title: High-throughput luminescent reporter of insulin secretion for discovering regulators of pancreatic Beta-cell function.
abstract: Defects in insulin secretion play a central role in the pathogenesis of type 2 diabetes, yet the mechanisms driving beta-cell dysfunction remain poorly understood, and therapies to preserve glucose-dependent insulin release are inadequate. We report a luminescent insulin secretion assay that enables large-scale investigations of beta-cell function, created by inserting Gaussia luciferase into the C-peptide portion of proinsulin. Beta-cell lines expressing this construct cosecrete luciferase and insulin in close correlation, under both standard conditions or when stressed by cytokines, fatty acids, or ER toxins. We adapted the reporter for high-throughput assays and performed a 1,600-compound pilot screen, which identified several classes of drugs inhibiting secretion, as well as glucose-potentiated secretagogues that were confirmed to have activity in primary human islets. Requiring 40-fold less time and expense than the traditional ELISA, this assay may accelerate the identification of pathways governing insulin secretion and compounds that safely augment beta-cell function in diabetes.
authors.affiliation: Diabetes Unit of the Department of Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Center for Human Genetic Research, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Molecular Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Medical and Population Genetics Program, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA. Electronic address: sburns@broadinstitute.org.
authors.forename: Sean M
authors.initials: SM
authors.lastname: Burns
authors.affiliation: Center for the Science of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Amedeo
authors.initials: A
authors.lastname: Vetere
authors.affiliation: Center for the Science of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Deepika
authors.initials: D
authors.lastname: Walpita
authors.affiliation: Center for the Science of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Vlado
authors.initials: V
authors.lastname: Dančík
authors.affiliation: Center for the Development of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Carol
authors.initials: C
authors.lastname: Khodier
authors.affiliation: Center for the Development of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Jose
authors.initials: J
authors.lastname: Perez
authors.affiliation: Center for the Science of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Paul A
authors.initials: PA
authors.lastname: Clemons
authors.affiliation: Center for the Science of Therapeutics, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: Bridget K
authors.initials: BK
authors.lastname: Wagner
authors.affiliation: Diabetes Unit of the Department of Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Center for Human Genetic Research, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Molecular Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Genetics, Harvard Medical School, Boston, MA 02115, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA; Medical and Population Genetics Program, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA.
authors.forename: David
authors.initials: D
authors.lastname: Altshuler
mesh_terms: D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005227:Fatty Acids / Q000494:pharmacology*; D017930:Genes, Reporter; D005947:Glucose / Q000494:pharmacology; D057166:High-Throughput Screening Assays; D006801:Humans; D007328:Insulin / Q000235:genetics / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D019284:Thapsigargin / Q000633:toxicity
chemical_list: D016207:Cytokines; D005227:Fatty Acids; D007328:Insulin; D011993:Recombinant Fusion Proteins; D019284:Thapsigargin; D008156:Luciferases; D005947:Glucose