Center for Tropical and Emerging Global Diseases and Department of Cellular Biology, University of Georgia, Athens, GA, 30602, USA; Departamento de Patologia Clínica, Faculdade de Ciências Médicas, Universidade Estadual de Campinas, Campinas, São Paulo, 13083, Brazil.
Center for Tropical and Emerging Global Diseases and Department of Cellular Biology, University of Georgia, Athens, GA, 30602, USA; Departamento de Patologia Clínica, Faculdade de Ciências Médicas, Universidade Estadual de Campinas, Campinas, São Paulo, 13083, Brazil.
Cell Calcium. 2020 Dec;92:102284. doi: 10.1016/j.ceca.2020.102284. Epub 2020 Sep 2.
In contrast to animal cells, the inositol 1,4,5-trisphosphate receptor of Trypanosoma cruzi (TcIPR) localizes to acidocalcisomes instead of the endoplasmic reticulum. Here, we present evidence that TcIPR is a Ca release channel gated by IP when expressed in DT40 cells knockout for all vertebrate IP receptors, and is required for Ca uptake by T. cruzi mitochondria, regulating pyruvate dehydrogenase dephosphorylation and mitochondrial O consumption, and preventing autophagy. Localization studies revealed its co-localization with an acidocalcisome marker in all life cycle stages of the parasite. Ablation of TcIPR by CRISPR/Cas9 genome editing caused: a) a reduction in O consumption rate and citrate synthase activity; b) decreased mitochondrial Ca transport without affecting the membrane potential; c) increased ammonia production and AMP/ATP ratio; d) stimulation of autophagosome formation, and e) marked defects in growth of culture forms (epimastigotes) and invasion of host cells by infective stages (trypomastigotes). Moreover, TcIPR overexpressing parasites showed decreased metacyclogenesis, trypomastigote host cell invasion and intracellular amastigote replication. In conclusion, the results suggest a modulatory activity of TcIPR-mediated acidocalcisome Ca release on cell bioenergetics in T. cruzi.
与动物细胞相反，克氏锥虫（Trypanosoma cruzi）的肌醇 1,4,5-三磷酸受体（TcIPR）定位于酸性钙囊泡而不是内质网。在这里，我们提供的证据表明，当在敲除所有脊椎动物 IP 受体的 DT40 细胞中表达时，TcIPR 是一种由 IP 门控的 Ca 释放通道，并且是克氏锥虫线粒体摄取 Ca 所必需的，调节丙酮酸脱氢酶去磷酸化和线粒体 O 消耗，并防止自噬。定位研究表明，它与寄生虫所有生命周期阶段的酸性钙囊泡标记物共定位。CRISPR/Cas9 基因组编辑对 TcIPR 的消融导致：a）O 消耗率和柠檬酸合酶活性降低；b）线粒体 Ca 转运减少而不影响膜电位；c）氨产量增加和 AMP/ATP 比增加；d）自噬体形成增加，以及 e）培养形式（epimastigotes）和感染阶段（trypomastigotes）入侵宿主细胞的生长明显缺陷。此外，TcIPR 过表达寄生虫显示出降低的 metacyclogenesis、trypomastigote 宿主细胞入侵和细胞内无鞭毛体复制。总之，结果表明 TcIPR 介导的酸性钙囊泡 Ca 释放对克氏锥虫细胞生物能量学具有调节活性。
