Protein kinase C (PKC) phosphorylation of the Ca2+ o-sensing receptor (CaR) modulates functional interaction of G proteins with the CaR cytoplasmic tail.
The extracellular calcium (Ca(2+)(o))-sensing receptor (CaR) activates Ca(2+) influx independent of the release of intracellular Ca(2+) stores. The latter can be negatively regulated by protein kinase C (PKC) through phosphorylation of Thr-888 of the CaR. In this study, we substituted Thr-888 with various amino acid residues or a stop codon to understand how PKC phosphorylation of the CaR inhibits receptor-mediated release of intracellular Ca(2+) stores. Substitutions of Thr-888 with hydrophobic and hydrophilic amino acid residues had various effects on CaR-mediated release of intracellular Ca(2+) stores as well as activation of Ca(2+) influx. Several point mutations, such as T888D, had marked negative effects on CaR-mediated release of intracellular Ca(2+) stores but not on phorbol myristate acetate-insensitive activation of Ca(2+) influx. Presumably, the negatively charged aspartate mimics phospho-threonine. Interestingly, truncating the receptor at 888 had an even more pronounced negative effect on CaR-elicited release of intracellular Ca(2+) stores without significantly affecting CaR-mediated activation of Ca(2+) influx. Therefore, truncation at position 888 of the CaR affects the activity of the receptor in a manner that resembles PKC phosphorylation of the CaR. This in turn suggests that PKC phosphorylation of the CaR prevents G protein subtypes from interacting with the region of the receptor critical for releasing Ca(2+) stores, which is missing in the truncated receptor.
Jiang
Yong-Feng
YF
Endocrine-Hypertension Division, Department of Medicine, Brigham and Women's Hospital, and Harvard Medical School, Boston, Massachusetts 02115, USA.
Zhang
Zaixiang
Z
Kifor
Olga
O
Lane
Charles R
CR
Quinn
Stephen J
SJ
Bai
Mei
M
D019943:Amino Acid Substitution; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D020013:Calcium Signaling / Q000502:physiology*; D002460:Cell Line; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017354:Point Mutation; D011493:Protein Kinase C / Q000378:metabolism*; D044169:Receptors, Calcium-Sensing; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D013096:Spermine / Q000494:pharmacology; D013912:Threonine; D014162:Transfection
D044169:Receptors, Calcium-Sensing; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D013096:Spermine; D013912:Threonine; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D002118:Calcium