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Scientific article
English

Conversion of proinsulin to insulin occurs coordinately with acidification of maturing secretory vesicles

Published inThe Journal of cell biology, vol. 103, no. 6 Pt 1, p. 2273-2281
Publication date1986
Abstract

Proinsulin is a single polypeptide chain composed of the B and A subunits of insulin joined by the C-peptide region. Proinsulin is converted to insulin during the maturation of secretory vesicles by the action of two proteases and conversion is inhibited by ionophores that disrupted intracellular H+ gradients. To determine if conversion of prohormone to hormone actually occurs in an acidic secretory vesicle, cultured rat islet cells were incubated in the presence of 3-(2,4-dinitroanilino)-3' amino-N-methyldipropylamine (DAMP), a basic congener of dinitrophenol that concentrates in acidic compartments and is retained there after aldehyde fixation. The cells were processed for indirect protein A-gold colocalization of DAMP, using a monoclonal antibody to dinitrophenol, and proinsulin, using a monoclonal antibody that exclusively reacts with the prohormone. The average density of DAMP-specific gold particles in immature secretory vesicles that contained proinsulin was 71/micron 2 (18 times cytoplasmic background), which indicated that this compartment was acidic. However, the density of DAMP-specific gold particles in the insulin-rich mature secretory vesicle averaged 433/micron 2. This suggests that although proinsulin conversion occurs in an acidic compartment, the secretory vesicles become more acidic as they mature. Since the concentration of anti-proinsulin IgG binding in secretory vesicles is inversely proportional to the conversion of proinsulin to insulin, we were able to determine that maturing secretory vesicles had to reach a critical pH before proinsulin conversion occurred.

Keywords
  • Animals
  • Antibodies, Monoclonal
  • Cytoplasmic Granules/ metabolism/ultrastructure
  • Fluorescent Antibody Technique
  • Insulin/analysis/ genetics
  • Islets of Langerhans/ metabolism/ultrastructure
  • Microscopy, Electron
  • Peptide Hydrolases/ metabolism
  • Proinsulin/analysis/ genetics
  • Protein Processing, Post-Translational
  • Rats
  • Rats, Inbred Strains
Citation (ISO format)
ORCI, Lelio et al. Conversion of proinsulin to insulin occurs coordinately with acidification of maturing secretory vesicles. In: The Journal of cell biology, 1986, vol. 103, n° 6 Pt 1, p. 2273–2281. doi: 10.1083/jcb.103.6.2273
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Identifiers
ISSN of the journal0021-9525
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