title: Preferential release of newly synthesized insulin assessed by a multi-label reporter system using pancreatic β-cell line MIN6.
abstract: Newly synthesized hormones have been suggested to be preferentially secreted by various neuroendocrine cells. This observation indicates that there is a distinct population of secretory granules containing new and old hormones. Recent development of fluorescent timer proteins used in bovine adrenal chromaffin cells revealed that secretory vesicles segregate into distinct age-dependent populations. Here, we verify the preferential release of newly synthesized insulin in the pancreatic β-cell line, MIN6, using a combination of multi-labeling reporter systems with both fluorescent and biochemical procedures. This system allows hormones or granules of any age to be labeled, in contrast to the timer proteins, which require fluorescence shift time. Pulse-chase labeling with different color probes distinguishes insulin secretory granules by age, with younger granules having a predominantly intracellular localization rather than at the cell periphery.
authors.affiliation: Institute for Molecular and Cellular Regulation, Gunma University, Maebashi, Gunma, Japan.
authors.forename: Ni
authors.initials: N
authors.lastname: Hou
authors.forename: Hideo
authors.initials: H
authors.lastname: Mogami
authors.forename: Chisato
authors.initials: C
authors.lastname: Kubota-Murata
authors.forename: Meng
authors.initials: M
authors.lastname: Sun
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Takeuchi
authors.forename: Seiji
authors.initials: S
authors.lastname: Torii
mesh_terms: D000818:Animals; D001671:Biochemistry / Q000379:methods; D002460:Cell Line; D055098:Endocrine Cells / Q000166:cytology; D005456:Fluorescent Dyes / Q000494:pharmacology; D017930:Genes, Reporter; D005947:Glucose / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000166:cytology*; D007340:Insulinoma / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods; D010957:Plasmids / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D007328:Insulin; D005947:Glucose