Bombesin activation of phospholipase C beta 1 in rat acinar pancreatic cells involves the pertussis toxin-sensitive G alpha i3 protein.
Bombesin stimulation of inositol 1,4,5-trisphosphate (Ins P3) formation in rat sonicated pancreatic acinar cells was inhibited by an antibody directed against the pertussis toxin (PTX)-sensitive GTP-binding G alpha i3 protein but not by an anti-G alpha q-11 antibody. After solubilization and gel filtration, [125I-Tyr4]bombesin binding sites were recovered in a peak of protein of 67 approximately 90 kDa with a maximal enrichment corresponding to a molecular mass of 83-kDa. Results obtained from the non-hydrolysable GTP analog guanosine-5'-[gamma-thio]triphosphate (GTP gamma S) binding, PTX-stimulated ADP-ribosylation and immunoblotting showed that the 83-kDa fraction contained the G alpha i3 protein but not the G alpha q-11 protein. Furthermore, GTP gamma S increased the bombesin binding dissociation constant (KD) from 0.32 to 0.60 nM, while the anti-G alpha i3 antibody decreased the maximal binding capacity (Bmax) from 50 to 25 fmol/mg protein without affecting the KD. Mixing solubilized bombesin binding sites with a phospholipase C (PLC) preparation from rat pancreas reconstituted a bombesin-stimulated PLC activity which was markedly inhibited by the anti-G alpha i3 antibody but unaffected by the anti-G alpha q-11 antibody. In addition, this stimulation was inhibited by an anti-PLC beta 1 antibody. This result supports the involvement of the PLC beta 1 isoform in bombesin receptor activation.
Pigeon
C
C
INSERM Unité 10, Institut Fédératif de Recherches Cellules Epithéliales, Hôpital Bichat-Claude Bernard, Paris, France.
Le Romancer
M
M
Linard
C
C
Lewin
M J
MJ
Reyl-Desmars
F
F
D000818:Animals; D001839:Bombesin / Q000494:pharmacology*; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D008297:Male; D010179:Pancreas / Q000378:metabolism*; D037342:Pertussis Toxin; D054799:Phospholipase C beta; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction / Q000187:drug effects*; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D007527:Isoenzymes; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D054799:Phospholipase C beta; C518035:Plcb1 protein, rat; D019204:GTP-Binding Proteins; D001839:Bombesin