Modulation of the Surface Proteome through Multiple Ubiquitylation Pathways in African Trypanosomes.
Modulation of the Surface Proteome through Multiple Ubiquitylation Pathways in African Trypanosomes.
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DOI：
10.1371/journal.ppat.1005236
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发表时间：
2015-10
期刊：
影响因子：
6.7
通讯作者：
Field MC
中科院分区：
文献类型：
--
作者：
Zoltner M;Leung KF;Alsford S;Horn D;Field MC
最近，我们确定了多个苏拉明敏感基因的基因组范围内筛选布氏锥虫，其中包括不变的表面糖蛋白ISG 75，适应蛋白-1（AP-1）复合物和两个去泛素化酶（DUBs）的orthopathic到ScUbp 15/HsHAUSP 1和pVHL相互作用DUB 1（I型），指定TbUsp 7和TbVdu 1，分别。在这里，我们研究了这些基因在ISG 75运输中的作用，ISG 75似乎是苏拉明摄取的关键。我们发现，虽然AP-1不影响ISG 75丰度，但TbUsp 7或TbVdu 1的敲低导致ISG 75丰度降低。沉默TbVdu 1也降低了ISG 65丰度。TbVdu 1是进化上保守的泛素化开关的一个组成部分，负责快速受体调节，表明T.布鲁塞。出乎意料的是，TbUsp 7敲低也阻断了内吞作用。为了整合这些观察结果，我们使用SILAC分析了TbUsp 7和TbVdu 1敲低对全局蛋白质组的影响。对于TbVdu 1，ISG 65和ISG 75是唯一显著调节的蛋白质，但对于TbUsp 7，一组整合的膜蛋白，包括内吞作用所需的酸性磷酸酶MBAP 1，以及其他ISG相关蛋白质被下调。此外，我们发现ESAG 6/7转铁蛋白受体和ESAG 5的表达增加，可能是由于内吞活性降低。因此，多种泛素化途径，与运输途径的复杂相互作用，控制锥虫表面蛋白质组的表达。病原体与其环境相互作用的机制对于满足寄生虫的基本需求和理解免疫逃避都非常重要。对于非洲锥虫来说，表面以变异表面糖蛋白（VSG）为主，但最近的数据表明，遍在化在介导不变表面糖蛋白（ISG）的转换和维持ISG拷贝数不依赖于VSG方面发挥着重要作用。此外，ISG表达是苏拉明敏感性所必需的。在这里，我们描述的机制介导ISG营业额，发现使用屏幕的敏感性苏拉明基因。这些涉及泛素化机制的多个方面，并将ISG周转与其他表面蛋白联系起来。我们的数据提供了第一次深入了解ISG家族调控的复杂性，确定锥虫中药物敏感性途径控制的进一步方面，并提供寄生虫表面蛋白质组代谢的见解。
Recently we identified multiple suramin-sensitivity genes with a genome wide screen in Trypanosoma brucei that includes the invariant surface glycoprotein ISG75, the adaptin-1 (AP-1) complex and two deubiquitylating enzymes (DUBs) orthologous to ScUbp15/HsHAUSP1 and pVHL-interacting DUB1 (type I), designated TbUsp7 and TbVdu1, respectively. Here we have examined the roles of these genes in trafficking of ISG75, which appears key to suramin uptake. We found that, while AP-1 does not influence ISG75 abundance, knockdown of TbUsp7 or TbVdu1 leads to reduced ISG75 abundance. Silencing TbVdu1 also reduced ISG65 abundance. TbVdu1 is a component of an evolutionarily conserved ubiquitylation switch and responsible for rapid receptor modulation, suggesting similar regulation of ISGs in T. brucei. Unexpectedly, TbUsp7 knockdown also blocked endocytosis. To integrate these observations we analysed the impact of TbUsp7 and TbVdu1 knockdown on the global proteome using SILAC. For TbVdu1, ISG65 and ISG75 are the only significantly modulated proteins, but for TbUsp7 a cohort of integral membrane proteins, including the acid phosphatase MBAP1, that is required for endocytosis, and additional ISG-related proteins are down-regulated. Furthermore, we find increased expression of the ESAG6/7 transferrin receptor and ESAG5, likely resulting from decreased endocytic activity. Therefore, multiple ubiquitylation pathways, with a complex interplay with trafficking pathways, control surface proteome expression in trypanosomes. The mechanisms by which pathogens interact with their environment are of major importance, both for fulfilling the basic needs of the parasite and understanding immune evasion. For African trypanosomes, the surface is dominated by the variant surface glycoprotein (VSG), but recent data has demonstrated an important role for ubiquitylation in mediating turnover of invariant surface glycoproteins (ISGs) and maintaining ISG copy number independent of VSG. Further, ISG expression is required for suramin-sensitivity. Here we describe mechanisms mediating ISG turnover, uncovered using a screen for genes involved in sensitivity to suramin. These involve multiple aspects of the ubiquitylation machinery, and connect ISG turnover with additional surface proteins. Our data provide a first insight into the complexity of regulation of the ISG family, identifying further aspects to the control of a drug-sensitivity pathway in trypanosomes, and offering insights into metabolism of the parasite surface proteome.