Faculty Research 1990 - 1999

Title

Beta-cell lines derived from transgenic Cpe(fat)/Cpe(fat) mice are defective in carboxypeptidase E and proinsulin processing.

Document Type

Article

Publication Date

1997

Keywords

Blotting-Western, Carboxypeptidases: ge, me, Cell-Line, Enzyme-Precursors: me, Immunohistochemistry, Insulin: bi, Islets-of-Langerhans: cy, me, Mice, Mice-Inbred-C57BL, Mice-Inbred-NOD, Mice-Transgenic: ge, me, Microscopy-Electron, Point-Mutation, Proinsulin: me, Protein-Processing-Post-Translational: ph, Rats, SUPPORT-NON-U-S-GOVT, SUPPORT-U-S-GOVT-P-H-S

JAX Source

Endocrinology 1997 Nov;138(11):4883-92

Grant

DK51271/DK/NIDDK, DA04494/DA/NIDA, DA00194/DA/NIDA

Abstract

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.

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