变异链球菌LuxS缺陷株甲基循环通路的恢复构建

上海口腔医学 ›› 2014, Vol. 23 ›› Issue (4): 385-390.

• 基础研究 • 下一篇

变异链球菌LuxS缺陷株甲基循环通路的恢复构建

王玉霞, 高丽, 江文欣, 马瑞, 唐子圣, 朱彩莲, 何智妍, 黄正蔚

上海交通大学医学院附属第九人民医院·口腔医学院牙体牙髓病科,上海市口腔医学重点实验室,上海 200011

收稿日期:2014-01-21 出版日期:2014-08-20 发布日期:2014-10-20
通讯作者: 黄正蔚,Fax:021-63135412,E-mail:huang_zhengwei@hotmail.com
作者简介:王玉霞(1987- ), 女, 学士, E-mail:wxy870111@163.com
基金资助:
国家自然科学基金(81070826,81371143,81300866); 上海市启明星计划(12QH1401400)

Complementation and function of methyl-metabolic pathway in Streptococcus mutans luxS null strain

WANG Yu-xia, GAO Li, JIANG Wen-xin, MA Rui, TANG Zi-sheng, ZHU Cai-lian, HE Zi-yan, HUANG Zheng-wei

Department of Endodontics, Ninth People’s Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine; Shanghai Key Laboratory of Stomatology. Shanghai 200011, China

Received:2014-01-21 Online:2014-08-20 Published:2014-10-20
Supported by:
Supported by National Natural Science Foundation of China (81070826, 81371143, 81300866) and partly supported by Shanghai Rising-Star Program (12QH1401400).

摘要/Abstract

摘要： 目的:以变异链球菌LuxS缺陷株为模型,构建其密度感应缺陷的甲基循环恢复株。方法:以LuxS为阳性表达对照,通过在变异链球菌LuxS缺陷株中异源表达SahH,实现其代谢通路的恢复构建。通过Western 印迹、反转录PCR和信号分子AI-2分泌检测,验证目的蛋白的表达。采用SAS8.0软件包对数据进行统计学分析。结果:Western 印迹检测2种蛋白在大肠杆菌中均得到正确表达,验证了克隆载体的正常表达能力;反转录PCR验证了变异链球菌LuxS缺陷株中luxS及sahH mRNA的存在;发光实验证实LuxS阳性表达对照株AI-2的分泌能力恢复。结论:成功构建变异链球菌LuxS缺陷的代谢恢复株,为探讨LuxS的调控机制提供了分子生物学工具及实验对照。

关键词: 变异链球菌, 生物膜, 密度感应, LuxS, SahH

Abstract: PURPOSE: To complement the activated methyl cycle (AMC) pathway at an AI-2 defect background in Streptococcus mutans (S. mutans) luxS null strain. METHODS: A sahH gene was amplified from Pseudomonas aeruginosa and introduced into the S. mutans luxS null strain to complement the methyl-metabolic disruption at an AI-2 defect background. Western blot, reverse-transcription PCR and AI-2 bioassay were performed to confirm the heterogenous expression of SahH in S. mutans luxS null strain. The data was statistically analyzed by SAS8.0 software package. RESULTS: LuxS and SahH were detected to express in Escherichia coli BL21 as well as their mRNA were confirmed to be successfully transcribed in S. mutans luxS null strain. AI-2 production was found in wide type S. mutans and its luxS-introduced luxS null strain but not found in the luxS null strain and its sahH and empty plasmid-introduced strains. CONCLUSIONS: A new S. mutans derivative with the AMC pathway complements while the AI-2 defect is constructed.

Key words: Streptococcus mutans, Biofilm, Quorum sensing, LuxS, SahH

中图分类号:

R780.2

王玉霞, 高丽, 江文欣, 马瑞, 唐子圣, 朱彩莲, 何智妍, 黄正蔚. 变异链球菌LuxS缺陷株甲基循环通路的恢复构建[J]. 上海口腔医学, 2014, 23(4): 385-390.

WANG Yu-xia, GAO Li, JIANG Wen-xin, MA Rui, TANG Zi-sheng, ZHU Cai-lian, HE Zi-yan, HUANG Zheng-wei. Complementation and function of methyl-metabolic pathway in Streptococcus mutans luxS null strain[J]. Shanghai Journal of Stomatology, 2014, 23(4): 385-390.

参考文献

[1] Waters CM, Bassler BL. Quorum sensing: cell-to-cell communication in bacteria [J]. Annu Rev Cell Dev Biol, 2005, 21: 319-346.
[2] Blehert DS, Palmer RJ Jr, Xavier JB, et al. Autoinducer 2 production by Streptococcus gordonii DL1 and the biofilm phenotype of a luxS mutant are influenced by nutritional conditions [J]. J Bacteriol, 2003, 185(16): 4851-4860.
[3] Camilli A, Bassler BL. Bacterial small-molecule signaling pathways [J]. Science, 2006, 311(5764): 1113-1116.
[4] Parveen N, Cornell KA. Methylthioadenosine/S-adenosylhomocysteine nucleosidase, a critical enzyme for bacterial metabolism [J]. Mol Microbiol, 2011, 79(1): 7-20.
[5] Cataldi TR, Bianco G, Abate S, et al. Analysis of S-adenosylmethionine and related sulfur metabolites in bacterial isolates of Pseudomonas aeruginosa (BAA-47) by liquid chromatography/electrospray ionization coupled to a hybrid linear quadrupole ion trap and Fourier transform ion cyclotron resonance mass spectrometry [J]. Rapid Commun Mass Spectrom, 2009, 23(21): 3465-3477.
[6] Wen ZT, Burne RA. LuxS-mediated signaling in Streptococcus mutans is involved in regulation of acid and oxidative stress tolerance and biofilm formation [J]. J Bacteriol, 2004, 186(9): 2682-2691.
[7] Yoshida A, Ansai T, Takehara T, et al. LuxS-based signaling affects Streptococcus mutans biofilm formation [J]. Appl Environ Microbiol, 2005, 71(5): 2372-2380.
[8] Sztajer H, Lemme A, Vilchez R, et al. Autoinducer-2-regulated genes in Streptococcus mutans UA159 and global metabolic effect of the luxS mutation [J]. J Bacteriol, 2008, 190(1): 401-415.
[9] Huang Z, Meric G, Liu Z, et al. luxS-based quorum-sensing signaling affects Biofilm formation in Streptococcus mutans[J]. J Mol Microbiol Biotechnol, 2009, 17(1): 12-19.
[10] 王倩, 唐子圣, 马瑞, 等. 变异链球菌luxS基因对多糖基质代谢的影响 [J]. 上海口腔医学, 2011, 20(1): 6-9.
[11] Biswas I, Jha JK, Fromm N. Shuttle expression plasmids for genetic studies in Streptococcus mutans [J]. Microbiology, 2008, 154(Pt 8): 2275-2282.
[12] Han X, Bai H, Liu L, et al. Cloning and expression of luxS and pfs and in vitro biosynthesis autoinducer 2 of avian pathogenic Escherichia coli from Anhui Province [J]. Wei Sheng Wu Xue Bao, 2012, 52(9): 1167-1172.
[13] Biswas I, Drake L, Johnson S, et al. Unmarked gene modification in Streptococcus mutans by a cotransformation strategy with a thermosensitive plasmid[J]. BioTechniques,2007,42(4):487-490.
[14] Merritt J, Qi F, Goodman SD, et al. Mutation of luxS affects biofilm formation in Streptococcus mutans[J]. Infect Immun, 2003, 71(4): 1972-1979.
[15] Redanz S, Standar K, Podbielski A, et al. Heterologous expression of sahH reveals that biofilm formation is autoinducer-2-independent in Streptococcus sanguinis but is associated with an intact activated methionine cycle[J]. J Biol Chem,2012,287(43): 36111-36122.
[16] Sun Z, He X, Brancaccio VF, et al. Bifidobacteria exhibit LuxS-dependent autoinducer 2 activity and biofilm formation[J]. PLoS One, 2014, 9(2): e88260.
[17] Bassler BL, Greenberg EP, Stevens AM. Cross-species induction of luminescence in the quorum-sensing bacterium Vibrio harveyi [J]. J Bacteriol, 1997, 179(12): 4043-4045.
[18] Xavier KB, Bassler BL. Interference with AI-2-mediated bacterial cell-cell communication [J]. Nature, 2005, 437(7059): 750-753.

变异链球菌LuxS缺陷株甲基循环通路的恢复构建

Complementation and function of methyl-metabolic pathway in Streptococcus mutans luxS null strain

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