den Hengst Chris D, van Hijum Sacha A F T, Geurts Jan M W, Nauta Arjen, Kok Jan, Kuipers Oscar P
Department of Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, Kerklaan 30, 9751 NN Haren and Friesland Foods Corporate Research, P. O. Box 87, 7400 AB Deventer, The Netherlands.
J Biol Chem. 2005 Oct 7;280(40):34332-42. doi: 10.1074/jbc.M502349200. Epub 2005 Jul 21.
CodY of Lactococcus lactis MG1363 is a transcriptional regulator that represses the expression of several genes encoding proteins of the proteolytic system. DNA microarray analysis, comparing the expression profiles of L. lactis MG1363 and an isogenic strain in which codY was mutated, was used to determine the CodY regulon. In peptide-rich medium and exponentially growing cells, where CodY exerts strong repressing activity, the expression of over 30 genes was significantly increased upon removal of codY. The differentially expressed genes included those predominantly involved in amino acid transport and metabolism. In addition, several genes belonging to other functional categories were derepressed, stressing the pleiotropic role of CodY. Scrutinizing the transcriptome data with bioinformatics tools revealed the presence of a novel over-represented motif in the upstream regions of several of the genes derepressed in L. lactis MG1363DeltacodY. Evidence is presented that this 15-bp cis-sequence, AATTTTCWGAAAATT, serves as a high affinity binding site for CodY, as shown by electrophoretic mobility shift assays and DNase I footprinting analyses. The presence of this CodY-box is sufficient to evoke CodY-mediated regulation in vivo. A copy of this motif is also present in the upstream region of codY itself. It is shown that CodY regulates its own synthesis and requires the CodY-box and branched-chain amino acids to interact with its promoter.
乳酸乳球菌MG1363的CodY是一种转录调节因子，可抑制几种编码蛋白水解系统蛋白质的基因的表达。利用DNA微阵列分析比较了乳酸乳球菌MG1363和一个codY发生突变的同基因菌株的表达谱，以确定CodY调控子。在富含肽的培养基和指数生长的细胞中，CodY发挥强大的抑制活性，去除codY后，30多个基因的表达显著增加。差异表达的基因包括那些主要参与氨基酸转运和代谢的基因。此外，其他功能类别的几个基因也被去抑制，这突出了CodY的多效性作用。用生物信息学工具仔细研究转录组数据发现，在乳酸乳球菌MG1363ΔcodY中被去抑制的几个基因的上游区域存在一个新的过度代表的基序。电泳迁移率变动分析和DNase I足迹分析表明，这个15个碱基的顺式序列AATTTTCWGAAAATT作为CodY的高亲和力结合位点。这个CodY框的存在足以在体内引发CodY介导的调控。这个基序的一个拷贝也存在于codY自身的上游区域。结果表明，CodY调节其自身的合成，并且需要CodY框和支链氨基酸与其启动子相互作用。
