Cobb David W, Kudyba Heather M, Villegas Alejandra, Hoopmann Michael R, Baptista Rodrigo P, Bruton Baylee, Krakowiak Michelle, Moritz Robert L, Muralidharan Vasant
Department of Cellular Biology, University of Georgia, Georgia, United States of America.
Center for Tropical and Emerging Global Diseases, University of Georgia, Georgia, United States of America.
PLoS Pathog. 2021 Feb 3;17(2):e1009293. doi: 10.1371/journal.ppat.1009293. eCollection 2021 Feb.
Malaria remains a major global health problem, creating a constant need for research to identify druggable weaknesses in P. falciparum biology. As important components of cellular redox biology, members of the Thioredoxin (Trx) superfamily of proteins have received interest as potential drug targets in Apicomplexans. However, the function and essentiality of endoplasmic reticulum (ER)-localized Trx-domain proteins within P. falciparum has not been investigated. We generated conditional mutants of the protein PfJ2-an ER chaperone and member of the Trx superfamily-and show that it is essential for asexual parasite survival. Using a crosslinker specific for redox-active cysteines, we identified PfJ2 substrates as PfPDI8 and PfPDI11, both members of the Trx superfamily as well, which suggests a redox-regulatory role for PfJ2. Knockdown of these PDIs in PfJ2 conditional mutants show that PfPDI11 may not be essential. However, PfPDI8 is required for asexual growth and our data suggest it may work in a complex with PfJ2 and other ER chaperones. Finally, we show that the redox interactions between these Trx-domain proteins in the parasite ER and their substrates are sensitive to small molecule inhibition. Together these data build a model for how Trx-domain proteins in the P. falciparum ER work together to assist protein folding and demonstrate the suitability of ER-localized Trx-domain proteins for antimalarial drug development.
疟疾仍然是一个重大的全球健康问题，这使得持续开展研究以确定恶性疟原虫生物学中可成药的薄弱环节成为必要。作为细胞氧化还原生物学的重要组成部分，硫氧还蛋白（Trx）超家族的蛋白质成员作为顶复门原虫潜在的药物靶点受到了关注。然而，恶性疟原虫内质网（ER）定位的Trx结构域蛋白的功能和必要性尚未得到研究。我们构建了蛋白质PfJ2（一种ER伴侣蛋白和Trx超家族成员）的条件性突变体，并表明它对无性寄生虫的存活至关重要。使用一种对氧化还原活性半胱氨酸特异的交联剂，我们鉴定出PfJ2的底物为PfPDI8和PfPDI11，它们也都是Trx超家族的成员，这表明PfJ2具有氧化还原调节作用。在PfJ2条件性突变体中敲低这些PDI表明PfPDI11可能不是必需的。然而，PfPDI8是无性生长所必需的，我们的数据表明它可能与PfJ2和其他ER伴侣蛋白形成复合物发挥作用。最后，我们表明寄生虫ER中这些Trx结构域蛋白与其底物之间的氧化还原相互作用对小分子抑制敏感。这些数据共同构建了一个模型，说明恶性疟原虫ER中的Trx结构域蛋白如何协同作用以协助蛋白质折叠，并证明了ER定位的Trx结构域蛋白作为抗疟药物开发靶点的适用性。
