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Phospholipase C‐related inactive protein is implicated in the constitutive internalization of GABA A receptors mediated by clathrin and AP2 adaptor complex
Author(s) -
Kanematsu Takashi,
Fujii Makoto,
Mizokami Akiko,
Kittler Josef T.,
Nabekura Junichi,
Moss Stephen J.,
Hirata Masato
Publication year - 2007
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.2006.04399.x
Subject(s) - internalization , clathrin , biology , microbiology and biotechnology , receptor , endocytosis , enzyme linked receptor , receptor mediated endocytosis , immunoprecipitation , signal transducing adaptor protein , phosphorylation , biochemistry , gene
A mechanism for regulating the strength of synaptic inhibition is enabled by altering the number of GABA A receptors available at the cell surface. Clathrin and adaptor protein 2 (AP2) complex‐mediated endocytosis is known to play a fundamental role in regulating cell surface GABA A receptor numbers. Very recently, we have elucidated that phospholipase C‐related catalytically inactive protein (PRIP) molecules are involved in the phosphorylation‐dependent regulation of the internalization of GABA A receptors through association with receptor β subunits and protein phosphatases. In this study, we examined the implications of PRIP molecules in clathrin‐mediated constitutive GABA A receptor endocytosis, independent of phospho‐regulation. We performed a constitutive receptor internalization assay using human embryonic kidney 293 (HEK293) cells transiently expressed with GABA A receptor α/β/γ subunits and PRIP. PRIP was internalized together with GABA A receptors, and the process was inhibited by PRIP‐binding peptide which blocks PRIP binding to β subunits. The clathrin heavy chain, μ2 and β2 subunits of AP2 and PRIP‐1, were complexed with GABA A receptor in brain extract as analyzed by co‐immunoprecipitation assay using anti‐PRIP‐1 and anti‐β2/3 GABA A receptor antibody or by pull‐down assay using β subunits of GABA A receptor. These results indicate that PRIP is primarily implicated in the constitutive internalization of GABA A receptor that requires clathrin and AP2 protein complex.