A Ratiometric Sensor for Imaging Insulin Secretion in Single β Cells

Martina Schifferer, Dmytro A. Yushchenko, Frank Stein, Andrey Bolbat, Carsten Schultz

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

Despite the urgent need for assays to visualize insulin secretion there is to date no reliable method available for measuring insulin release from single cells. To address this need, we developed a genetically encoded reporter termed RINS1 based on proinsulin superfolder GFP (sfGFP) and mCherry fusions for monitoring insulin secretion. RINS1 expression in MIN6 β cells resulted in proper processing yielding single-labeled insulin species. Unexpectedly, glucose or drug stimulation of insulin secretion in β cells led to the preferential release of the insulin-sfGFP construct, while the mCherry-fused C-peptide remained trapped in exocytic granules. This physical separation was used to monitor glucose-stimulated insulin secretion ratiometrically by total internal reflection fluorescence microscopy in single MIN6 and primary mouse β cells. Further, RINS1 enabled parallel monitoring of pulsatile insulin release in tolbutamide-treated β cells, demonstrating the potential of RINS1 for investigations of antidiabetic drug candidates at the single-cell level. Schifferer et al. describe a genetically encoded reporter termed RINS1 for ratiometric sensing of insulin secretion. RINS1 is based on proinsulin-sfGFP and -mCherry fusions and enables monitoring of insulin dynamics upon drug treatment at different subcellular sites within β cells.

Original languageEnglish (US)
JournalCell Chemical Biology
DOIs
StateAccepted/In press - Mar 22 2016

Keywords

  • Biosensor
  • Calcium
  • Fluorescence
  • Glucose
  • Granule
  • Insulin
  • MCherry
  • Oscillation
  • Potassium channel
  • Superfolder GFP
  • TIRF
  • Tolbutamide

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry
  • Molecular Biology
  • Molecular Medicine
  • Drug Discovery
  • Pharmacology

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