Mechanisms of aquaporin-4 vesicular trafficking in mammalian cells
Mechanisms of aquaporin-4 vesicular trafficking in mammalian cells
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DOI：
10.1111/jnc.16029
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发表时间：
2023-12-16
影响因子：
4.7
通讯作者：
Balklava，Zita
中科院分区：
文献类型：
--
作者：
Markou，Andrea;Kitchen，Philip;Balklava，Zita
水通道蛋白-4（AQP 4）水通道在中枢神经系统的神经胶质细胞中大量表达，并在各种损伤（如创伤性损伤或中风）后促进脑肿胀。缺乏特异性和治疗性AQP 4抑制剂突出了探索替代途径来控制神经胶质细胞膜透水性的需要。哺乳动物细胞中AQP 4的细胞表面丰度响应于氧水平和张力的变化而迅速波动，这表明囊泡运输在其易位到细胞表面和从细胞表面转运中的作用。然而，AQP 4运输的分子机制尚未完全阐明。在这项工作中，早期和再循环内体作为可能的候选人的快速AQP 4易位与细胞骨架动力学的变化一起进行了研究。在瞬时转染的HEK 293细胞中，大量AQP‐eGFP与mCherry‐ Rab 5阳性早期内体和mCherry‐ Rab 11阳性再循环内体共定位。当暴露于低渗条件下时，AQP 4 ‐eGFP迅速从细胞内囊泡转移到细胞表面。显性阴性形式的mCherry-Rab 5和-Rab 11与AQP 4-eGFP的共表达阻止了低渗诱导的AQP 4-eGFP运输，并分别导致在细胞表面或细胞内囊泡处的浓缩。内吞抑制药物的使用表明AQP 4内化是动力蛋白依赖性的。细胞骨架动力学修饰药物也影响AQP 4往返细胞表面的转运。在表达高水平内源性AQP 4的原代人星形胶质细胞中验证了AQP 4运输机制。结果突出了早期和回收内体和细胞骨架动力学在响应低渗和缺氧应激的AQP 4易位中的作用，并表明在生理条件下胞内囊泡和细胞表面之间的AQP 4的连续循环。
The aquaporin‐4 (AQP4) water channel is abundantly expressed in the glial cells of the central nervous system and facilitates brain swelling following diverse insults, such as traumatic injury or stroke. Lack of specific and therapeutic AQP4 inhibitors highlights the need to explore alternative routes to control the water permeability of glial cell membranes. The cell surface abundance of AQP4 in mammalian cells fluctuates rapidly in response to changes in oxygen levels and tonicity, suggesting a role for vesicular trafficking in its translocation to and from the cell surface. However, the molecular mechanisms of AQP4 trafficking are not fully elucidated. In this work, early and recycling endosomes were investigated as likely candidates of rapid AQP4 translocation together with changes in cytoskeletal dynamics. In transiently transfected HEK293 cells a significant amount of AQP‐eGFP colocalised with mCherry‐Rab5‐positive early endosomes and mCherry‐Rab11‐positive recycling endosomes. When exposed to hypotonic conditions, AQP4‐eGFP rapidly translocated from intracellular vesicles to the cell surface. Co‐expression of dominant negative forms of the mCherry‐Rab5 and ‐Rab11 with AQP4‐eGFP prevented hypotonicity‐induced AQP4‐eGFP trafficking and led to concentration at the cell surface or intracellular vesicles respectively. Use of endocytosis inhibiting drugs indicated that AQP4 internalisation was dynamin‐dependent. Cytoskeleton dynamics‐modifying drugs also affected AQP4 translocation to and from the cell surface. AQP4 trafficking mechanisms were validated in primary human astrocytes, which express high levels of endogenous AQP4. The results highlight the role of early and recycling endosomes and cytoskeletal dynamics in AQP4 translocation in response to hypotonic and hypoxic stress and suggest continuous cycling of AQP4 between intracellular vesicles and the cell surface under physiological conditions.