Berho Nathalie, Duret Sybille, Renaudin Joël
UMR 1090 Génomique Développement et Pouvoir Pathogène, INRA et Université de Bordeaux 2, IBVM, Centre INRA de Bordeaux, 71 avenue Edouard Bourlaux, BP 81, 33883 Villenave d'Ornon Cedex, France.
Microbiology (Reading). 2006 Mar;152(Pt 3):873-886. doi: 10.1099/mic.0.28541-0.
In the plant-pathogenic mollicute Spiroplasma citri, spiralin is the major lipoprotein at the cell surface and is thought to be one of the components involved in the interactions of the spiroplasma with its insect vector. With the aim of identifying surface proteins other than spiralin, monoclonal antibodies (mAbs) were produced by immunization of mice with the spiralin-defective S. citri mutant GII3-9a2. mAb 10G3 was found to react with several polypeptides of 43-47 and 80-95 kDa, all of which were detected in the detergent phase after Triton X-114 partitioning of proteins. Mass spectrometry (MALDI-TOF) analyses of the two major polypeptides P47 and P80 of GII3-9a2, reacting with mAb 10G3, revealed that P47 was a processed product and represented the C-terminal moiety of P80. Search for sequence homologies revealed that P80 shared strong similarities with the S. citri adhesion-related protein P89 (Sarp1) of S. citri BR3, and is one (named Scarp4a) of the eight Scarps encoded by the S. citri GII-3 genome. The eight scarp genes are carried by plasmids pSci1-5. Western immunoblotting of proteins with mAb 10G3 revealed that, in contrast to the insect-transmissible S. citri strain GII-3, the non-insect-transmissible strains ASP-1, R8A2 and 44 did not express Scarps. Southern blot hybridization experiments indicated that these strains possessed no scarp genes, and did not carry plasmids pSci1-5. However, S. citri strain GII3-5, lacking pSci5, was still efficiently transmitted, showing that, in the genetic background of S. citri GII-3, the pSci5-encoded genes, and in particular scarp2b, 3b and 5a, are not essential for insect transmission. Whether plasmid-encoded genes are involved in transmission of S. citri by its leafhopper vector remains to be determined.
在植物致病性柔膜菌柑橘螺原体中，螺旋蛋白是细胞表面的主要脂蛋白，被认为是参与螺原体与其昆虫载体相互作用的成分之一。为了鉴定除螺旋蛋白之外的表面蛋白，用螺旋蛋白缺陷型柑橘螺原体突变体GII3 - 9a2免疫小鼠制备了单克隆抗体（mAb）。发现单克隆抗体10G3能与43 - 47 kDa和80 - 95 kDa的几种多肽发生反应，在蛋白质经Triton X - 114分配后的去污剂相中均能检测到这些多肽。对与单克隆抗体10G3反应的GII3 - 9a2的两种主要多肽P47和P80进行质谱（MALDI - TOF）分析，结果表明P47是一种加工产物，代表P80的C末端部分。序列同源性搜索显示，P80与柑橘螺原体BR3的柑橘螺原体黏附相关蛋白P89（Sarp1）有很强的相似性，并且是柑橘螺原体GII - 3基因组编码的八个Sarp（Scarp）之一（命名为Scarp4a）。这八个scarp基因由质粒pSci1 - 5携带。用单克隆抗体10G3对蛋白质进行Western免疫印迹分析表明，与可经昆虫传播的柑橘螺原体菌株GII - 3不同，不可经昆虫传播的菌株ASP - 1、R8A2和44不表达Scarp。Southern印迹杂交实验表明，这些菌株没有scarp基因，也不携带质粒pSci1 - 5。然而，缺乏pSci5的柑橘螺原体菌株GII3 - 5仍然能高效传播，这表明在柑橘螺原体GII - 3的遗传背景下，pSci5编码的基因，特别是scarp2b、3b和5a，对于昆虫传播并非必不可少。质粒编码的基因是否参与柑橘螺原体通过其叶蝉载体的传播仍有待确定。
