Molecular cloning, developmental expression and pharmacological characterization of the CCKB/gastrin receptor in the calf pancreas.
We have cloned the calf predominant pancreatic cholecystokinin B (CCKB)/gastrin receptor cDNA. It encodes a 454 amino acid protein with 90% identity with the CCKB/gastrin receptor cloned in other species and tissues. However, the calf pancreatic CCKB/gastrin receptor contains a pentapeptide cassette within the third intracellular loop which is absent in the cloned human brain and stomach receptor. Quantification of the CCKB/gastrin receptor mRNA levels by reverse transcription polymerase chain reaction demonstrated the same level of transcripts at birth, +7 and +28 days. On the other hand, binding study with pancreatic membranes showing a dramatic increase (600-fold) in the number of CCKB/gastrin receptor sites between at birth and +28 days indicates that the development of the calf pancreatic CCKB/gastrin receptor occurs during the first 4 weeks of post-natal life. COS monkey cells (COS-7 cells) transiently transfected by the cloned cDNA exhibit binding of 125I-Bolton-Hunter-[Thr28,Ahx31]CCK-(25-33) and 125I-Bolton-Hunter-[Leu15]human gastrin-(2-17) to two affinity classes of sites. Kd values of the high affinity binding components indicate a 4-fold higher affinity of the receptor for sulfated gastrin than for CCK. Finally, the recombinant receptor is coupled to G proteins and [Ca2+]i mobilization, and is expressed as a glycoprotein of 82 kDa.
Dufresne
M
M
Institut National de la Santé et de la Recherche Médicale, Unité 151, Institut Louis Bugnard, Centre Hospitalier Universitaire de Rangueil, Toulouse, France.
Escrieut
C
C
Clerc
P
P
Le Huerou-Luron
I
I
Prats
H
H
Bertrand
V
V
Le Meuth
V
V
Guilloteau
P
P
Vaysse
N
N
Fourmy
D
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002460:Cell Line; D003001:Cloning, Molecular; D019204:GTP-Binding Proteins / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010179:Pancreas / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis; D011869:Radioligand Assay; D011949:Receptors, Cholecystokinin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D012333:RNA, Messenger; D011949:Receptors, Cholecystokinin; D019204:GTP-Binding Proteins