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Construction, detection and microarray analysis on Shigella dysenteriae A1 IroN, ShuA single, double mutants

Science china(Life Sciences) 2006年第3期49卷 251-258页

作者： bin wen, liu moqing, peng junping, sun lilian, xu xingye, zhang jinghai& jin qi shengyang pharmaceutical university, school of pharmaceutical engineering, shengyang 110016, china State Key Laboratory for Molecular Virology and Genetic engineering, Beijing 100176, china shengyang pharmaceutical university School of Pharmaceutical Engineering Shengyang 110016 China State Key Laboratory for Molecular Virology and Genetic engineering Beijing 100176 China State Key Laboratory for Molecular Virology and Genetic engineering Beijing 100176 China shengyang pharmaceutical university School of Pharmaceutical Engineering Shengyang 110016 China

In this study, we constructed single mutants MTS-1, MTS-2 of IroN and ShuA gene and double mutant MTS of them in Shigella dysenteriae A1 strain 51197 by insert and absence. The functional detection of every mutant was performed at the level of culture medium and cell experiment. The gene expression profiles of the mutants and the wild-type strains under iron- enriched and iron-limited conditions were analyzed by the SD51197 whole genomic microarray. The results showed that all the mutants grew obviously less well than the wild-type strains in L broth appending iron chelator DIP. The addition of iron to the cultures can stimulate the growth of mutants back to wild-type levels. In either the experiments on the ability of intracellular multiplication or the cell-to-cell spread in HeLa and U937 cell lines, mutants showed no obvious change in virulence compared with the parental strain SD51197. However when DIP was added to the cultured HeLa cells, the ability of intracellular multiplication of MTS-1, MTS-2, MTS has reduced about 23.4%, 25.2%, 43.6% respectively. The analysis of expression profiles under the iron-limited condition showed that the mutants were more sensitive for the changes of iron deficiency than the wild-type strains, many genes have been altered. Up-regulated genes mainly involved genes of transcription, coenzyme metabolism, amino acid transport and metabolism, and unknown functional genes, while down-regulated genes mainly involved genes of energy and carbohydrate metabolism and unknown function genes; the expression levels of known iron-transport associated genes generally showed up-regulated. The results demonstrated that iron-transport associated genes IroN, ShuA were likely to have some effects on the virulence and growth of S. dysenteriae.

关键词： Shigella dysenteriae A1, gene knockout, functional detection, microarray analysis.

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Genomic compositions and phylogenetic analysis of Shigella boydii subgroup

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Science china(Life Sciences) 2006年第1期49卷 46-52页

作者： WANG jing zhang Xiaobing peng junping YANG E bin wen YANG Jian DONG Jie sun lilian xu xingye jin qi State Key Laboratory for Molecular Virology and Genetic engineering Beijing 100176China school of Medicine Xi’an Jiaotong UniversityXi’an 710061China school of pharmaceutical engineering Shengyang Pharmaceutical UniversityShengyang 110016China

Comparative Genomic Hybridization (CGH) microarray analysis was used tocompare the genomic compositions of all eighteen Shigella boydii serotype representative *** results indicated the genomic ''backbone'' of this subgroup contained 2552 ORFs homologous tononpatho-genic E. coli K12. Compared with the genome of K12199 ORFs were found to be absent in allS. boydii serotype representatives, including mainly outer membrane protein genes and O-antigenbiosynthesis genes. Yet the specific ORFs of S. boydii subgroup contained basically bacteriophagegenes and the function unknown (FUN) genes. Some iron metabolism, transport and type Ⅱ secretionsystem related genes were found in most representative strains. According to the CGH phylogeneticanalysis, the eighteen S. boydii serotype representatives were divided into four groups, in whichserotype C13 strain was remarkably distinguished from the other serotype strains. This groupingresult corresponded to the distribution of some metabolism related genes. Furthermore, the analysisof genome backbone genes, specific genes, and the phylogenetic trees allowed us to discover theevolution laws of S. boydii and to find out important clues to pathogenesis research, vaccinationand the therapeutic medicine development.

关键词： S.boydii comparative genomic hybridization(CGH) strain-specific ORFs evolution

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