Expression of G protein alpha-subunits in bovine parathyroid.
A G protein-coupled Ca(2+)-sensing receptor was recently cloned from bovine parathyroid and shown to mediate divalent cation regulation of PTH secretion. To define which G proteins might be coupled to the Ca(2+)-sensing receptor in parathyroid cells, we determined which G protein alpha-subunit messenger RNAs (mRNAs) are expressed in the parathyroid. We also considered the possibility that a novel parathyroid-specific G alpha might be present. We, therefore, used the reverse transcription-polymerase chain reaction to study the expression of G alpha subunits in a bovine parathyroid mRNA preparation. Degenerate primers, corresponding to two regions conserved in every G alpha subunit, the G3 and G4 sequences, were used to amplify G alpha complementary DNA fragments that were subcloned and sequenced. We found that mRNAs corresponding to G alpha s, G alpha i2, G alpha 11, G alpha 12, and G alpha z are the predominant G alpha mRNAs expressed in the bovine parathyroid. No novel G alpha mRNA was identified. Northern blots confirmed the expression of the cloned G alpha subunits and showed lower expression of G alpha o and G alpha i1 mRNAs. Immunoblots confirmed abundant expression of G alpha s, G alpha i2, and G alpha 11 and provided evidence for expression of G alpha i1 and G alpha i3, but not G alpha o. G alpha q mRNA was not identified by the degenerate primer reverse transcription-polymerase chain reaction strategy, but the immunoblot detected G alpha q protein, albeit at considerably lower levels than G alpha 11. The abundance of G alpha 11 relative to G alpha q in bovine parathyroid is consistent with but does not prove a role for G alpha 11 in coupling the Ca(2+)-sensing receptor to phospholipase C.
Varrault
A
A
Metabolic Diseases Branch, National Institute of Diabetes and Digestive and Kidney Disease, National Institutes of Health, Bethesda, Maryland 20892, USA.
Pena
M S
MS
Goldsmith
P K
PK
Mithal
A
A
Brown
E M
EM
Spiegel
A M
AM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D003001:Cloning, Molecular; D019204:GTP-Binding Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D010280:Parathyroid Glands / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis
D012333:RNA, Messenger; D019204:GTP-Binding Proteins; D002118:Calcium