Characterization of vesicles secreted from insulinoma NIT-1 cells.
Insulinoma NIT-1, an insulin-secreting mouse cell line, secretes vesicles in response to glucose or calcium. These vesicles, like exosomes, are relatively homogeneous (30-100 nm). We analyzed their protein profiles employing one-dimensional SDS gel electrophoresis combined with nanoLC-ESI-q-TOF tandem mass spectrometry, and searched for post-translational modifications (PTMs) using MOD(i) algorithm. We identified 270 proteins which matched at least two peptides reproducibly in duplicate runs. These proteins included metabolic proteins, endocytosis/exocytosis related proteins, chaperones, cytoskeletal proteins, membrane transporters/ion channels, signaling molecules, and nucleic acid binding proteins. Over 200 of these are newly identified proteins for the first time in secreted vesicles, and included RNA- and translation-related proteins, ubiquitin- and protein-degradation related proteins and post-translationally modified proteins. The rest of the proteins identified in this study were similar to those reported by others to be present in exosomes of various origins. The present study demonstrates that vesicles secreted from insulinoma NIT-1 cells have some properties, common to exosomes from lymphocytes and cancer cells, and some differing from those of other types of exosomes. We believe that the modified and newly identified proteins we identified in secreted vesicles from insulinoma NIT-1 cells have the potential to provide insights into mechanisms of biogenesis and function of secreted vesicles and may help explain the impairment of insulin secretion in islets from type-2 diabetes.
Center for Cell Signaling & Drug Discovery Research, College of Pharmacy and Division of Life & Pharmaceutical Sciences, Ewha Womans University, Seoul, Korea 120-750.
Hyo Sun
HS
Lee
Jaeho
J
Jeong
Kong-Joo
KJ
Lee
D000818:Animals; D015153:Blotting, Western; D002458:Cell Fractionation / Q000379:methods; D045744:Cell Line, Tumor; D050417:Insulin-Secreting Cells / Q000378:metabolism; D007340:Insulinoma / Q000378:metabolism*; D051379:Mice; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000032:analysis* / Q000145:classification; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D022142:Secretory Vesicles / Q000737:chemistry*; D053719:Tandem Mass Spectrometry
D011506:Proteins; D020543:Proteome