Department of Genetics, Biology and Biochemistry, University of Turin, Turin, Italy.
Proteomics. 2010 Oct;10(19):3469-79. doi: 10.1002/pmic.201000269.
Phosphorylation of erythrocyte membrane proteins has been previously documented following infection and intracellular growth of the malarial parasite, Plasmodium falciparum in red cells. Much of this data dealt with phosphorylation of serine residues. In this study, we report detailed characterization of phosphorylation of serine and tyrosine residues of red cell membrane proteins following infection by P falciparum. Western blot analysis using anti-phosphotyrosine and anti-phosphoserine antibodies following 2-DE in conjunction with double channel laser-induced infrared fluorescence enabled accurate assessment of phosphorylation changes. Tyrosine phosphorylation of band 3 represented the earliest modification observed during parasite development. Band 3 tyrosine phosphorylation observed at the ring stage appears to be under the control of Syk kinase. Serine and tyrosine phosphorylation of additional cytoskeletal, trans-membrane and membrane associated proteins was documented as intracellular development of parasite progressed. Importantly, during late schizont stage of parasite maturation, we observed widespread protein dephosphorylation. In vitro treatments that caused distinct activation of red cell tyrosine and serine kinases elicited phosphorylative patterns similar to what observed in parasitized red blood cell, suggesting primary involvement of erythrocyte kinases. Identification of tyrosine phosphorylations of band 3, band 4.2, catalase and actin which have not been previously described in P. falciparum infected red cells suggests new potential regulatory mechanisms that could modify the functions of the host cell membrane.
先前有研究报道，疟原虫（Plasmodium falciparum）在红细胞内感染和增殖后，其红细胞膜蛋白会发生磷酸化。这些数据主要涉及丝氨酸残基的磷酸化。在本研究中，我们详细描述了疟原虫感染后红细胞膜蛋白丝氨酸和酪氨酸残基的磷酸化情况。利用抗磷酸酪氨酸和抗磷酸丝氨酸抗体进行 Western blot 分析，结合二维电泳和双通道激光诱导红外荧光技术，可准确评估磷酸化变化。带 3 的酪氨酸磷酸化是在寄生虫发育过程中观察到的最早修饰。环期寄生虫发育过程中观察到的带 3 酪氨酸磷酸化似乎受到 Src 相关酪氨酸激酶（Syk 激酶）的控制。随着寄生虫的发育，还记录到了其他细胞骨架、跨膜和膜相关蛋白的丝氨酸和酪氨酸磷酸化。重要的是，在寄生虫成熟的晚期裂殖体阶段，我们观察到广泛的蛋白质去磷酸化。体外处理可明显激活红细胞酪氨酸和丝氨酸激酶，其磷酸化模式与寄生红细胞中观察到的相似，提示红细胞激酶的主要参与。鉴定了以前在感染疟原虫的红细胞中未描述过的带 3、带 4.2、过氧化氢酶和肌动蛋白的酪氨酸磷酸化，提示了可能改变宿主细胞膜功能的新的潜在调节机制。
