4 research outputs found
Osmotic swelling-induced activation of the extracellular-signal-regulated protein kinases Erk-1 and Erk-2 in intestine 407 cells involves the Ras/Raf-signalling pathway
Human Intestine 407 cells respond to hypo-osmotic stress with a rapid
stimulation of compensatory ionic conductances accompanied by a transient
increase in the activity of the extracellular-signal-regulated protein
kinases Erk-1 and Erk-2. In this study, we examined the upstream
regulators of hypotonicity-induced Erk-1/Erk-2 activation and their
possible role in cell-volume regulation. The hypotonicity-provoked
Erk-1/Erk-2 activation was greatly reduced in cells pretreated with the
specific mitogen-activated/Erk-activating kinase inhibitor PD098059 and
was preceded by a transient stimulation of Raf-1. Pretreatment of the
cells with PMA, GF109203X, wortmannin or Clostridium botulinum C3
exoenzyme did not appreciably affect the hypotonicity-provoked Erk-1/Erk-2
stimulation, suggesting the osmosensitive signalling pathway to be largely
independent of protein kinase C and p21(rho). In contrast, expression of
dominant negative RasN17 completely abolished the hypotonicity-induced
Erk-1/Erk-2 activation. Stimulation of the swelling-induced ion efflux was
independent of activation of these mitogen-activated protein kinases, as
revealed by hypotonicity-provoked isotope efflux from 125I-- and
86Rb+-loaded cells after pretreatment with PD098059 and after
An insulin receptor mutant (Asp707 → Ala), involved in leprechaunism, is processed and transported to the cell surface but unable to bind insulin
We have identified a homozygous mutation near the carboxyl terminus of the insulin receptor (IR) α subunit from a leprechaun patient, changing Asp707 into Ala. Fibroblasts from this patient had no high affinity insulin binding sites. To examine the effect of the mutation on IR properties, the mutant IR was stably expressed in Chinese hamster ovary cells. Western blot analysis and metabolic labeling showed a normal processing of the mutant receptor to α and β subunits. No increase in high affinity insulin binding sites was observed on Chinese hamster ovary cells expressing the mutant receptor, and also, affinity cross-linking of 125I- labeled insulin by disuccinimidyl suberate to these cells failed to label the mutant α subunit. Biotinylation of cell surface proteins by biotin succinimidyl ester resulted in efficient biotinylation of the mutant IR α and β subunits, showing its presence on the cell surface. On solubilization of the mutant insulin receptor in Triton X. 100-containing buffers, 125I- insulin was efficiently cross-linked to the receptor a subunit by disuccinimidyl suberate. These studies demonstrate that Ala707 IR is normally processed and transported to the cell surface and that the mutation distorts the insulin binding site. Detergent restores this site. This is an example of a naturally occurring mutation in the insulin receptor that affects insulin binding without affecting receptor transport and processing. This mutation points to a major contribution of the a subunit carboxyl terminus to insulin binding
Genetic factors and insulin secretion: gene variants in the IGF genes
IGFs are important regulators of pancreatic beta-cell development, growth, and maintenance. Mutations in the IGF genes have been found to be associated with type 2 diabetes, myocardial infarction, birth weight, and obesity. These associations could result from changes in insulin secretion. We have analyzed glucose-stimulated insulin secretion using hyperglycemic clamps in carriers of a CA repeat in the IGF-I promoter and an ApaI polymorphism in the IGF-II gene. Normal and impaired glucose-tolerant subjects (n = 237) were independently recruited from three different populations in the Netherlands and Germany to allow independent replication of associations. Both first- and second-phase insulin secretion were not significantly different between the various IGF-I or IGF-II genotypes. Remarkably, noncarriers of the IGF-I CA repeat allele had both a reduced insulin sensitivity index (ISI) and disposition index (DI), suggesting an altered balance between insulin secretion and insulin action. Other diabetes-related parameters were not significantly different for both the IGF-I and IGF-II gene variant. We conclude that gene variants in the IGF-I and IGF-II genes are not associated with detectable variations in glucose-stimulated insulin secretion in these three independent populations. Further studies are needed to examine the exact contributions of the IGF-I CA repeat alleles to variations in ISI and DI
Evidence that the mitochondrial leucyl tRNA synthetase (LARS2) gene represents a novel type 2 diabetes susceptibility gene
Previously, we have shown that a mutation in the mitochondrial DNA-encoded tRNA(Leu(UUR)) gene is associated with type 2 diabetes. One of the consequences of this mutation is a reduced aminoacylation of tRNA(Leu(UUR)). In this study, we have examined whether variants in the leucyl tRNA synthetase gene (LARS2), involved in aminoacylation of tRNA(Leu(UUR)), associate with type 2 diabetes. Direct sequencing of LARS2 cDNA from 25 type 2 diabetic subjects revealed eight single nucleotide polymorphisms. Two of the variants were examined in 7,836 subjects from four independent populations in the Netherlands and Denmark. A -109 g/a variant was not associated with type 2 diabetes. Allele frequencies for the other variant, H324Q, were 3.5% in type 2 diabetic and 2.7% in control subjects, respectively. The common odds ratio across all four studies was 1.40 (95% CI 1.12-1.76), P = 0.004. There were no significant differences in clinical variables between carriers and noncarriers. In this study, we provide evidence that the LARS2 gene may represent a novel type 2 diabetes susceptibility gene. The mechanism by which the H324Q variant enhances type 2 diabetes risk needs to be further established. This is the first report of association between an aminoacyl tRNA synthetase gene and disease. Our results further highlight the important role of mitochondria in glucose homeostasis