Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo, Japan.
Department of Pharmacology and Toxicology, Kyorin University School of Medicine, Tokyo, Japan.
Front Cell Infect Microbiol. 2021 Sep 15;11:737457. doi: 10.3389/fcimb.2021.737457. eCollection 2021.
The quality control and export of mRNA by RNA-binding proteins are necessary for the survival of malaria parasites, which have complex life cycles. Nuclear poly(A) binding protein 2 (NAB2), THO complex subunit 4 (THO4), nucleolar protein 3 (NPL3), G-strand binding protein 2 (GBP2) and serine/arginine-rich splicing factor 1 (SR1) are involved in nuclear mRNA export in malaria parasites. However, their roles in asexual and sexual development, and in cellular localization, are not fully understood. In this study using the rodent malaria parasite, , we found that NAB2 and SR1, but not THO4, NPL3 or GBP2, played essential roles in the asexual development of malaria parasites. By contrast, GBP2 but not NPL3 was involved in male and female gametocyte production. THO4 was involved in female gametocyte production, but had a lower impact than GBP2. In this study, we focused on GBP2 and NAB2, which play important roles in the sexual and asexual development of malaria parasites, respectively, and examined their cellular localization. GBP2 localized to both the nucleus and cytoplasm of malaria parasites. Using immunoprecipitation coupled to mass spectrometry (IP-MS), GBP2 interacted with the proteins ALBA4, DOZI, and CITH, which play roles in translational repression. IP-MS also revealed that phosphorylated adapter RNA export protein (PHAX) domain-containing protein, an adaptor protein for exportin-1, also interacted with GBP2, implying that mRNA export occurs the PHAX domain-containing protein pathway in malaria parasites. Live-cell fluorescence imaging revealed that NAB2 localized at the nuclear periphery. Moreover, IP-MS indicated that NAB2 interacted with transportin. RNA immunoprecipitation coupled to RNA sequencing revealed that NAB2 bound directly to 143 mRNAs, including those encoding 40S and 60S ribosomal proteins. Our findings imply that malaria parasites use an evolutionarily ancient mechanism conserved throughout eukaryotic evolution.
RNA 结合蛋白对 mRNA 的质量控制和输出对于疟原虫的生存是必要的，疟原虫具有复杂的生命周期。核多聚（A）结合蛋白 2（NAB2）、THO 复合物亚基 4（THO4）、核仁蛋白 3（NPL3）、G-链结合蛋白 2（GBP2）和丝氨酸/精氨酸丰富剪接因子 1（SR1）参与疟原虫的核 mRNA 输出。然而，它们在无性和有性发育以及细胞定位中的作用尚未完全阐明。在这项使用啮齿动物疟原虫的研究中，我们发现 NAB2 和 SR1，但不是 THO4、NPL3 或 GBP2，在疟原虫的无性发育中发挥着重要作用。相比之下，GBP2 而不是 NPL3 参与了雄配子体和雌配子体的产生。THO4 参与了雌配子体的产生，但影响低于 GBP2。在这项研究中，我们重点关注分别在疟原虫的有性和无性发育中发挥重要作用的 GBP2 和 NAB2，并研究了它们的细胞定位。GBP2 定位于疟原虫的细胞核和细胞质中。通过免疫沉淀结合质谱（IP-MS），GBP2 与在翻译抑制中起作用的蛋白质 ALBA4、DOZI 和 CITH 相互作用。IP-MS 还表明，磷酸化适配器 RNA 输出蛋白（PHAX）结构域包含蛋白，一种用于 exportin-1 的衔接蛋白，也与 GBP2 相互作用，这意味着在疟原虫中，mRNA 输出发生在 PHAX 结构域包含蛋白途径中。活细胞荧光成像显示 NAB2 定位于核周。此外，IP-MS 表明 NAB2 与 transportin 相互作用。RNA 免疫沉淀结合 RNA 测序显示，NAB2 直接与 143 个 mRNA 结合，包括编码 40S 和 60S 核糖体蛋白的 mRNA。我们的研究结果表明，疟原虫使用了一种在整个真核生物进化中保守的古老的进化机制。
