Differences in signaling properties of the cytoplasmic domains of the insulin receptor and insulin-like growth factor receptor in 3T3-L1 adipocytes.

Insulin and insulin-like growth factors (IGFs) elicit distinct but overlapping biological effects in vivo. To investigate whether differences in intrinsic signaling capacity of receptors contribute to biological specificity, we constructed chimeric receptors containing the extracellular portion of the neurotrophin receptor TrkC fused to the intracellular portion of the insulin or IGF-I receptors. Chimeras were stably expressed in 3T3-L1 adipocytes at levels comparable to endogenous insulin receptors and were efficiently activated by neurotrophin-3. The wild-type insulin receptor chimera mediated approximately 2-fold greater phosphorylation of insulin receptor substrate 1 (IRS-1), association of IRS-1 with phosphoinositide 3-kinase, stimulation of glucose uptake, and GLUT4 translocation, compared with the IGF-I receptor chimera. In contrast, the IGF-I receptor chimera mediated more effective Shc phosphorylation, association of Shc with Grb2, and activation of mitogen-activated protein kinase compared with the insulin receptor chimera. The two receptors elicited similar activation of protein kinase B, p70S6 kinase, and glycogen synthesis. We conclude that the insulin receptor mediates some aspects of metabolic signaling in adipocytes more effectively than the IGF-I receptor, as a consequence of more efficient phosphorylation of IRS-1 and greater recruitment/activation of phosphoinositide 3-kinase.

Differential signaling to glycogen synthesis by the intracellular domain of the insulin versus the insulin-like growth factor-1 receptor. Evidence from studies of TrkC-chimeras.

Insulin and insulin-like growth factor-1 (IGF-1) have similar cell-surface receptors yet subserve different physiological functions. To examine whether these differences relate to intrinsic signaling properties of the intracellular domains of their respective receptors, chimeric receptors were constructed using the extracellular domain of the neurotrophin-3 (NT-3) receptor, TrkC, and the intracellular domain of either the insulin receptor or the IGF-1 receptor. TrkC-IR (TIR) and TrkC-IGF-1R (TIGR) were stably expressed in 3T3-L1 cells. While TIR and TIGR cell lines expressing similar numbers of chimeric receptors showed a similar dose-response relationship in NT-3 stimulated DNA synthesis, NT-3 stimulated glycogen synthesis was greater in TIR than in TIGR cells (maximum TIGR response was 35% of maximum TIR response). Additionally, the concentration of NT-3 at which significant stimulation of glycogen synthesis was seen was 0.1 ng/ml in TIR and 1 ng/ml in TIGR cells. Basal levels of thymidine incorporation but not glycogen synthesis were consistently higher in TIR than in TIGR expressing cells. No detectable basal autophosphorylation of chimeric receptors was seen in any of the cell lines. However, exposure of cell lines to the phosphatase inhibitor bisperoxovanadate resulted in greater basal autophosphorylation of the TIR and endogenous murine IR than the TIGR and endogenous murine IGF-1R. Thus, in this system, the intracellular domain of the IR appears to couple more effectively to glycogen synthesis than that of the IGF-1R, whereas the intracellular domains of both receptors have a similar capacity to stimulate DNA synthesis.

Homozygosity for a common polymorphism in the islet-specific promoter of the glucokinase gene is associated with a reduced early insulin response to oral glucose in pregnant women.

A commonly occurring sequence variant in the islet-specific promoter of the glucokinase gene (-30 G to A) has been variably reported to be associated with reduced insulin secretory responses to oral glucose. The effect of this promoter variant may be subtle and only become apparent under conditions of beta-cell 'stress'. As late pregnancy is a time of increased insulin secretory demand, we have examined whether this common genetic variant was associated with impairment of insulin secretory responses to oral glucose in 92 women in the third trimester of pregnancy. The three women who were homozygous for the variant sequence had a markedly diminished 30' insulin incremental response to oral glucose (10.4, 11.4, and 17.2 pmol insulin mmol-1 glucose, respectively) compared to either heterozygous (49.3 (37.6-64.6 pmol insulin mmol-1 glucose)) (p < 0.002) or homozygous wild-type (51.4 (40.9-64.7 pmol insulin mmol-1 glucose)) (p < 0.002) Mann-Whitney U test) women. In a subset of 35 British Caucasian women with gestational diabetes, no mutations resulting in a change of amino acid sequence were detected by molecular scanning of all exons of the glucokinase gene. In summary, in a cohort of 35 British Caucasian women with gestational diabetes neither missense nor nonsense glucokinase mutations were found. However, in women in the third trimester of pregnancy, homozygosity for a common polymorphic variant in the islet-specific promoter of the glucokinase gene was associated with a highly significant reduction of early insulin secretory responsiveness to oral glucose. Under the conditions of increased secretory demand represented by late pregnancy, a promoter variant in the glucokinase gene may influence the early insulin secretory response to oral glucose.