Wu Y, Egerton G, Ball A, Tanguay R M, Bianco A E
Division of Molecular Biology and Immunology, Liverpool School of Tropical Medicine, Liverpool, UK.
Mol Biochem Parasitol. 2000 Apr 15;107(2):155-68. doi: 10.1016/s0166-6851(99)00227-3.
Chaperonin 60 (cpn60) belongs to the group of ubiquitous molecular chaperones that comprise the heat shock proteins, nucleoplasmins and chaperonins. Antibodies to recombinant CPN60 from humans was used to screen a cDNA library of Onchocerca volvulus and antigen-positive clones were selected. Sequencing of the DNA inserts confirmed their identity as cpn60 transcripts. These are distinct from a cpn60 sequence recorded previously from O. volvulus (GenBank accession number Y09416) that appears to be of endobacterial origin, rather than derived from the parasite itself. The full-length sequence of the cDNA (designated Ov-cpn60) codes for a protein of 64.3kDa (598 amino acid residues) and shares significant identity with homologous gene products from Caenorhabditis elegans (72%), humans (69%), yeast (53%) and Escherichia coli (50%). The endobacterial and parasite sequences are 41% conserved. Ov-CPN60 migrates with an apparent molecular mass of 65kDa on SDS-PAGE and is present in all life-cycle stages, as determined by immunoblotting with rabbit antibodies raised against the recombinant protein. Immunogold electron microscopy identified the protein within mitochondria, as expected, but also in extra-mitochondrial sites, including inclusion bodies of the glandular oesophagus (in infective larvae), the uterine wall, cytosol of developing spermatids, and the hypodermis and cuticle. Endobacteria were also labelled, indicating cross-reactivity between CPN60 from the parasite and its intracellular symbiont. In human infections, serum antibodies to Ov-CPN60 were present in only 11% of cases from Ecuador, but in 81-89% of subjects in three separate foci from West Africa. There was no relationship between antibody levels and age, sex, or infection intensity, and no consistent association between the serological response and immune status. An evaluation of antibody specificities in individual sera revealed a mixture of parasite-specific and host crossreactive anti-CPN60 antibodies, the ratio of which varied amongst geographic areas. It is concluded that antibody responses to Ov-CPN60 are unlikely to contribute either to host protection or pathology in onchocerciasis.
伴侣蛋白60（cpn60）属于普遍存在的分子伴侣家族，该家族包括热休克蛋白、核质蛋白和伴侣蛋白。用人重组CPN60抗体筛选盘尾丝虫的cDNA文库，并选择抗原阳性克隆。对DNA插入片段进行测序，证实它们是cpn60转录本。这些转录本与先前从盘尾丝虫记录的cpn60序列（GenBank登录号Y09416）不同，后者似乎源自内共生菌，而非来自寄生虫本身。cDNA的全长序列（命名为Ov-cpn60）编码一个64.3kDa的蛋白质（598个氨基酸残基），与秀丽隐杆线虫（72%）、人类（69%）、酵母（53%）和大肠杆菌（50%）的同源基因产物具有显著同源性。内共生菌和寄生虫的序列保守性为41%。Ov-CPN60在SDS-PAGE上的表观分子量为65kDa，通过用针对重组蛋白产生的兔抗体进行免疫印迹分析确定其存在于所有生命周期阶段。免疫金电子显微镜观察发现，正如预期的那样，该蛋白存在于线粒体中，但也存在于线粒体外部位，包括腺食管的包涵体（感染性幼虫中）、子宫壁、发育中的精子细胞的细胞质以及皮下组织和角质层。内共生菌也被标记，表明寄生虫的CPN60与其细胞内共生体之间存在交叉反应。在人类感染中，厄瓜多尔仅有11%的病例血清中存在抗Ov-CPN60抗体，但在西非三个不同疫源地的81%-89%的受试者中存在该抗体。抗体水平与年龄、性别或感染强度之间没有关系，血清学反应与免疫状态之间也没有一致的关联。对个体血清中抗体特异性的评估显示，存在寄生虫特异性和宿主交叉反应性抗CPN60抗体的混合物，其比例在不同地理区域有所不同。得出的结论是，抗Ov-CPN60抗体反应不太可能对盘尾丝虫病的宿主保护或病理状况产生影响。
