JCS02658 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. Adenovirus-mediated overexpression of SV2A and SV2C. (A) INS-1E cells were infected with the control AdCALacZ (LacZ), AdCASV2A (SV2A) or AdCASV2C (SV2C) adenoviruses at ~50 and 100 virus particles/cell. The next day cellular homogenates were collected and the expression of the exogenous proteins were analysed by western blotting using anti-SV2A or anti-SV2C antibodies. (B) INS-1E cells were infected with AdCALacZ (a,b), AdCASV2A (c) or AdCASV2C (d) at 50 virus particles/cell. One day later, the cells were double stained with antibodies directed against insulin (green) and against SV2A or SV2C (red). The right panels correspond to the overlays of the anti-insulin and anti-SV2A or -SV2C signals.

• Supplemental Figure 2 -

  Fig. S2. Impact of overexpressed SV2A and SV2C on insulin release in islets. (Top panel) Immunoblotting of SV2A and -C overexpression in rat islets performed with the anti-SV2 antibody. Additional immunofluorescence studies revealed that exogenous SV2 proteins are correctly targeted to insulin granules (not shown). (Lower panels) Islets infected with adenoviruses containing the control LacZ, SV2A or SV2C cDNAs, were incubated at basal 2.8 mM glucose (glc) and stimulated for 15 minutes with 30 mM KCl or for 30 minutes with 16.7 mM glucose. Values show the meanąs.e. of four independent experiments (*P<0.05). The more marked secretory defect in pancreatic islets compared to INS 1E cells could be explained by the approximately fivefold higher endogenous protein level in the cell line diminishing the impact of the extrinsic SV2 isoforms.