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G‐Protein Receptor Kinase 4γ interacts preferentially with inactive Gα s and Gα 13
Author(s) -
Keever Lindsay B,
Andresen Bradley T
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a429-b
Subject(s) - heterotrimeric g protein , g protein coupled receptor kinase , g protein coupled receptor , regulator of g protein signaling , g protein , beta adrenergic receptor kinase , microbiology and biotechnology , protein subunit , g beta gamma complex , internalization , phosphorylation , gi alpha subunit , immunoprecipitation , hek 293 cells , signal transduction , agonist , receptor , chemistry , kinase , effector , biology , biochemistry , gtpase activating protein , gene
G‐protein Receptor Kinases (GRKs) are responsible for agonist‐mediated G‐protein coupled receptor (GPCR) phosphorylation and consequently desensitization and internalization. This process is initiated by the activated Gα subunit of heterotrimeric G‐proteins interacting with the regulator of G‐protein signaling homology domain of the GRK in a manner similar to an effector. Our preliminary data indicated that wild type GRK4γ interacted with both inactive and active Gα s and Gα 13 . We hypothesized that this interaction is unique to GRK4 and that it affects agonist‐mediated signaling. To test this hypothesis, 293T cells were cotransfected with a Gα subunit and GRK4γ, or GRK2 as a positive control, and protein‐protein interactions were assessed by coimmunoprecipitation of the GRK with the Gα subunit. In control, GDP‐treated, as well as activated, AlF 4 − ‐treated, samples, GRK4γ preferentially binds to Gα s and Gα 13 irrespective of activation state. In contrast, GRK2 interacted with Gα q only in the activated state. Utilizing constitutively active mutants (QL mutations) of Gα s and Gα 13 as well as wild type controls, GRK4γ interacted with the wild type Gα subunit to a much greater extent than the constitutively active QL mutant. This interaction was preserved in detergent free buffer. These data suggest that GRK4γ acts in an opposite manner compared to GRK2. Functional assays indicated that both GRK2 and GRK4 inhibit agonist‐mediated cAMP accumulation; however, only GRK4γ increased basal levels of cAMP through a kinase dependent mechanism. In conclusion, GRK4γ interacts with Gα s in a novel manner, yet still regulates GPCR signal transduction similar to GRK2.