Inhibition of WNT signaling by G protein-coupled receptor (GPCR) kinase 2 (GRK2).
Activation of Wnt signaling pathways causes release and stabilization of the transcription regulator beta-catenin from a destruction complex composed of axin and the adenomatous polyposis coli (APC) protein (canonical signaling pathway). Assembly of this complex is facilitated by a protein-protein interaction between APC and a regulator of G protein signaling (RGS) domain in axin. Because G protein-coupled receptor kinase 2 (GRK2) has a RGS domain that is closely related to the RGS domain in axin, we determined whether GRK2 regulated canonical signaling. We found that GRK2 inhibited Wnt1-induced activation of a reporter construct as well as reduced Wnt3a-dependent stabilization and nuclear translocation of beta-catenin. GRK2 enzymatic activity was required for this negative regulatory effect, and depletion of endogenous GRK2 using small interfering RNA enhanced canonical signaling. GRK2-dependent inhibition of canonical signaling is relevant to osteoblast (OB) biology because overexpression of GRK2 attenuated Wnt/beta-catenin signaling in calvarial OBs. Coimmunoprecipitation studies found that: 1) GRK2 bound APC; 2) The GRK2-APC interaction was promoted by GRK2 enzymatic activity; and 3) Deletion of the RGS domain in GRK2 prevented both the GRK2-APC interaction and GRK2-dependent inhibition of canonical signaling. These data suggest that: 1) GRK2 negatively regulates Wnt signaling; 2) GRK2-dependent inhibition of canonical signaling requires a protein-protein interaction between the RGS domain in GRK2 and APC; and 3) Enzymatic activity promotes the GRK2-APC interaction and is required for the negative regulatory effect on canonical signaling. We speculate that inhibiting GRK2 activity in bone-forming OBs might be a useful therapeutic strategy for increasing bone mass.
Wang
Liming
L
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Gesty-Palmer
Diane
D
Fields
Timothy A
TA
Spurney
Robert F
RF
D000818:Animals; D001842:Bone and Bones / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D017077:Culture Media, Conditioned; D054769:G-Protein-Coupled Receptor Kinase 2 / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D051379:Mice; D008954:Models, Biological; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D051153:Wnt Proteins / Q000037:antagonists & inhibitors*; D051176:beta Catenin / Q000378:metabolism
D017077:Culture Media, Conditioned; D034741:RNA, Small Interfering; D051153:Wnt Proteins; D051176:beta Catenin; D054769:G-Protein-Coupled Receptor Kinase 2