title: Not all secretory granules are created equal: Partitioning of soluble content proteins.
abstract: Secretory granules carrying fluorescent cargo proteins are widely used to study granule biogenesis, maturation, and regulated exocytosis. We fused the soluble secretory protein peptidylglycine alpha-hydroxylating monooxygenase (PHM) to green fluorescent protein (GFP) to study granule formation. When expressed in AtT-20 or GH3 cells, the PHM-GFP fusion protein partitioned from endogenous hormone (adrenocorticotropic hormone, growth hormone) into separate secretory granule pools. Both exogenous and endogenous granule proteins were stored and released in response to secretagogue. Importantly, we found that segregation of content proteins is not an artifact of overexpression nor peculiar to GFP-tagged proteins. Neither luminal acidification nor cholesterol-rich membrane microdomains play essential roles in soluble content protein segregation. Our data suggest that intrinsic biophysical properties of cargo proteins govern their differential sorting, with segregation occurring during the process of granule maturation. Proteins that can self-aggregate are likely to partition into separate granules, which can accommodate only a few thousand copies of any content protein; proteins that lack tertiary structure are more likely to distribute homogeneously into secretory granules. Therefore, a simple "self-aggregation default" theory may explain the little acknowledged, but commonly observed, tendency for both naturally occurring and exogenous content proteins to segregate from each other into distinct secretory granules.
authors.affiliation: Department of Neuroscience, University of Connecticut Health Center, Farmington, CT 06030-3401, USA.
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Sobota
authors.forename: Francesco
authors.initials: F
authors.lastname: Ferraro
authors.forename: Nils
authors.initials: N
authors.lastname: Bäck
authors.forename: Betty A
authors.initials: BA
authors.lastname: Eipper
authors.forename: Richard E
authors.initials: RE
authors.lastname: Mains
mesh_terms: D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000378:metabolism; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism; D011506:Proteins / Q000378:metabolism* / Q000648:ultrastructure; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D022142:Secretory Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D012995:Solubility; D014162:Transfection; D014617:Vacuoles / Q000648:ultrastructure
chemical_list: D009097:Multienzyme Complexes; D011506:Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D000324:Adrenocorticotropic Hormone; D002784:Cholesterol; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase