橙色粘球菌四型菌毛膜复合物的发现

doi:10.3976/j.issn.1002-4026.2014.06.004

摘要/Abstract

摘要： 细菌四型菌毛 (T4P) 不仅在生物膜形成、DNA吸收和侵染致病等生物过程中发挥着重要的作用，同样也能够介导固体界面的滑动运动。尽管T4P具有非常重要的作用，但橙色粘球菌T4P系统中蛋白间组装机制的研究仍处于初级阶段。T4P系统pilN/O/P/Q 单基因的敲除与回补实验显示敲除菌株导致橙色粘球菌群体运动完全丧失，而pilN/O/P/Q相应基因回补则恢复突变株群体运动。PilN/O/P/Q蛋白通过酵母双杂交（Y2H）检测蛋白间的相互作用发现，定位于周质空间的PilP不仅与外膜蛋白PilQ的C末端相互作用，也与内膜蛋白PilO相互作用。遗传学与酵母双杂实验结果显示着PilP蛋白是联通内外膜的桥梁，暗示着PilN/O/P/Q蛋白形成跨越内膜、细胞周质和外膜的整合复合物结构，并在菌毛延伸和收缩过程中发挥着重要的作用，为阐明橙色粘球菌菌毛的延伸与收缩的分子机制奠定坚实基础。

关键词: 四型菌毛, 酵母双杂交, 橙色粘球菌, 群体运动

Abstract: Bacterial type IV pili or pilus (T4P) not only plays an important role in such biological processes as biofilm formation, DNA intake and bacterial pathogenesis but also enables bacterial motility on solid surfaces. Despite its importance, we know very little about the assembly mechanism among T4P proteins of Myxococcus xanthus.Deletion and supplement experiments of single gene pilN/O/P/Q demonstrate that strain removal causes Myxococcus xanthus colony to totally lose social motility, but strain supplement can restore their social motility. We employed yeast two hybrid (Y2H) to detect the interactions among M. xanthus T4P proteins. We discovered that PilP at periplasm interacted not only with C end of outerfilm protein PilQ, but also with innerfilm protein PilO. Results of yeast two hybrid (Y2H) and genetics experiments show that protein PilP is the bridge connecting innerfilm and outerfilm. This indicates that proteins PilN/O/P/Q may form a total complex including periplasm, innerfilm and outerfilm, and play an important role in pilus extension and contraction. Our research lays a solid foundation for the exploration of molecular mechanism of the extension and contraction of Myxococcus xanthus pilus.

Key words: Myxococcus xanthus, yeast two hybrid, type IV pili, social motility

中图分类号:

R378.1

李成云，李红梅，李纪顺，杨合同,Bazhanov DMITRY. 橙色粘球菌四型菌毛膜复合物的发现[J]. 山东科学, 2014, 27(6): 22-30.

LI Cheng-yun,LI Hong-mei,LI Ji-shun,YANG He-tong,Bazhanov DMITRY. Discovery of type IV pilus membrane compound of Myxococcus xanthus[J]. SHANDONG SCIENCE, 2014, 27(6): 22-30.

参考文献

［1］REICHENBACH H.The ecology of the myxobacteria ［J］. Environmental microbiology,1999,1（1）: 15-21. ［2］SHIMKETS L J. Intercellular signaling during fruitingbody development of Myxococcus xanthus ［J］. Annual review of microbiology, 1999,53:525-549. ［3］KAISER D. Coupling cell movement to multicellular development in myxobacteria ［J］. Microbiology, 2003,1（1）: 45-54. ［4］KAISER D. A microbial genetic journey ［J］. Annual review of microbiology, 2006, 60: 1-25. ［5］HODGKIN KAISERD. Genetics of gliding motility in Myxococcus xanthus (Myxobacterales): Two gene systems control movement ［J］. Molecular and General Genetics 1979, 171（1432-1874）:177-191. ［6］SHI W, ZUSMAN DR. The two motility systems of Myxococcus xanthus show different selective advantages on various surfaces ［J］. Proceedings of the National Academy of Sciences of the United States of America, 1993,90（8）:3378-3382. ［7］SHIMKETS L J. Correlation of energydependent cell cohesion with social motility in Myxococcus xanthus［J］. Journal of bacteriology, 1986,166（3）: 837-841. ［8］SHIMKETS L J. Role of cell cohesion in Myxococcus xanthus fruiting body formation ［J］. Journal of Bacteriology, 1986,166（3）:842-848. ［9］BOWDEN M G, KAPLAN H B. The Myxococcus xanthus lipopolysaccharide Oantigen is required for social motility and multicellular development ［J］. Molecular microbiology, 1998,30（2）:275-284. ［10］KAISER D. Social gliding is correlated with the presence of pili in Myxococcus xanthus ［J］. Proceedings of the National Academy of Sciences of the United States of America, 1979,76（11）:5952-5956. ［11］WU S S, KAISER D. Genetic and functional evidence that Type IV pili are required for social gliding motility in Myxococcus xanthus ［J］. Molecular microbiology, 1995,18（3）: 547-558. ［12］WALL D, KAISER D. Type IV pili and cell motility ［J］. Molecular microbiology, 1999,32（1）:1-10. ［13］PELICIC V. Type IV pili:e pluribus unum? ［J］. Molecular microbiology, 2008,68（4）:827-837. ［14］CLAUSEN M, JAKOVLJEVIC V, SOGAARDANDERSEN L, et al. Highforce generation is a conserved property of type IV pilus systems ［J］. Journal of bacteriology, 2009,191（14）:4633-4638. ［15］MATTICK JS. Type IV pili and twitching motility ［J］. Annual review of microbiology, 2002, 56: 289-314. ［16］WEIMER R M, CREIGHTON C, STASSINOPOULOS A,et al. A chaperone in the HSP70 family controls production of extracellular fibrils in Myxococcus xanthus ［J］. Journal of bacteriology,1998,180（20）:5357-5368. ［17］YANG Z, MA X, TONG L,et al.Myxococcus xanthus dif genes are required for biogenesis of cell surface fibrils essential for social gliding motility ［J］. Journal of bacteriology, 2000,182（20）:5793-5798. ［18］WANG B, HU W, LIU H,et al. Adaptation of salttolerant Myxococcus strains and their motility systems to the ocean conditions ［J］. Microbial ecology, 2007,54（1）:43-51. ［19］AYERS M, SAMPALEANU L M, TAMMAM S,et al. PilM/N/O/P proteins form an inner membrane complex that affects the stability of the Pseudomonas aeruginosa type IV pilus secretin ［J］. Journal of molecular biology, 2009,394（1）:128-142. ［20］KAISER D. Social gliding is correlated with the presence of pili in Myxococcus xanthus［J］. Proc Natl Acad Sci USA, 1979,76（11）:5952-5956. ［21］WALL D, KOLENBRANDER PE, KAISER D. The Myxococcus xanthus pilQ (sglA) gene encodes a secretin homolog required for type IV pilus biogenesis, social motility, and development ［J］. Journal of bacteriology, 1999,181（1）:24-33. ［22］WU S S,WU J, KAISER D. The Myxococcus xanthus pilT locus is required for social gliding motility although pili are still produced ［J］. Molecular microbiology, 1997, 23（1）:109-121. ［23］JULIEN B,KAISER AD, GARZA A. Spatial control of cell differentiation in Myxococcus xanthus［J］. Proc Natl Acad Sci USA, 2000,97（16）: 9098-9103. ［24］BLACK W P, XU Q, CADIEUX C L,et al. Isolation and characterization of a suppressor mutation that restores Myxococcus xanthus exopolysaccharide production ［J］. Microbiology, 2009,155（Pt11）:3599-3610. ［25］SESHADRI R, JOSEPH SW, CHOPRA A K,et al.Genome sequence of Aeromonas hydrophila ATCC 7966T:Jack of all trades ［J］. J Bacteriol, 2006,188（23）: 8272-8282. ［26］UEKI T, INOUYE S, INOUYE M. Positivenegative KG cassettes for construction of multigene deletions using a single drug marker ［J］. Gene, 1996, 183（12）: 153-157. ［27］BLACK W P, YANG Z. Myxococcus xanthus chemotaxis homologs DifD and DifG negatively regulate fibril polysaccharide production ［J］. Journal of bacteriology, 2004, 186（4）: 1001-1008. ［28］GIETZ R D,WOODS R A. Screening for proteinprotein interactions in the yeast twohybrid system ［J］. Methods in molecular biology, 2002, 185: 471-486. ［29］JAMES P, HALLADAY J, CRAIG EA. Genomic libraries and a host strain designed for highly efficient twohybrid selection in yeast ［J］. Genetics, 1996, 144（4）:1425-1436. ［30］KARUPPIAH V, DERRICK J P. Structure of the PilMPilN inner membrane type IV pilus biogenesis complex from Thermus thermophilus［J］. The Journal of biological chemistry, 2011,286（27）:24434-24442. ［31］TAMMAM S, SAMPALEANU LM, KOO J,et al. PilM/N/O/P/Q from the Pseudomonas aeruginosa type IV pilus system form a transenvelope protein interaction network that interacts with PilA ［J］. Journal of bacteriology,2013,195（10）: 2126-2135. ［32］GILTNER C L, NGUYEN Y, BURROWS LL. Type IV pilin proteins: Versatile molecular modules ［J］. Microbiology and molecular biology reviews, 2012,76（4）:740-772. ［33］BERRY J L,PHELAN M M, COLLINS RF,et al. Structure and assembly of a transperiplasmic channel for type IV pili in Neisseria meningitidis［J］. PLoS pathogens, 2012,8（9）:e1002923.