Aggregation and lack of secretion of most newly synthesized proinsulin in non-beta-cell lines.
Myoblasts transfected with HB10D insulin secrete more hormone than those transfected with wild-type insulin, as published previously, indicating that production of wild-type insulin is not efficient in these cells. The ability of non-beta-cells to produce insulin was examined in several cell lines. In clones of neuroendocrine GH(4)C(1) cells stably transfected with proinsulin, two thirds of (35)S-proinsulin was degraded within 3 h of synthesis, whereas (35)S-prolactin was stable. In transiently transfected neuroendocrine AtT20 cells, half of (35)S-proinsulin was degraded within 3 h after synthesis, whereas (35)S-GH was stable. In transiently transfected fibroblast COS cells, (35)S-proinsulin was stable for longer, but less than 10% was secreted 8 h after synthesis. Proinsulin formed a concentrated patch detected by immunofluorescence in transfected cells that did not colocalize with calreticulin or BiP, markers for the endoplasmic reticulum, but did colocalize with membrin, a marker for the cis-medial Golgi complex. Proinsulin formed a Lubrol-insoluble aggregate within 30 min after synthesis in non-beta-cells but not in INS-1E cells, a beta-cell line that normally produces insulin. More than 45% of (35)S-HB10D proinsulin was secreted from COS cells 3 h after synthesis, and this mutant formed less Lubrol-insoluble aggregate in the cells than did wild-type hormone. These results indicate that proinsulin production from these non-beta-cells is not efficient and that proinsulin aggregates in their secretory pathways. Factors in the environment of the secretory pathway of beta-cells may prevent aggregation of proinsulin to allow efficient production.
Zhu
Yong Lian
YL
Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520-8066, USA.
Abdo
Alexander
A
Gesmonde
Joan F
JF
Zawalich
Kathleen C
KC
Zawalich
Walter
W
Dannies
Priscilla S
PS
D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D007150:Immunohistochemistry; D011384:Proinsulin / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D014162:Transfection; D015032:Zinc / Q000494:pharmacology
D011384:Proinsulin; D015032:Zinc