Water channel aquaporin-2 directly binds to actin.
Water channel aquaporin-2 (AQP2) strictly regulates body water homeostasis in mammals. Trafficking of AQP2 to the apical membrane is critical to the reabsorption of water in renal collecting ducts. Controlled apical positioning of AQP2 suggests the interaction of AQP2 with other proteins. To isolate AQP2-binding proteins, immunoaffinity chromatography of extracts from rat kidney papilla was performed using a column covalently coupled with anti-AQP2 antibody. Using this method 42-kDa protein was purified and subsequently identified as beta- and gamma-isoforms of actin by two-dimensional gel analysis and matrix-assisted laser desorption-ionization time-of-flight mass spectrometry. AQP2 was indeed coimmunoprecipitated with actin from cell lysates of rat kidney papilla. In addition, surface plasmon resonance analyses showed that the C-terminal fragment of AQP2 strongly bound to actin and the K(D) value was 3.18x10(-8)M. In this experiment we have elucidated the direct binding of channel protein AQP2 to cytoskeletal protein actin, providing a novel mechanism for trafficking of not only AQP2 but also recycling channel proteins.
Noda
Yumi
Y
Department of Nephrology, Tokyo Medical and Dental University, Tokyo 113-8519, Japan. ynodmed@tmd.ac.jp
Horikawa
Saburo
S
Katayama
Yoshifumi
Y
Sasaki
Sei
S
D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D051399:Aquaporin 2; D020346:Aquaporins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002846:Chromatography, Affinity; D015180:Electrophoresis, Gel, Two-Dimensional; D007668:Kidney / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D051381:Rats
D000199:Actins; C496025:Aqp2 protein, rat; D051399:Aquaporin 2; D020346:Aquaporins; D010446:Peptide Fragments