Wang Lijuan, Perpich John, Driks Adam, Kroos Lee
Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI 48824, USA.
J Bacteriol. 2007 Oct;189(20):7302-9. doi: 10.1128/JB.00839-07. Epub 2007 Aug 10.
During sporulation of Bacillus subtilis, four regulatory proteins act in the order sigma(E), SpoIIID, sigma(K), and GerE to temporally control gene expression in the mother cell. sigma(E) and sigma(K) work sequentially with core RNA polymerase to transcribe different sets of genes. SpoIIID and GerE are small, sequence-specific DNA-binding proteins that activate or repress transcription of many genes. Previous studies showed that transcriptionally active sigma(K) RNA polymerase inhibits early mother cell gene expression, reducing accumulation of SpoIIID late in sporulation. Here, the effects of perturbing the mother cell gene regulatory network by maintaining the SpoIIID level late during sporulation are reported. Persistent expression was obtained by fusing spoIIID to the sigma(K)-controlled gerE promoter on a multicopy plasmid. Fewer heat- and lysozyme-resistant spores were produced by the strain with persistent spoIIID expression, but the number of spores resistant to organic solvents was unchanged, as was their germination ability. Transmission electron microscopy showed structural defects in the spore coat. Reporter fusions to sigma(K)-dependent promoters showed lower expression of gerE and cotC and higher expression of cotD. Altered expression of cot genes, which encode spore coat proteins, may account for the spore structural defects. These results suggest that one role of negative feedback by sigma(K) RNA polymerase on early mother cell gene expression is to lower the level of SpoIIID late during sporulation in order to allow normal expression of genes in the sigma(K) regulon.
在枯草芽孢杆菌的芽孢形成过程中，四种调节蛋白按σ(E)、SpoIIID、σ(K)和GerE的顺序发挥作用，以在时间上控制母细胞中的基因表达。σ(E)和σ(K)依次与核心RNA聚合酶协同作用，转录不同的基因集。SpoIIID和GerE是小的、序列特异性的DNA结合蛋白，可激活或抑制许多基因的转录。先前的研究表明，具有转录活性的σ(K) RNA聚合酶会抑制母细胞早期基因表达，减少芽孢形成后期SpoIIID的积累。在此，报告了在芽孢形成后期维持SpoIIID水平对扰乱母细胞基因调控网络的影响。通过将spoIIID与多拷贝质粒上受σ(K)控制的gerE启动子融合，实现了持续表达。持续表达spoIIID的菌株产生的耐热和耐溶菌酶的芽孢较少，但对有机溶剂有抗性的芽孢数量及其萌发能力未发生变化。透射电子显微镜显示芽孢外壳存在结构缺陷。与依赖σ(K)的启动子的报告基因融合显示，gerE和cotC的表达较低，而cotD的表达较高。编码芽孢外壳蛋白的cot基因表达改变可能是芽孢结构缺陷的原因。这些结果表明，σ(K) RNA聚合酶对母细胞早期基因表达的负反馈作用之一是在芽孢形成后期降低SpoIIID的水平，以便σ(K)调控子中的基因能够正常表达。
