Handford Jennifer I, Ize Bérengère, Buchanan Grant, Butland Gareth P, Greenblatt Jack, Emili Andrew, Palmer Tracy
Department of Molecular Microbiology, John Innes Centre, Norwich, United Kingdom.
J Bacteriol. 2009 Aug;191(15):4732-49. doi: 10.1128/JB.00136-09. Epub 2009 Apr 17.
The yjeE, yeaZ, and ygjD genes are highly conserved in the genomes of eubacteria, and ygjD orthologs are also found throughout the Archaea and eukaryotes. In this study, we have constructed conditional expression strains for each of these genes in the model organism Escherichia coli K12. We show that each gene is essential for the viability of E. coli under laboratory growth conditions. Growth of the conditional strains under nonpermissive conditions results in dramatic changes in cell ultrastructure. Deliberate repression of the expression of yeaZ results in cells with highly condensed nucleoids, while repression of yjeE and ygjD expression results in at least a proportion of very enlarged cells with an unusual peripheral distribution of DNA. Each of the three conditional expression strains can be complemented by multicopy clones harboring the rstA gene, which encodes a two-component-system response regulator, strongly suggesting that these proteins are involved in the same essential cellular pathway. The results of bacterial two-hybrid experiments show that YeaZ can interact with both YjeE and YgjD but that YgjD is the preferred interaction partner. The results of in vitro experiments indicate that YeaZ mediates the proteolysis of YgjD, suggesting that YeaZ and YjeE act as regulators to control the activity of this protein. Our results are consistent with these proteins forming a link between DNA metabolism and cell division.
yjeE、yeaZ和ygjD基因在真细菌基因组中高度保守,并且在古细菌和真核生物中也能找到ygjD的直系同源基因。在本研究中,我们在模式生物大肠杆菌K12中构建了这些基因各自的条件表达菌株。我们发现,在实验室生长条件下,每个基因对于大肠杆菌的生存能力都是必需的。条件菌株在非允许条件下生长会导致细胞超微结构发生显著变化。特意抑制yeaZ的表达会导致细胞出现高度浓缩的类核,而抑制yjeE和ygjD的表达则会导致至少一部分细胞异常增大,且DNA呈异常的外周分布。这三种条件表达菌株中的每一种都可以被携带rstA基因的多拷贝克隆所互补,rstA基因编码一种双组分系统应答调节因子,这强烈表明这些蛋白质参与了相同的基本细胞途径。细菌双杂交实验结果表明,YeaZ可以与YjeE和YgjD都发生相互作用,但YgjD是更倾向的相互作用伙伴。体外实验结果表明,YeaZ介导YgjD的蛋白水解,这表明YeaZ和YjeE作为调节因子来控制该蛋白的活性。我们的结果与这些蛋白质在DNA代谢和细胞分裂之间形成联系是一致的。