The regulation of proinsulin biosynthesis in pancreatic β-cells is vital for maintaining optimal insulin stores for glucose-induced insulin release. The majority of nutrient fuels that induce insulin release also stimulate proinsulin biosynthesis, but since insulin exocytosis and proinsulin synthesis involve different cellular mechanisms, a point of divergence in the respective metabolic stimulus–response coupling pathways must exist. A parallel examination of the metabolic regulation of proinsulin biosynthesis and insulin secretion was undertaken in the same β-cells. In MIN6 cells, glucose-induced proinsulin biosynthesis and insulin release shared a requirement for glycolysis to generate stimulus-coupling signals. Pyruvate stimulated both proinsulin synthesis (threshold 0.13–0.2 mM) and insulin release (threshold 0.2–0.3 mM) in MIN6 cells, which was eliminated by an inhibitor of pyruvate transport (1 mM α-cyano-4-hydroxycinnamate). A combination of α-oxoisohexanoate and glutamine also stimulated proinsulin biosynthesis and insulin release in MIN6 cells, which, together with the effect of pyruvate, indicated that anaplerosis was necessary for instigating secondary metabolic stimulus-coupling signals in the β-cell. A consequence of increased anaplerosis in β-cells is a marked increase in malonyl-CoA, which in turn inhibits β-oxidation and elevates cytosolic fatty acyl-CoA levels. In the β-cell, long-chain fatty acyl moieties have been strongly implicated as metabolic stimulus-coupling signals for regulating insulin exocytosis. Indeed, it was found in MIN6 cells and isolated rat pancreatic islets that exogenous oleate, palmitate and 2-bromopalmitate all markedly potentiated glucose-induced insulin release. However, in the very same β-cells, these fatty acids in contrast inhibited glucose-induced proinsulin biosynthesis. This implies that neither fatty acyl moieties nor β-oxidation are required for the metabolic stimulus–response coupling pathway specific for proinsulin biosynthesis, and represent an early point of divergence of the two signalling pathways for metabolic regulation of proinsulin biosynthesis and insulin release. Therefore alternative metabolic stimulus-coupling factors for the specific control of proinsulin biosynthesis at the translational level were considered. One possibility examined was an increase in glycerophosphate shuttle activity and change in cytosolic redox state of the β-cell, as reflected by changes in the ratio of α-glycerophosphate to dihydroxyacetone phosphate. Although 16.7 mM glucose produced a significant rise in the α-glycerophosphate/dihydroxyacetone phosphate ratio, 1 mM pyruvate did not. It follows that the cytosolic redox state and fatty acyl moieties are not necessarily involved as secondary metabolic stimulus-coupling factors for regulation of proinsulin biosynthesis. However, the results indicate that glycolysis and the subsequent increase in anaplerosis are indeed necessary for this signalling pathway, and therefore an extramitochondrial product of β-cell pyruvate metabolism (that is upstream of the increased cytosolic fatty acyl-CoA) acts as a key intracellular secondary signal for specific control of proinsulin biosynthesis by glucose at the level of translation.

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