Lavazec Catherine, Moreira Cristina K, Mair Gunnar R, Waters Andrew P, Janse Chris J, Templeton Thomas J
Department of Microbiology and Immunology, Weill Cornell Medical College, New York, NY 10021, USA.
Mol Biochem Parasitol. 2009 Jan;163(1):1-7. doi: 10.1016/j.molbiopara.2008.09.002. Epub 2008 Sep 19.
Plasmodium encodes a family of six secreted multi-domain adhesive proteins, termed PCCps, which are released from gametocytes during emergence within the mosquito midgut. The expression and cellular localization of PCCp proteins predict a role either in gametocyte development or within the mosquito midgut during the transition from gametes into the ookinete stage. However, mutant parasites lacking expression of any single PCCp protein show a phenotype at the oocyst stage with a failure of oocyst maturation and sporozoite formation. In this study we investigated the stage-specific transcription of the PCCp genes of the rodent malaria parasite, Plasmodium berghei, and analyzed their promoter activities. Transcript expression analysis by quantitative real time RT-PCR showed that as in the human malaria parasite, Plasmodium falciparum, all PbCCp genes are predominantly transcribed in the gametocyte stage with a low level of transcription in the oocyst stage. Transgenic P. berghei parasites that contain the reporter protein GFP driven by the promoter regions of PbCCps showed pronounced GFP expression exclusively in gametocytes, in agreement with the RT-PCR data. To determine whether functional redundancies of different PCCp family members could explain the lack of a phenotype in gametocytes or gametes in single knockout mutant parasites, double gene null mutant P. berghei parasites were generated lacking either PCCp1 and PCCp3, or PCCp1 and PCCp4. The phenotype of these double knockout mutants was similar to that observed for single gene knockout mutants and manifest at the oocyst rather than the gametocyte or other stages within the mosquito midgut lumen.
疟原虫编码一个由六种分泌型多结构域粘附蛋白组成的家族，称为PCCps，它们在配子体出现在蚊子中肠时从配子体中释放出来。PCCp蛋白的表达和细胞定位预示其在配子体发育过程中或在从配子到动合子阶段的转变过程中在蚊子中肠内发挥作用。然而，缺乏任何一种PCCp蛋白表达的突变寄生虫在卵囊阶段表现出一种表型，即卵囊成熟和子孢子形成失败。在本研究中，我们调查了啮齿动物疟原虫伯氏疟原虫PCCp基因的阶段特异性转录，并分析了它们的启动子活性。通过定量实时RT-PCR进行的转录本表达分析表明，与人类疟原虫恶性疟原虫一样，所有PbCCp基因主要在配子体阶段转录，在卵囊阶段转录水平较低。含有由PbCCps启动子区域驱动的报告蛋白GFP的转基因伯氏疟原虫寄生虫仅在配子体中显示出明显的GFP表达，这与RT-PCR数据一致。为了确定不同PCCp家族成员的功能冗余是否可以解释单基因敲除突变寄生虫中配子体或配子中缺乏表型的现象，我们构建了缺失PCCp1和PCCp3或PCCp1和PCCp4的双基因缺失突变伯氏疟原虫寄生虫。这些双敲除突变体的表型与单基因敲除突变体观察到的表型相似，并且在卵囊阶段而不是在蚊子中肠腔中的配子体或其他阶段表现出来。
