120分钟开始。21个转录本中的15个在公共序列数据库中没有同源物，因此被归类为新基因。其余6个转录本分别对应recE、stpA、sulA和umuC基因，以及大肠杆菌lysU-cadA基因间隔区中一个编码假定蛋白的基因；recE基因由不重叠的片段代表了两次。为了确定这20个转录本的诱导是否构成经典SOS调节子的一部分，我们评估了recA突变体中这些基因的诱导情况。除了一个例外，这些基因对丝裂霉素C的表达增加依赖于功能性recA等位基因的存在。这个例外在没有功能性RecA蛋白的情况下被诱导了五倍，这表明除了SOS诱导的RecA-LexA途径外，对丝裂霉素C的应答还存在另一层调控。我们的数据揭示了几个属于已知直接参与致病过程的操纵子的基因。此外，我们还发现了几个噬菌体样序列，其中一些可能是致病性决定因素的标志。基于这些观察结果，我们提出，结合差异基因表达分析使用DNA损伤剂可能是一种在多种生物体中鉴定致病性决定因素的有用且简便的方法。
