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Aquaporin‐2 interactome in rat inner medullary collecting duct (1137.5)
Author(s) -
Chou ChungLin,
Hageman Daniel,
Knepper Mark
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.1137.5
Subject(s) - aquaporin 2 , vasopressin , aquaporin , immunoprecipitation , microbiology and biotechnology , rab , chemistry , biochemistry , biology , gtpase , water channel , endocrinology , gene , mechanical engineering , engineering , inlet
Vasopressin regulates membrane trafficking of the water channel aquaporin‐2 (AQP2). We ran LC‐MS/MS analysis to identify AQP2 interacting proteins in native IMCD cells. Freshly isolated rat IMCD suspensions were treated with a vasopressin analog (dDAVP) or vehicle followed by in‐cell crosslinking using a homobifunctional crosslinker BSOCOES, detergent solubilization, AQP2 immunoprecipitation, SDS‐PAGE separation, and quantitative mass spectrometry of trypsin digests of gel pieces. The peptides passing stringent spectral quality thresholds were quantified (label‐free) to identify those with high signal:noise (AQP2 antibody/preimmune IgG > 4). The overall analysis identified a total of 89 replicable AQP2‐interacting proteins distributed throughout several cellular compartments (based on Gene Ontology analysis). 76 were found in dDAVP‐treated samples and 66 in vehicle‐treated samples (overlap 53). 16 have previously been found to undergo changes in phosphorylation in response to vasopressin including proteins involved in membrane trafficking (Lrba, Ndrg1, Sec61b), signaling (β‐catenin, inositol 3‐phosphate synthase, wolframin), cell‐cell adhesion (α‐catenin, β‐catenin, periplakin) and transport (Slc14a2, Atp1a1). Additionally, four distinct Rab proteins were identified (5C,7A,14,17). These findings provide extensive data for modeling vasopressin‐regulated AQP2 trafficking. Grant Funding Source : Supported by NHLBI