Aims/hypothesis:  

Syntaxin-1A (Syn-1A) is known to play a negative regulatory role in insulin secretion but the precise mechanisms for its action are not clear. Syn-2, –3 and –4 are also present in islet beta cells but their functions are not known. Here, we investigated the role of these syntaxins in the insulin secretory process.

Methods:  

We examined the following effects of Syn-1, –2, –3 and –4 expression in insulinoma beta-cell lines. Endogenous insulin secretion was measured by batch radioimmunoassay (RIA) and single cell patch clamp capacitance measurements. The l-type Ca²⁺ channel activity was studied by patch clamp electrophysiology. Insulin gene transcription was examined by Northern blotting and measurement of insulin gene promoter activity by the co-expression of cyan fluorescent protein-labelled rat insulin promoter.

Results:  

Syn-1A or –3, but not Syn-2 or –4 overexpression, inhibited K⁺-induced insulin release as determined by RIA (49.7 ± 5.5 % and 49.1 ± 6.2 %, respectively) and electrophysiologic membrane capacitance measurements (68.0 ± 21.0 % and 58.0 ± 13.2 %, respectively). Overexpressed Syn-1A and –3, but not Syn-2, inhibited Ca²⁺ channel current amplitude by 39.5 ± 11.6 % and 52.7 ± 6.0 %, respectively. Of note, overexpression of Syn-1A and –3 also reduced single cell (by confocal microscopy) and total cellular endogenous insulin content (by RIA) by 24.8 ± 4.2 % and 31.8 ± 3.9 %, respectively. This correlated to a reduction in endogenous insulin mRNA by 24.5 ± 4.2 % and 25.7 ± 4.2 %, respectively. This inhibition of insulin biosynthesis is mainly at the level of insulin gene transcription as demonstrated by an inhibition of insulin gene promoter activity (53.3 ± 9.15 % and 39.0 ± 6.8 %, respectively).

Conclusions/interpretation:  

These results demonstrate that Syn-1A and –3 possess strong inhibitory actions on both insulin exocytosis and insulin biosynthesis whereas Syn-2 and –4 do not inhibit the insulin secretory process. [Diabetologia (2002) 45: 231–241]