G Protein-coupled receptor kinases phosphorylate LRP6 in the Wnt pathway.
Wnt ligands conduct their functions in canonical Wnt signaling by binding to two receptors, the single transmembrane low density lipoprotein receptor-related proteins 5 and 6 (LRP5/6) and seven transmembrane (7TM) Frizzled receptors. Subsequently, phosphorylation of serine/threonine residues within five repeating signature PPPSP motifs on LRP6 is responsible for LRP6 activation. GSK3beta, a cytosolic kinase for phosphorylation of a downstream effector beta-catenin, was proposed to participate in such LRP6 phosphorylation. Here, we report a new class of membrane-associated kinases for LRP6 phosphorylation. We found that G protein-coupled receptor kinases 5 and 6 (GRK5/6), traditionally known to phosphorylate and desensitize 7TM G protein-coupled receptors, directly phosphorylate the PPPSP motifs on single transmembrane LRP6 and regulate Wnt/LRP6 signaling. GRK5/6-induced LRP6 activation is inhibited by the LRP6 antagonist Dickkopf. Depletion of GRK5 markedly reduces Wnt3A-stimulated LRP6 phosphorylation in cells. In zebrafish, functional knock-down of GRK5 results in reduced Wnt signaling, analogous to LRP6 knock-down, as assessed by decreased abundance of beta-catenin and lowered expression of the Wnt target genes cdx4, vent, and axin2. Expression of GRK5 rescues the diminished beta-catenin and axin2 response caused by GRK5 depletion. Thus, our findings identify GRK5/6 as novel kinases for the single transmembrane receptor LRP6 during Wnt signaling.
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Minyong
M
Chen
Melanie
M
Philipp
Jiangbo
J
Wang
Richard T
RT
Premont
Tiffany R
TR
Garrison
Marc G
MG
Caron
Robert J
RJ
Lefkowitz
Wei
W
Chen
D000595:Amino Acid Sequence; D000818:Animals; D060466:Axin Protein; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D051157:Frizzled Receptors / Q000235:genetics / Q000378:metabolism; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000235:genetics / Q000378:metabolism*; D054768:G-Protein-Coupled Receptor Kinases / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D060488:Low Density Lipoprotein Receptor-Related Protein-6; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism*; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
D060466:Axin Protein; D051157:Frizzled Receptors; D018398:Homeodomain Proteins; C555475:LRP6 protein, human; D060488:Low Density Lipoprotein Receptor-Related Protein-6; C555476:Lrp6 protein, mouse; D016376:Oligonucleotides, Antisense; D011973:Receptors, LDL; D012097:Repressor Proteins; D051153:Wnt Proteins; D029961:Zebrafish Proteins; D051176:beta Catenin; D054774:G-Protein-Coupled Receptor Kinase 5; D054768:G-Protein-Coupled Receptor Kinases; C089176:G-protein-coupled receptor kinase 6