This applicant has proposed a program of research to prepare her for a career in academic nephrology and basic science research in the field of renal physiology/pathophysiology, specifically, AQP2 trafficking. This applicant will propose to characterize AQP2 interacting proteins and investigate the physiological significance of these interactions, to better understand the molecular mechanism underlying AQP2 trafficking, and explore the possible novel role of AQP2 in cellular biological and pathophysiological processes. The research will be conducted in the laboratory of Dr. Dennis Brown at the Program in Membrane Biology (PMB) and Division of Nephrology, Massachusetts General Hospital. Vasopressin (VP) is the major antidiuretic hormone involved in the regulation of water reabsorption by mammalian kidney. It functions by recruiting the AQP2 water channel from cytoplasmic vesicles to the plasma membrane of collecting duct principal cells. The impairment of VP-AQP2 signaling pathways results in fluid retention seen in congestive heart failure, cirrhosis, as well as concentrating defect seen in diabetes insipidus. AQP2 is regulated through complex trafficking pathways which have not been well characterized. Our hypothesis is that regulated trafficking of AQP2 requires direct and indirect protein-protein interactions during intracellular translocation, exocytosis as well as endocytosis. Specifically, we will 1) extend our current study on the interaction of AQP2 and heat shock protein 70 (hsc70) by tracking further down to subcellular compartments/step(s) in AQP2 trafficking pathways, and continue to characterize other AQP2 binding candidates identified from our yeast-two hybrid screen;2) study the molecular mechanism underlying the role of phosphorylation and dephosphorylation on AQP2 trafficking;3) explore novel potential roles of AQP2 in cell adhesion, migration, and renal tubular formation/malformation. We will investigate the distribution and trafficking of AQP2 during the general biological processes as well as the effect of alteration of AQP2 trafficking on these biological processes. We will using stable or primary cell culture, tissue slice culture, 3D culture system, a zebrafish model system, and AQP2 mutant mouse model. Multidisciplinary approaches and powerful technologies will be used to carry out these studies that will shed light on the mechanism of AQP2 trafficking and function.
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