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Scientific
Publications - Work Done by Microbiology Reader
P.C.Y. Lau, and D.G. Cvitkovitch, Identification and functional analysis of two-component signal transduction systems in Streptococcus mutans, The IADR/AADR/CADR 80th General Session (March 6-9, 2002), San Diego, California, USA ABSTRACT Two-component signal transduction systems (TCSTS) are present
across divergent bacterial species and function importantly in both
environmental sensing and cell-cell communication. Each system is composed of a
histidine kinase (HK) sensor protein coupled to a cognate response regulator
(RR) element. Objectives: The present study was carried out to identify
the complete complement of TCSTS in the oral pathogen Streptococcus mutans,
and to determine the functional role of each component gene product in these
systems. Methods: Published TCSTS seqeunce data from the related organism
Streptococcus pneumoniae were compared with the S. mutans genome
database available from the University of Oklahoma to identify putative TCSTS. A
rapid PCR-based gene deletion strategy involving restriction-ligation and
allelic replacement was employed for the systematic creation of knockout mutant
strains, each with a different HK or RR component deleted. An automated, high
throughput Bioscreen C Microbiology Analyzer was employed to assay the deletion
mutants derived from wild-type NG8 or UA159 strains for growth in different
stress conditions, including sodium chloride, sodium dodecyl sulphate, ethanol,
hydrogen peroxide, acid, etc. Results: Thirteen HK/RR pairs, believed to
represent the complete complement of TCSTS, were identified and located in the S.
mutans genome. Specific primers were designed to individually delete 24
genes of 12 TCSTS by PCR ligation mutagenesis (one system consisted of comD
and comE, genes that were previously deleted via insertion-duplication
mutagenesis). Comparison of growth data of the 26 mutants with the wild-type
strains indicated the physiological role of the TCSTS genes in cellular
function. Conclusions: Using a combination of bioinformatics tools,
molecular biology techniques and high-throughput screening, the insight into
functional significance of all 13 TCSTS in S. mutans can be elucidated.
Supported by PHS grant DE 013230 from the NIDCR.
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