Jochmann Nina, Kurze Anna-Katharina, Czaja Lisa F, Brinkrolf Karina, Brune Iris, Hüser Andrea T, Hansmeier Nicole, Pühler Alfred, Borovok Ilya, Tauch Andreas
International NRW Graduate School in Bioinformatics and Genome Research, Centrum für Biotechnologie, Universität Bielefeld, Universitätsstraße 25, D-33615 Bielefeld, Germany.
Institut für Genomforschung und Systembiologie, Centrum für Biotechnologie, Universität Bielefeld, Universitätsstraße 27, D-33615 Bielefeld, Germany.
Microbiology (Reading). 2009 May;155(Pt 5):1459-1477. doi: 10.1099/mic.0.025841-0. Epub 2009 Apr 16.
The lexA gene of Corynebacterium glutamicum ATCC 13032 was deleted to create the mutant strain C. glutamicum NJ2114, which has an elongated cell morphology and an increased doubling time. To characterize the SOS regulon in C. glutamicum, the transcriptomes of NJ2114 and a DNA-damage-induced wild-type strain were compared with that of a wild-type control using DNA microarray hybridization. The expression data were combined with bioinformatic pattern searches for LexA binding sites, leading to the detection of 46 potential SOS boxes located upstream of differentially expressed transcription units. Binding of a hexahistidyl-tagged LexA protein to 40 double-stranded oligonucleotides containing the potential SOS boxes was demonstrated in vitro by DNA band shift assays. It turned out that LexA binds not only to SOS boxes in the promoter-operator region of upregulated genes, but also to SOS boxes detected upstream of downregulated genes. These results demonstrated that LexA controls directly the expression of at least 48 SOS genes organized in 36 transcription units. The deduced genes encode a variety of physiological functions, many of them involved in DNA repair and survival after DNA damage, but nearly half of them have hitherto unknown functions. Alignment of the LexA binding sites allowed the corynebacterial SOS box consensus sequence TcGAA(a/c)AnnTGTtCGA to be deduced. Furthermore, the common intergenic region of lexA and the differentially expressed divS-nrdR operon, encoding a cell division suppressor and a regulator of deoxyribonucleotide biosynthesis, was characterized in detail. Promoter mapping revealed differences in divS-nrdR expression during SOS response and normal growth conditions. One of the four LexA binding sites detected in the intergenic region is involved in regulating divS-nrdR transcription, whereas the other sites are apparently used for negative autoregulation of lexA expression.
谷氨酸棒杆菌ATCC 13032的lexA基因被删除，以创建突变菌株谷氨酸棒杆菌NJ2114，该菌株具有拉长的细胞形态和延长的倍增时间。为了表征谷氨酸棒杆菌中的SOS调节子，使用DNA微阵列杂交将NJ2114和DNA损伤诱导的野生型菌株的转录组与野生型对照的转录组进行了比较。表达数据与用于LexA结合位点的生物信息学模式搜索相结合，从而检测到位于差异表达转录单元上游的46个潜在SOS框。通过DNA带移分析在体外证明了六聚组氨酸标记的LexA蛋白与40个含有潜在SOS框的双链寡核苷酸的结合。结果表明，LexA不仅与上调基因的启动子-操纵子区域中的SOS框结合，而且还与在下调基因上游检测到的SOS框结合。这些结果表明，LexA直接控制至少48个SOS基因的表达，这些基因组织在36个转录单元中。推导的基因编码多种生理功能，其中许多与DNA损伤后的DNA修复和存活有关，但其中近一半迄今功能未知。LexA结合位点的比对使得能够推导棒状杆菌SOS框共有序列TcGAA(a/c)AnnTGTtCGA。此外，还详细表征了lexA与差异表达的divS-nrdR操纵子的共同基因间区域，该操纵子编码细胞分裂抑制因子和脱氧核糖核苷酸生物合成的调节因子。启动子图谱分析揭示了SOS反应和正常生长条件下divS-nrdR表达的差异。在基因间区域检测到 的四个LexA结合位点之一参与调节divS-nrdR转录，而其他位点显然用于lexA表达的负自调节。
