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.
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.
Ikuko
I
Kadohira
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan. Electronic address: yoabe@sc.itc.keio.ac.jp.
Yoichiro
Y
Abe
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Mutsuo
M
Nuriya
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Kazumi
K
Sano
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Shoji
S
Tsuji
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.
Takeshi
T
Arimitsu
Department of Obstetrics and Gynecology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Yasunori
Y
Yoshimura
Department of Pharmacology, School of Medicine, Keio University, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
Masato
M
Yasui
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