739 Original Paper Cell Physiol Biochem 2011;27:739-748 Accepted: May 03, 2011 Cellular Physiology Cellular Physiology Cellular Physiology Cellular Physiology Cellular Physiology and Biochemistr and Biochemistr and Biochemistr and Biochemistr and Biochemistry Copyright © 2011 S. Karger AG, Basel Fax +41 61 306 12 34 E-Mail karger@karger.ch www.karger.com © 2011 S. Karger AG, Basel 1015-8987/11/0276-0739$38.00/0 Accessible online at: www.karger.com/cpb Integrin Signaling Modulates AQP2 Trafficking via Arg-Gly-Asp (RGD) Motif Grazia Tamma 1 , Domenica Lasorsa 1 , Marianna Ranieri 1 , Lisa Mastrofrancesco 1 , Giovanna Valenti 1 and Maria Svelto 1,2 1 Department of General and Environmental Physiology, University of Bari, Bari, 2 Centro di Eccellenza di Genomica in campo Biomedico ed Agrario (CEGBA) Dr. Grazia Tamma Department of General and Environmental Physiology University of Bari, Via Amendola 165/A, 70125 Bari (Italy) Tel. +39 080 5443414, Fax+39 080 5443388 E-Mail g.tamma@biologia.uniba.it Key Words Aquaporins • Integrin • Trafficking Abstract Aquaporin-2 (AQP2) increases the water permeability of renal collecting ducts in response to vasopressin. Vasopressin stimulation is accompanied by a profound remodeling of actin cytoskeleton whose dynamics are regulated by crosstalk between intracellular and extracellular signals. Here, we report that AQP2 contains a conserved RGD domain in its external C- loop. Co-immunoprecipitation experiments demonstrated that AQP2 binds integrin 1 in renal tissue and in MCD4 cells. To investigate the role of this interaction on AQP2 trafficking, cells were exposed to synthetic RGD-containing peptides, GRGDNP or GRGDSP, able to bind certain integrins. Incubation with these peptides increased the membrane expression of AQP2 in the absence of hormonal stimulation as assessed by confocal analysis and cell surface biotinylation. To identify the signals underlying the effects of peptides on AQP2 trafficking, some possible intracellular messengers were evaluated. Exposure of MCD4 cells to GRGDNP increased intracellular cAMP as assessed by FRET studies while GRGDSP increased intracellular calcium concentration. Taken together, these data propose integrins as new players controlling the cellular localization of AQP2, via two distinct signal transduction pathways dependent on cAMP and calcium respectively. Introduction Renal collecting duct water permeability is regulated by the hormone vasopressin which controls the expression [1] and cellular distribution of the water channel AQP2 [2-4]. Vasopressin binds its cognate receptor V2R stimulating a cAMP-dependent signal transduction cascade leading to the activation of protein kinase A (PKA) which phosphorylates AQP2 at serine 256 [5-7]. This event is essential to promote the translocation of AQP2-bearing vesicles to the apical plasma membrane, thus ensuring collecting duct water reabsorption [5].