Department of Microbiology and Molecular Genetics, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, Texas 77030.
Department of Microbiology and Molecular Genetics, McGovern Medical School, University of Texas Health Science Center at Houston, Houston, Texas 77030
J Biol Chem. 2019 Nov 8;294(45):16672-16683. doi: 10.1074/jbc.RA119.010538. Epub 2019 Sep 20.
The evolutionarily early divergent human parasite proliferates through binary cell fission in both its tsetse fly vector and mammalian host. The parasite divides unidirectionally along the longitudinal cell axis from the anterior cell tip toward the posterior cell tip through a mechanism distinct from that in the cells of its human host. Initiation of cytokinesis in is regulated by two evolutionarily conserved protein kinases, the Polo-like kinase TbPLK and the Aurora B kinase TbAUK1, and a cohort of trypanosome-specific proteins, including the three cytokinesis initiation factors CIF1, CIF2, and CIF3. Here, using RNAi, epitope tagging of proteins, GST pulldown, and coimmunoprecipitation assays, and immunofluorescence and scanning electron microscopy analyses, we report the identification and functional characterization of two trypanosome-specific proteins, flagellum attachment zone tip-localizing protein required for cytokinesis (FPRC) and CIF4. We found that the two proteins colocalize to the distal tips of the new and the old flagellum attachment zones and are required for cytokinesis initiation. Knockdown of FPRC or CIF4 disrupted the localization of CIF1, suggesting that they function upstream of CIF1. Moreover, depletion of CIF4 abolished FPRC localization, indicating that CIF4 acts upstream of FPRC. Together, these results identify two new cytokinesis regulators in and integrate them into the CIF1-mediated cytokinesis regulatory pathway. These findings highlight the existence of a cytokinesis pathway in that is different from that of its mammalian host and therefore suggest that cytokinesis in could potentially be exploited as a new drug target.
原生动物寄生虫 在其采采蝇媒介和哺乳动物宿主中通过二分裂进行增殖。寄生虫沿着纵向细胞轴从前细胞尖端单向分裂到后细胞尖端，其分裂机制与宿主细胞不同。 在 中，有丝分裂的起始受两个进化上保守的蛋白激酶（Polo 样激酶 TbPLK 和 Aurora B 激酶 TbAUK1）和一组原虫特异性蛋白（包括三个细胞分裂起始因子 CIF1、CIF2 和 CIF3）的调节。在这里，我们使用 RNAi、蛋白的表位标记、GST 下拉和共免疫沉淀分析以及免疫荧光和扫描电子显微镜分析，鉴定并研究了两种原虫特异性蛋白——鞭毛附着区尖端定位蛋白（flagellum attachment zone tip-localizing protein required for cytokinesis，FPRC）和 CIF4 的功能特征。我们发现这两种蛋白都定位于新老鞭毛附着区的远端尖端，并且对于有丝分裂的起始是必需的。FPRC 或 CIF4 的敲低会破坏 CIF1 的定位，表明它们在 CIF1 之前起作用。此外，CIF4 的耗竭会消除 FPRC 的定位，表明 CIF4 在 FPRC 之前起作用。综上所述，这些结果确定了 中的两个新的有丝分裂调控因子，并将它们整合到 CIF1 介导的有丝分裂调控途径中。这些发现突显了 在有丝分裂中存在与哺乳动物宿主不同的途径，因此表明 中的有丝分裂可能成为新的药物靶点。
