Assentoft, Mette5; Larsen, Brian R5; Olesen, Emma T B5; Fenton, Robert A4; MacAulay, Nanna5
1 Section VIII, Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, Københavns Universitet2 Morphogenesis and Differentiation Program, Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, Københavns Universitet3 Neuronal Signalling Lab, Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, Københavns Universitet4 unknown5 Neuronal Signalling Lab, Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, Københavns Universitet
Aquaporin 4 (AQP4) is the predominant water channel in the mammalian brain and is mainly expressed in the perivascular glial endfeet at the brain-blood interface. AQP4 serves as a water entry site during brain edema formation, and regulation of AQP4 may therefore be of therapeutic interest. Phosphorylation of aquaporins can regulate plasma membrane localization and, possibly, the unit water permeability via gating of the AQP channel itself. In vivo phosphorylation of six serine residues in the COOH terminus of AQP4 has been detected by mass spectrometry: Ser(276), Ser(285), Ser(315), Ser(316), Ser(321), and Ser(322). To address the role of these phosphorylation sites for AQP4 function, serine-to-alanine mutants were created to abolish the phosphorylation sites. All mutants were detected at the plasma membrane of transfected C6 cells, with the fraction of the total cellular AQP4 expressed at the plasma membrane of transfected C6 cells being similar between the wild-type (WT) and mutant forms of AQP4. Activation of protein kinases A, C, and G in primary astrocytic cultures did not affect the plasma membrane abundance of AQP4. The unit water permeability was determined for the mutant AQP4s upon heterologous expression in Xenopus laevis oocytes (along with serine-to-aspartate mutants of the same residues to mimic a phosphorylation). None of the mutant AQP4 constructs displayed alterations in the unit water permeability. Thus phosphorylation of six different serine residues in the COOH terminus of AQP4 appears not to be required for proper plasma membrane localization of AQP4 or to act as a molecular switch to gate the water channel.
American Journal of Physiology: Cell Physiology, 2014, Vol 307, Issue 10