Phosphorylation in the C-terminal domain of Aquaporin-4 is required for Golgi transition in primary cultured astrocytes.
Aquaporin-4 (AQP4) is expressed in the perivascular and subpial astrocytes end-feet in mammalian brain, and plays a critical component of an integrated water and potassium homeostasis. Here we examine whether AQP4 is phosphorylated in primary cultured mouse astrocytes. Astrocytes were metabolically labeled with [(32)P]phosphoric acid, then AQP4 was immunoprecipitated with anti-AQP4 antibody. We observed that AQP4 was constitutively phosphorylated, which is reduced by treatment with protein kinase CK2 inhibitors. To elucidate the phosphorylation of AQP4 by CK2, myc-tagged wild-type or mutant AQP4 was transiently transfected in primary cultured astrocytes. Substitution of Ala residues for four putative CK2 phosphorylation sites in the C terminus abolished the phosphorylation of AQP4. Immunofluorescent microscopy revealed that the quadruple mutant was localized in the Golgi apparatus. These observations indicate that the C-terminal domain of AQP4 is constitutively phosphorylated at least in part by protein kinase CK2 and it is required for Golgi transition.
Kadohira
Ikuko
I
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan; Department of Obstetrics and Gynecology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Abe
Yoichiro
Y
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan. Electronic address: yoabe@sc.itc.keio.ac.jp.
Nuriya
Mutsuo
M
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Sano
Kazumi
K
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Tsuji
Shoji
S
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Arimitsu
Takeshi
T
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan; Department of Pediatrics, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Yoshimura
Yasunori
Y
Department of Obstetrics and Gynecology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Yasui
Masato
M
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D051401:Aquaporin 4 / Q000235:genetics / Q000378:metabolism*; D001253:Astrocytes / Q000378:metabolism / Q000648:ultrastructure; D047390:Casein Kinase II / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D051379:Mice; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D021381:Protein Transport
C496030:AQP4 protein, human; D051401:Aquaporin 4; D047390:Casein Kinase II