Beta-cell lines derived from transgenic Cpe(fat)/Cpe(fat) mice are defective in carboxypeptidase E and proinsulin processing.
A spontaneous point mutation in the coding region of the carboxypeptidase E (CPE) gene in Cpe(fat)/Cpe(fat) mice affects proinsulin processing. Cell lines derived from the pancreatic beta-cells of Cpe(fat)/Cpe(fat) mice were generated by crossing C57BLKS/J-Cpe(fat)/+ mice with NOD mice expressing the simian virus 40 large T oncogene under the control of the rat insulin II promoter. Two cell lines, designated NIT-2 and NIT-3, were cultured from adenomatous islets obtained from F2 littermates and were compared with the NIT-1 cell line previously developed from mice with wild-type CPE. Electron microscopy of the cultured NIT-2 and -3 cells showed increased numbers of enlarged and electron-lucent granules compared with NIT-1 cells. Pro-CPE, but not the mature form of CPE, is present in NIT-2 and -3 cells, and neither pro-CPE nor CPE are secreted into the medium. Immunocytochemistry shows the pro-CPE to be localized to an endoplasmic reticulum-like structure in NIT-3 cells. Proinsulin is less extensively processed in NIT-2 and -3 cells than in NIT-1 cells, indicating that the Cpe(fat) mutation affects both the endopeptidase and carboxypeptidase reactions. The secretion of insulin/proinsulin from NIT-2 and -3 cells is significantly elevated by secretagogues, indicating that CPE is not required for sorting proinsulin into the regulated pathway.
Varlamov
O
O
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Fricker
L D
LD
Furukawa
H
H
Steiner
D F
DF
Langley
S H
SH
Leiter
E H
EH
D000818:Animals; D015153:Blotting, Western; D043423:Carboxypeptidase H; D002268:Carboxypeptidases / Q000235:genetics* / Q000378:metabolism*; D002460:Cell Line; D004792:Enzyme Precursors / Q000378:metabolism; D007150:Immunohistochemistry; D007328:Insulin / Q000096:biosynthesis; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D016688:Mice, Inbred NOD; D008822:Mice, Transgenic / Q000235:genetics* / Q000378:metabolism*; D008854:Microscopy, Electron; D017354:Point Mutation; D011384:Proinsulin / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D051381:Rats
D004792:Enzyme Precursors; D007328:Insulin; D011384:Proinsulin; D002268:Carboxypeptidases; C094214:procarboxypeptidase E; D043423:Carboxypeptidase H