Baker Rosanna P, Wijetilaka Ruvini, Urban Sinisa
Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.
PLoS Pathog. 2006 Oct;2(10):e113. doi: 10.1371/journal.ppat.0020113.
Invasion of host cells by the malaria pathogen Plasmodium relies on parasite transmembrane adhesins that engage host-cell receptors. Adhesins must be released by cleavage before the parasite can enter the cell, but the processing enzymes have remained elusive. Recent work indicates that the Toxoplasma rhomboid intramembrane protease TgROM5 catalyzes this essential cleavage. However, Plasmodium does not encode a direct TgROM5 homolog. We examined processing of the 14 Plasmodium falciparum adhesins currently thought to be involved in invasion by both model and Plasmodium rhomboid proteases in a heterologous assay. While most adhesins contain aromatic transmembrane residues and could not be cleaved by nonparasite rhomboid proteins, including Drosophila Rhomboid-1, Plasmodium falciparum rhomboid protein (PfROM)4 (PFE0340c) was able to process these adhesins efficiently and displayed novel substrate specificity. Conversely, PfROM1 (PF11_0150) shared specificity with rhomboid proteases from other organisms and was the only PfROM able to cleave apical membrane antigen 1 (AMA1). PfROM 1 and/or 4 was thus able to cleave diverse adhesins including TRAP, CTRP, MTRAP, PFF0800c, EBA-175, BAEBL, JESEBL, MAEBL, AMA1, Rh1, Rh2a, Rh2b, and Rh4, but not PTRAMP, and cleavage relied on the adhesin transmembrane domains. Swapping transmembrane regions between BAEBL and AMA1 switched the relative preferences of PfROMs 1 and 4 for these two substrates. Our analysis indicates that PfROMs 1 and 4 function with different substrate specificities that together constitute the specificity of TgROM5 to cleave diverse adhesins. This is the first enzymatic analysis of Plasmodium rhomboid proteases and suggests an involvement of PfROMs in all invasive stages of the malaria lifecycle, in both the vertebrate host and the mosquito vector.
疟原虫病原体疟原虫对宿主细胞的侵袭依赖于与宿主细胞受体结合的寄生虫跨膜粘附素。在寄生虫能够进入细胞之前，粘附素必须通过切割释放，但加工酶一直难以捉摸。最近的研究表明，弓形虫菱形膜内蛋白酶TgROM5催化这种关键的切割。然而，疟原虫并不编码直接的TgROM5同源物。我们在异源试验中，研究了目前认为参与侵袭的14种恶性疟原虫粘附素被模型和疟原虫菱形蛋白酶的加工情况。虽然大多数粘附素含有芳香族跨膜残基，不能被非寄生虫菱形蛋白（包括果蝇菱形蛋白-1）切割，但恶性疟原虫菱形蛋白（PfROM）4（PFE0340c）能够有效地加工这些粘附素，并表现出新颖的底物特异性。相反，PfROM1（PF11_0150）与其他生物体的菱形蛋白酶具有共同的特异性，并且是唯一能够切割顶端膜抗原1（AMA1）的PfROM。因此，PfROM 1和/或4能够切割多种粘附素，包括TRAP、CTRP、MTRAP、PFF0800c、EBA-175、BAEBL、JESEBL、MAEBL、AMA1、Rh1、Rh2a、Rh2b和Rh4，但不能切割PTRAMP，并且切割依赖于粘附素的跨膜结构域。在BAEBL和AMA1之间交换跨膜区域，改变了PfROMs 1和4对这两种底物的相对偏好。我们的分析表明，PfROMs 1和4以不同的底物特异性发挥作用，共同构成了TgROM5切割多种粘附素的特异性。这是对疟原虫菱形蛋白酶的首次酶学分析，并表明PfROMs参与了疟原虫生命周期在脊椎动物宿主和蚊媒中的所有侵袭阶段。
