Secretogranin III binds to cholesterol in the secretory granule membrane as an adapter for chromogranin A.
Granin-family proteins, including chromogranin A (CgA) and secretogranin III (SgIII), are transported to secretory granules (SGs) in neuroendocrine cells. We previously showed that SgIII binds strongly to CgA in an intragranular milieu and targets CgA to SGs in pituitary and pancreatic endocrine cells. In this study, we demonstrated that with a sucrose density gradient of rat insulinoma-derived INS-1 cell homogenates, SgIII was localized to the SG fraction and was fractionated to the SG membrane (SGM) despite lacking the transmembrane region. With depletion of cholesterol from the SGM using methyl-beta-cyclodextrin, SgIII was impaired to bind to the SGM. Both SgIII and CgA were solubilized from the SGM by Triton X-100 in contrast to the Triton X-100 insolubility of carboxypeptidase E. SgIII and carboxypeptidase E strongly bound to the SGM-type liposome in intragranular conditions, but CgA did not. Instead, CgA bound to the SGM-type liposome only in the presence of SgIII. Immunocytochemical and pulse-chase experiments revealed that SgIII deleting the N-terminal lipid-binding region missorted to the constitutive pathway in mouse corticotroph-derived AtT-20 cells. Thus, we suggest that SgIII directly binds to cholesterol components of the SGM and targets CgA to SGs in pituitary and pancreatic endocrine cells.
Department of Molecular Medicine, Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512, Japan.
Masahiro
M
Hosaka
Masayuki
M
Suda
Yuko
Y
Sakai
Tetsuro
T
Izumi
Tsuyoshi
T
Watanabe
Toshiyuki
T
Takeuchi
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000378:metabolism; D000818:Animals; D043423:Carboxypeptidase H / Q000378:metabolism; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D002784:Cholesterol / Q000378:metabolism; D053379:Chromogranin A; D002864:Chromogranins / Q000737:chemistry* / Q000378:metabolism; D003505:Cyclodextrins / Q000737:chemistry; D003902:Detergents / Q000494:pharmacology; D004703:Endocrine System / Q000378:metabolism; D007150:Immunohistochemistry; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D017830:Octoxynol / Q000494:pharmacology; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D022142:Secretory Vesicles / Q000378:metabolism*; D013347:Subcellular Fractions; D013395:Sucrose / Q000494:pharmacology; D014981:Xenopus; D047392:beta-Cyclodextrins
D053379:Chromogranin A; D002864:Chromogranins; D003505:Cyclodextrins; D003902:Detergents; D008055:Lipids; D008081:Liposomes; D011506:Proteins; D012333:RNA, Messenger; D047392:beta-Cyclodextrins; C506155:chromogranin A, mouse; C506153:chromogranin A, rat; C108732:methyl-beta-cyclodextrin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D013395:Sucrose; D017830:Octoxynol; D002784:Cholesterol; D043423:Carboxypeptidase H