Intracellular transport and sorting of mutant human proinsulins that fail to form hexamers.
Human proinsulin and insulin oligomerize to form dimers and hexamers. It has been suggested that the ability of prohormones to self associate and form aggregates may be responsible for the sorting process at the trans-Golgi. To examine whether insulin oligomerization is required for proper sorting into regulated storage granules, we have constructed point mutations in human insulin B chain that have been previously shown to prevent formation of insulin hexamers (Brange, J., U. Ribel, J. F. Hansen, G. Dodson, M. T. Hansen, S. Havelund, S. G. Melberg, F. Norris, K. Norris, L. Snel, A. R. Sorensen, and H. O. Voight. 1988. Nature [Lond.]. 333:679-682). One mutant (B10His----Asp) allows formation of dimers but not hexamers and the other (B9Ser----Asp) prevents formation of both dimers and hexamers. The mutants were transfected into the mouse pituitary AtT-20 cells, and their ability to be sorted into regulated secretory granules was compared to wild-type insulin. We found that while B10His----Asp is sorted somewhat less efficiently than wild-type insulin as reported previously (Carroll, R. J., R. E. Hammer, S. J. Chan, H. H. Swift, A. H. Rubenstein, and D. F. Steiner. 1988. Proc. Natl. Acad. Sci. USA. 85:8943-8947; Gross, D. J., P. A. Halban, C. R. Kahn, G. C. Weir, and L. Villa-Kumaroff. 1989. Proc. Natl. Acad. Sci. USA. 86:4107-4111). B9Ser----Asp is targeted to granules as efficiently as wild-type insulin. These results indicate that self association of proinsulin into hexamers is not required for its targeting to the regulated secretory pathway.
Quinn
D
D
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Orci
L
L
Ravazzola
M
M
Moore
H P
HP
D000595:Amino Acid Sequence; D001483:Base Sequence; D002451:Cell Compartmentation; D002460:Cell Line; D006056:Golgi Apparatus / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D007328:Insulin / Q000235:genetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D010957:Plasmids; D011384:Proinsulin / Q000235:genetics* / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D014162:Transfection
D007328:Insulin; D046911:Macromolecular Substances; D015345:Oligonucleotide Probes; D011384:Proinsulin