A function for a specific zinc metalloprotease of African trypanosomes.
A function for a specific zinc metalloprotease of African trypanosomes.
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DOI：
10.1371/journal.ppat.0030150
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发表时间：
2007-10-19
期刊：
影响因子：
6.7
通讯作者：
Donelson JE
中科院分区：
文献类型：
--
作者：
Grandgenett PM;Otsu K;Wilson HR;Wilson ME;Donelson JE
布氏锥虫基因组编码三组锌金属蛋白酶，每组与利什曼原虫的主要表面蛋白酶（MSP，也称为GP 63）具有约30%的氨基酸同一性。这些蛋白酶之一，TbMSP-B，是由四个几乎相同的，串联基因在血液和前环锥虫转录编码。早期的工作表明，针对TbMSP-B的RNA干扰阻止了重组变体表面糖蛋白（VSG）从原环锥虫中释放。在这里，我们使用基因缺失来显示TbMSP-B和磷脂酶C（GPI-PLC）在血流锥虫分化为前环形式期间协同作用以去除天然VSG。当四个串联TbMSP-B基因从两个染色体等位基因中删除时，血流B −/−锥虫仍然可以分化为前环形式，但与野生型生物体的分化相比，VSG的去除更慢，并且是以非截短的形式。同样，当单拷贝GPI-PLC基因的两个等位基因都被删除时，血流PLC −/−细胞仍然可以分化。然而，当TbMSP-B和GPI-PLC的所有基因都从二倍体基因组中删除时，血流B-/-PLC-/-锥虫在分化培养基中不增殖，60%的VSG保留在细胞表面。半胱氨酸蛋白酶抑制剂不影响这一结果。这些发现表明，在从血流分化为前环形式的过程中去除60%的VSG是由于GPI-PLC和TbMSP-B的协同活性。非洲锥虫引起昏睡病，这是非洲人类和牲畜的致命疾病。在它们的生命周期中，这些原生动物寄生虫在哺乳动物和采采蝇的血液中循环。它们的两个主要发育阶段是采采蝇的血流型和原循环型。血流锥虫通过周期性地转换其主要表面蛋白，即变体表面糖蛋白（VSG）来阻碍宿主的免疫反应。当采采蝇摄取血流锥虫时，它们必须迅速摆脱VSG，并将其替换为一种不相关的不变蛋白质，该蛋白质更适合它们作为苍蝇中肠中的前循环生物体生存。在这里，我们研究锥虫使用的机制，以消除VSG在其分化过程中，从血流中的培养物中的前循环形式。我们从锥虫基因组中删除了一种锥虫蛋白酶的基因，发现血流锥虫仍然可以分化为前环形式，但VSG去除减少。删除磷脂酶的基因也有类似的效果--它们仍然可以分化，但VSG的去除受损。当蛋白酶和磷脂酶的基因都被删除时，血流锥虫不能分化为前环形式，它们在表面保留了约60%的VSG，并且它们在分化培养基中死亡。这些结果突出了这两种酶在分化过程中的协同作用。
The Trypanosoma brucei genome encodes three groups of zinc metalloproteases, each of which contains ∼30% amino acid identity with the major surface protease (MSP, also called GP63) of Leishmania. One of these proteases, TbMSP-B, is encoded by four nearly identical, tandem genes transcribed in both bloodstream and procyclic trypanosomes. Earlier work showed that RNA interference against TbMSP-B prevents release of a recombinant variant surface glycoprotein (VSG) from procyclic trypanosomes. Here, we used gene deletions to show that TbMSP-B and a phospholipase C (GPI-PLC) act in concert to remove native VSG during differentiation of bloodstream trypanosomes to procyclic form. When the four tandem TbMSP-B genes were deleted from both chromosomal alleles, bloodstream B −/− trypanosomes could still differentiate to procyclic form, but VSG was removed more slowly and in a non-truncated form compared to differentiation of wild-type organisms. Similarly, when both alleles of the single-copy GPI-PLC gene were deleted, bloodstream PLC −/− cells could still differentiate. However, when all the genes for both TbMSP-B and GPI-PLC were deleted from the diploid genome, the bloodstream B −/− PLC −/− trypanosomes did not proliferate in the differentiation medium, and 60% of the VSG remained on the cell surface. Inhibitors of cysteine proteases did not affect this result. These findings demonstrate that removal of 60% of the VSG during differentiation from bloodstream to procyclic form is due to the synergistic activities of GPI-PLC and TbMSP-B. African trypanosomes cause sleeping sickness, a fatal disease of humans and livestock in Africa. During their life cycle, these protozoan parasites cycle between the bloodstream of mammals and tsetse flies. Their two main developmental stages are the bloodstream form and the procyclic form in the tsetse fly. Bloodstream trypanosomes thwart their host's immune response by periodically switching their major surface protein, the variant surface glycoprotein (VSG). When bloodstream trypanosomes are ingested by a tsetse fly, they must quickly shed the VSG and replace it with an unrelated invariant protein more suited to their survival as procyclic organisms in the fly midgut. Here, we examine the mechanisms used by trypanosomes to remove the VSG during their differentiation from bloodstream to procyclic form in culture. We deleted the genes for one of the trypanosome's protease enzymes from the trypanosome genome and found that bloodstream trypanosomes could still differentiate to the procyclic form, but VSG removal was diminished. Deleting the genes for a phospholipase enzyme had a similar effect—they could still differentiate but VSG removal was impaired. When the genes for both the protease and the phospholipase were deleted, bloodstream trypanosomes could not differentiate to the procyclic form, they retained about 60% of the VSG on their surface, and they died in the differentiation medium. These results highlight the synergistic roles of these two enzymes in the differentiation process.
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影响因子：
5.7
作者：
Schlagenhauf, E;Etges, R;Metcalf, P
通讯作者：
Metcalf, P
影响因子：
4.8
作者：
ROBERTS, SC;WILSON, ME;DONELSON, JE
通讯作者：
DONELSON, JE
影响因子：
7.8
作者：
Webb, H;Carnall, N;Carrington, M
通讯作者：
Carrington, M
影响因子：
2.9
作者：
BOUVIER, J;SCHNEIDER, P;BORDIER, C
通讯作者：
BORDIER, C
DOI：
10.1186/1475-9292-2-3
发表时间：
2003-06-25
期刊：
Kinetoplastid Biology and Disease
影响因子：
--
作者：
Seed, John Richard;Wenck, Mary Anne
通讯作者：
Wenck, Mary Anne