Functional characterization of an insulin-responsive glucose transporter (GLUT4) from fish adipose tissue

Self Cite

SUMMARY In brown trout, red and white skeletal muscle express the insulin-regulatable glucose transporter 4 (btGLUT4). We have previously shown that the mRNA expression of btGLUT4 in red muscle, but not white muscle, is altered under experimental conditions designed to cause changes in the plasma levels of insulin, such as fasting, insulin and arginine administration. In order to determine whether changes of btGLUT4 expression at the mRNA level are correlated with changes at the protein level, we performed in vivoexperiments to alter blood insulin concentrations and determined the abundance of btGLUT4 protein in trout red and white skeletal muscle by immunoblotting using an antibody to salmon GLUT4. In the present study we show that btGLUT4 protein content in red muscle decreases after fasting and increases after insulin administration. By contrast, btGLUT4 protein content in white muscle decreases after fasting but is not affected by insulin treatment. Our results show a good correlation between the changes observed in btGLUT4 protein and the previously reported changes in mRNA levels in response to alterations in circulating insulin, indicating that the regulation of btGLUT4 in brown trout takes place predominantly in the red skeletal muscle.

Section: Introductionmentioning

confidence: 87%

Physiological regulation of glucose transporter (GLUT4) protein content in brown trout (Salmo trutta) skeletal muscle

Dı́az

Capilla

Planas

2007

Self Cite

“…In mammalian cells, the FQQI domain is important in the sorting process that separates GLUT4 from the endosome system (Holman and Sandoval, 2001;Lalioti et al, 2001). In Atlantic cod and coho salmon (Capilla et al, 2004) the FQQ motif is present, but the I is replaced with another hydrophobic amino acid, L. In brown trout, the motif is FQHL (Capilla et al, 2004).…”

Section: Amino Acid Sequencesmentioning

confidence: 99%

“…In brown trout GLUT4 was expressed primarily in red and white skeletal muscle, gill, kidney and adipose tissue but only to a limited extent in heart (Planas et al, 2000). Also, GLUT4 is insulin sensitive in adipocytes from coho salmon (Capilla et al, 2004). In salmonids, red muscle shows the hallmarks of insulin responsiveness but white muscle does not.…”

Section: Introductionmentioning

confidence: 99%

Sequence of Atlantic cod (Gadus morhua) GLUT4, GLUT2 and GPDH:developmental stage expression, tissue expression and relationship to starvation-induced changes in blood glucose

Hall

Short

Driedzic

2006

SUMMARY cDNAs of putative glucose transporters, GLUT4 and GLUT2, were cloned from Atlantic cod (Gadus morhua). The GLUT4 cDNA encodes a 503 amino acid and the GLUT2 cDNA a 506 amino acid protein. Phylogenetic analysis, amino acid sequence alignment, and tissue distribution support categorizing them as homologues of mammalian GLUT4 and 2. GLUT4 clusters with GLUT4s from fish and other vertebrates. It shows 84% amino acid identity to GLUT4 from coho salmon and brown trout and 65% identity with other vertebrates. It is most highly expressed in heart, strongly expressed in red and white skeletal muscle and present at lower levels in gill, gonad, intestine, and kidney. GLUT2 clusters with GLUT2 from rainbow trout and other vertebrates. It shows 75% amino acid identity with rainbow trout and 62% identity with chicken GLUT2. In Atlantic cod, GLUT2 is most highly expressed in liver with lower levels noted in intestine and kidney. Food deprivation for 2 months was used as a vehicle to monitor GLUT expression at different blood glucose levels. Starvation resulted in a decrease in blood glucose and liver glycogen that recovered following 20 days of re-feeding. GLUT4 expression in heart was decreased with starvation and increased with re-feeding. GLUT4 mRNA level in heart correlated with blood glucose. It is suggested that this relationship is related to insulin responsiveness. GLUT4 expression in white muscle increased with starvation and decreased with re-feeding. It is proposed that this is due to the necessity to maintain high levels of the glucose transporter protein in the face of starvation-associated proteolysis. GLUT2 expression in liver correlated with blood glucose, consistent with higher rates of glucose transport from liver to blood in the fed state than in the food-deprived state. Glycerol-3-phosphate dehydrogenase (GPDH) cDNA was also cloned. It encodes a 351 amino acid protein, which is 73-90% identical to GPDH from numerous other fish species. GPDH is ubiquitously expressed. Expression in heart decreased with starvation and increased with refeeding, whereas expression in liver did not change with starvation. In other studies, gene expression was monitored at nine time points from fertilization of eggs to larval development. GLUT4 is detectable in fertilized eggs and is fully expressed by the halfway to hatching point. GLUT2 is not evident at fertilization, is detectable at halfway to hatching, and fully expressed at hatching. GPDH expression was evident from fertilization.

“…Fish glucose transporters GLUT2 and GLUT4 are characterized by a lower affinity for glucose, which may at least partly explain the persistent post-prandial hyperglycemia observed in trout fed with carbohydrates (Capilla et al, 2004a;Krasnov et al, 2001). At the metabolic level, most of the key enzymes involved in carbohydrate metabolism have been described in fish (Cowey and Walton, 1989).…”

Section: Introductionmentioning

confidence: 99%

Insulin regulates the expression of several metabolism-related genes in the liver and primary hepatocytes of rainbow trout (Oncorhynchus mykiss)

Plagnes-Juan

Lansard

Seiliez

et al. 2008

108

SUMMARYRainbow trout have a limited ability to use dietary carbohydrates efficiently and are considered to be glucose intolerant. Administration of carbohydrates results in persistent hyperglycemia and impairs post-prandial down regulation of gluconeogenesis despite normal insulin secretion. Since gluconeogenic genes are mainly under insulin control, we put forward the hypothesis that the transcriptional function of insulin as a whole may be impaired in the trout liver. In order to test this hypothesis, we performed intraperitoneal administration of bovine insulin to fasted rainbow trout and also subjected rainbow trout primary hepatocytes to insulin and/or glucose stimulation. We demonstrate that insulin was able to activate Akt, a key element in the insulin signaling pathway, and to regulate hepatic metabolism-related target genes both in vivo and in vitro. In the same way as in mammals, insulin decreased mRNA expression of gluconeogenic genes, including glucose 6-phosphatase (G6Pase), fructose 1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK). Insulin also limited the expression of carnitine palmitoyltransferase 1 (CPT1), a limiting enzyme of fatty acid β-oxidation. In vitro studies revealed that, as in mammals, glucose is an important regulator of some insulin target genes such as the glycolytic enzyme pyruvate kinase (PK) and the lipogenic enzyme fatty acid synthase (FAS). Interestingly, glucose also stimulates expression of glucokinase (GK), which has no equivalent in mammals. This study demonstrates that insulin possesses the intrinsic ability to regulate hepatic gene expression in rainbow trout, suggesting that other hormonal or metabolic factors may counteract some of the post-prandial actions of insulin.