Abstract：
Na+/H+ exchanger isoform 1 (NHE1), a member of a large family of integral membrane proteins, plays a role in regulating the cortical actin cytoskeleton. Trypanosoma cruzi, the agent of Chagas disease, depends on F-actin rearrangement and lysosome mobilization to invade host cells. To determine the involvement of NHE1 in T. cruzi metacyclic trypomastigote (MT) internalization, the effect of treatment in cells with NHE1 inhibitor amiloride or of NHE1 depletion was examined in human epithelial cells. MT invasion decreased in amiloride-treated and NHE1-depleted cells. The phosphorylation profile of diverse protein kinases, whose activation is associated with remodeling of actin fibers, was analyzed in amiloride-treated and NHE1-depleted cells. In amiloride-treated cells, the phosphorylation levels of protein kinase C (PKC), focal adhesion kinase (FAK) and Akt were similar to those of untreated cells, whereas those of extracellular signal-regulated protein kinases (ERK1/2) increased. In NHE1-deficient cells, with marked alteration in the actin cytoskeleton architecture and in lysosome distribution, the levels of phospho-PKC and phospho-FAK decreased, whereas those of phospho-Akt and phospho-ERK1/2 increased. These data indicate that NHE1 plays a role in MT invasion, by maintaining the activation status of diverse protein kinases in check and preventing the inappropriate F-actin arrangement that affects lysosome distribution.
摘要：
Na+/H+交换子亚型1(NHE1)，一个完整的膜蛋白家族的成员，在调节皮质肌动蛋白细胞骨架中起作用。克氏锥虫,查加斯病的代理人，依赖于F-肌动蛋白重排和溶酶体动员来侵入宿主细胞。为了确定NHE1参与克氏虫脂环色素动物（MT）内化，在人上皮细胞中检查了用NHE1抑制剂阿米洛利或NHE1耗竭对细胞的治疗效果。在阿米洛利处理和NHE1耗尽的细胞中MT侵袭减少。不同蛋白激酶的磷酸化谱，其激活与肌动蛋白纤维的重塑有关，在阿米洛利处理和NHE1耗尽的细胞中进行分析。在阿米洛利处理的细胞中，蛋白激酶C（PKC）的磷酸化水平，粘着斑激酶（FAK）和Akt与未处理的细胞相似，而细胞外信号调节蛋白激酶（ERK1/2）的增加。在NHE1缺陷细胞中，随着肌动蛋白细胞骨架结构和溶酶体分布的显著改变，磷酸-PKC和磷酸-FAK的水平下降，而磷酸化-Akt和磷酸化-ERK1/2的增加。这些数据表明NHE1在MT入侵中起作用，通过维持多种蛋白激酶的激活状态来检查并防止影响溶酶体分布的不适当的F-肌动蛋白排列。
