Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo, São Paulo, Brazil.
Departamento de Biofísica, Universidade Federal de São Paulo, São Paulo, Brazil.
Biochem Biophys Res Commun. 2018 Sep 5;503(2):722-728. doi: 10.1016/j.bbrc.2018.06.067. Epub 2018 Jun 20.
Calcium signaling has an essential role in fundamental processes of Plasmodium life cycle, including migration, cell invasion and parasite development. Two important players in calcium homeostasis, the Histidine Triad (HIT) protein that is implicated in calcium signaling in mammalian cells and calmodulin, which is a classic calcium sensor in eukaryotes are present in Plasmodium falciparum, however theirs function is unknown in the parasite. Here, we investigated the involvement of the P. falciparum Histidine Triad protein (PfHint-1) and calmodulin (PfCaM) in calcium signaling and intracellular proteolysis. For this, we targeted PfHint-1 with a hemagglutinin tail and overexpressed both proteins. We observed that PfHint-1 is expressed throughout the erythrocytic stages and partially colocalizes to the endoplasmic reticulum. Parasites overexpressing PfHint-1 displayed lower ER Ca content and a higher [Ca] rise in the parasite cytosol upon Ca addition to the extracellular medium after depletion of ER calcium store. PfCaM-overexpressing parasites exhibit a higher [Ca] rise after challenge with the calmodulin inhibitor, calmidazolium. The calcium-dependent proteolytic activity in PfCaM- and PfHint-1-overexpressing parasites was increased and correlated to alterations in calcium homeostasis. Taken together, our results indicate the participation of these proteins in P. falciparum fundamental cellular processes and highlights promising targets for the development of antimalarial drugs.
钙信号在疟原虫生命周期的基本过程中起着至关重要的作用，包括迁移、细胞入侵和寄生虫发育。钙稳态的两个重要参与者，即参与哺乳动物细胞钙信号的组氨酸三肽（HIT）蛋白和钙调蛋白，在恶性疟原虫中都存在，然而它们在寄生虫中的功能尚不清楚。在这里，我们研究了恶性疟原虫组氨酸三肽蛋白（PfHint-1）和钙调蛋白（PfCaM）在钙信号和细胞内蛋白水解中的作用。为此，我们用一个血凝素尾巴靶向 PfHint-1，并过表达这两种蛋白。我们观察到 PfHint-1 在整个红细胞阶段表达，并部分与内质网共定位。PfHint-1 过表达的寄生虫在耗尽内质网钙库后，向细胞外培养基中添加 Ca 时，细胞溶质中的 ER Ca 含量降低，[Ca]升高。用钙调蛋白抑制剂 calmidazolium 刺激 PfCaM 过表达的寄生虫时，[Ca]升高更高。PfCaM 和 PfHint-1 过表达的寄生虫中的钙依赖性蛋白水解活性增加，并与钙稳态的改变相关。总之，我们的结果表明这些蛋白参与了恶性疟原虫的基本细胞过程，并强调了针对抗疟药物开发的有前途的靶标。
