Plasmodium falciparum guanylyl cyclase-alpha and the activity of its appended P4-ATPase domain are essential for cGMP synthesis and blood stage egress
Plasmodium falciparum guanylyl cyclase-alpha and the activity of its appended P4-ATPase domain are essential for cGMP synthesis and blood stage egress
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恶性疟原虫鸟苷酸环化酶-α 及其附加 P4-ATP 酶结构域的活性对于 cGMP 合成和血液阶段排出至关重要
DOI：
10.1101/2020.09.07.285734
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Nofal S
中科院分区：
文献类型：
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作者：
Nofal S
鸟苷酸环化酶（GC）合成环GMP（cGMP），并且与环核苷酸磷酸二酯酶一起负责调节这种细胞内信使的水平，所述细胞内信使介导跨真核生物的无数功能。在疟疾寄生虫（疟原虫属），以及他们的apicomplexan和纤毛虫亲属，GC与一个独特的双功能配置的P4-ATP酶样结构域。P4-ATP酶通过将磷脂从外叶移位到内叶来产生膜双层脂质不对称性。在这里，我们研究恶性疟原虫鸟苷酸环化酶α（GCα）及其相关的P4-ATP酶模块的作用，表明缺乏环化酶和P4-ATP酶结构域的无性血液阶段的寄生虫无法从宿主红细胞中排出。GCα无效寄生虫不能合成cGMP或动员钙，这是cGMP依赖性蛋白激酶（PKG）驱动的外出要求。使用化学互补与cGMP类似物和点突变的一个关键的保守残基内的P4-ATP酶结构域，我们表明，P4-ATP酶活性的上游，并连接到cGMP合成。总的来说，我们的研究结果表明，GCα是PKG的一个重要的调节剂，其相关的P4-ATP酶结构域在产生cGMP裂殖子egression.IMPORTANCEThe疟疾的临床表现出现由于连续轮的复制ofPlasmodiumparasites内红细胞中起着主要作用。一旦成熟，子裂殖子从受感染的红细胞中释放出来，在一个被称为出口的严格调控的过程中侵入新细胞。以前的研究表明，激活环GMP（cGMP）信号是启动出口的关键。在这里，我们证明了GCα，一种独特的双功能酶，是唯一的酶负责cGMP生产在恶性疟原虫的无性血液阶段，并需要导致裂殖子出口的细胞事件。我们进一步证明，除了GC结构域，GCα的附加ATP酶样结构域也参与cGMP的产生。我们的研究结果强调了GCα在协调疟原虫外出所需的cGMP信号传导中的关键作用。
Guanylyl cyclases (GCs) synthesize cyclic GMP (cGMP) and, together with cyclic nucleotide phosphodiesterases, are responsible for regulating levels of this intracellular messenger which mediates myriad functions across eukaryotes. In malaria parasites (Plasmodium spp), as well as their apicomplexan and ciliate relatives, GCs are associated with a P4-ATPase-like domain in a unique bifunctional configuration. P4-ATPases generate membrane bilayer lipid asymmetry by translocating phospholipids from the outer to the inner leaflet. Here, we investigate the role of Plasmodium falciparum guanylyl cyclase alpha (GCα) and its associated P4-ATPase module, showing that asexual blood-stage parasites lacking both the cyclase and P4-ATPase domains are unable to egress from host erythrocytes. GCα-null parasites cannot synthesize cGMP or mobilize calcium, a cGMP-dependent protein kinase (PKG)-driven requirement for egress. Using chemical complementation with a cGMP analogue and point mutagenesis of a crucial conserved residue within the P4-ATPase domain, we show that P4-ATPase activity is upstream of and linked to cGMP synthesis. Collectively, our results demonstrate that GCα is a critical regulator of PKG and that its associated P4-ATPase domain plays a primary role in generating cGMP for merozoite egress.IMPORTANCEThe clinical manifestations of malaria arise due to successive rounds of replication ofPlasmodiumparasites within red blood cells. Once mature, daughter merozoites are released from infected erythrocytes to invade new cells in a tightly regulated process termed egress. Previous studies have shown that the activation of cyclic GMP (cGMP) signaling is critical for initiating egress. Here, we demonstrate that GCα, a unique bifunctional enzyme, is the sole enzyme responsible for cGMP production during the asexual blood stages of Plasmodium falciparum and is required for the cellular events leading up to merozoite egress. We further demonstrate that in addition to the GC domain, the appended ATPase-like domain of GCα is also involved in cGMP production. Our results highlight the critical role of GCα in cGMP signaling required for orchestrating malaria parasite egress.