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Key glycolytic enzyme activities of skeletal muscle are decreased under fed and fasted states in mice with knocked down levels of Shc proteins.

Hagopian K, Tomilov AA, Kim K, Cortopassi GA, Ramsey JJ - PLoS ONE (2015)

Bottom Line: Shc proteins interact with the insulin receptor, indicating a role in regulating glycolysis.Changes in metabolite levels were consistent with the observed changes in enzyme activities.These studies indicate that decreased levels of Shc proteins in skeletal muscle lead to a decreased glycolytic capacity in both fed and fasted states.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California Davis, Davis, CA 95616, United States of America.

ABSTRACT
Shc proteins interact with the insulin receptor, indicating a role in regulating glycolysis. To investigate this idea, the activities of key glycolytic regulatory enzymes and metabolites levels were measured in skeletal muscle from mice with low levels of Shc proteins (ShcKO) and wild-type (WT) controls. The activities of hexokinase, phosphofructokinase-1 and pyruvate kinase were decreased in ShcKO versus WT mice under both fed and fasted conditions. Increased alanine transaminase and branched-chain amino acid transaminase activities were also observed in ShcKO mice under both fed and fasting conditions. Protein expression of glycolytic enzymes was unchanged in the ShcKO and WT mice, indicating that decreased activities were not due to changes in their transcription. Changes in metabolite levels were consistent with the observed changes in enzyme activities. In particular, the levels of fructose-2,6-bisphosphate, a potent activator of phosphofructokinase-1, were consistently decreased in the ShcKO mice. Furthermore, the levels of lactate (inhibitor of hexokinase and phosphofructokinase-1) and citrate (inhibitor of phosphofructokinase-1 and pyruvate kinase) were increased in fed and fasted ShcKO versus WT mice. Pyruvate dehydrogenase activity was lower in ShcKO versus WT mice under fed conditions, and showed inhibition under fasting conditions in both ShcKO and WT mice, with ShcKO mice showing less inhibition than the WT mice. Pyruvate dehydrogenase kinase 4 levels were unchanged under fed conditions but were lower in the ShcKO mice under fasting conditions. These studies indicate that decreased levels of Shc proteins in skeletal muscle lead to a decreased glycolytic capacity in both fed and fasted states.

No MeSH data available.


Related in: MedlinePlus

Glycolytic and other metabolite levels in hindlimb skeletal muscle from WT and ShcKO mice.The levels of several metabolites from fed and fasted ShcKO and WT mice were determined under fed and fasting conditions, as described in the experimental procedures. The glycolytic metabolites determined were G6P (A), F6P (B), F1,6BP (C), the G6P/F1,6BP ratio (D), PYR (E), LAC (F) and LAC/PYR ratio (G). Also determined were the levels of F2,6BP (H), Acetyl-CoA (I), CoA (J), Acetyl-CoA/CoA ratio (K) and citrate (L). The following comparisons were made: within a genotype, fed versus fasted; across genotypes, fed versus fed and fasted versus fasted. Bars that do not share a common symbol differ significantly (P < 0.05). Data presented as mean ± SEM (n = 6). The symbol (~) is used to indicate a trend towards an increase or a decrease (P < 0.10) when the difference is not significant.
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pone.0124204.g004: Glycolytic and other metabolite levels in hindlimb skeletal muscle from WT and ShcKO mice.The levels of several metabolites from fed and fasted ShcKO and WT mice were determined under fed and fasting conditions, as described in the experimental procedures. The glycolytic metabolites determined were G6P (A), F6P (B), F1,6BP (C), the G6P/F1,6BP ratio (D), PYR (E), LAC (F) and LAC/PYR ratio (G). Also determined were the levels of F2,6BP (H), Acetyl-CoA (I), CoA (J), Acetyl-CoA/CoA ratio (K) and citrate (L). The following comparisons were made: within a genotype, fed versus fasted; across genotypes, fed versus fed and fasted versus fasted. Bars that do not share a common symbol differ significantly (P < 0.05). Data presented as mean ± SEM (n = 6). The symbol (~) is used to indicate a trend towards an increase or a decrease (P < 0.10) when the difference is not significant.

Mentions: Several metabolites of the glycolytic pathway from muscle were measured (Fig 4). In the ShcKO mice, G6P, F6P and F1,6BP (Fig 4A–4C) were lower (P < 0.05) than the WT animals under both fed and fasting conditions. However, under fasting conditions, G6P and F6P levels in both WT and ShcKO mice were higher (P < 0.05) while F1,6BP was lower (P < 0.05) when compared with fed conditions. Taking the ratio of G6P/F1,6BP as indicator of PFK-1 inhibition (Fig 4D), the results showed increased ratios (P < 0.05) in both WT and ShcKO mice under fasting compared with fed conditions. However, under fasting conditions, the ratio was higher (P < 0.05) in the ShcKO mice compared to WT mice while under fed conditions a trend toward an increase (P = 0.062) was observed in the ShcKO compared to WT mice. The levels of PYR and LAC were also measured. Under fed conditions, ShcKO mice showed lower (P < 0.05) PYR levels compared to WT animals (Fig 4E), while under fasting conditions the opposite was the case (P < 0.05). When fasted WT and ShcKO mice were compared with their fed counterparts, PYR levels were lower (P < 0.05) in the WT animals and did not differ in the ShcKO mice. In the case of LAC (Fig 4F), under both fed and fasted conditions, the ShcKO mice had higher (P < 0.05) levels of LAC than their WT counterparts. When fed WT and ShcKO mice were compared with their fasted counterparts, the fasted animals had higher (P < 0.05) levels of LAC. The LAC/PYR ratio (Fig 4G), as indicator of the NADH/NAD ratio, was higher (P < 0.05) in the fasted compared to fed WT mice while the fasted ShcKO did not differ from their fed counterparts. Under fed conditions, the ratio was higher (P < 0.05) in the ShcKO compared to WT mice while under fasting there was a trend (P = 0.09) toward a decrease in the ShcKO versus WT animals.


Key glycolytic enzyme activities of skeletal muscle are decreased under fed and fasted states in mice with knocked down levels of Shc proteins.

Hagopian K, Tomilov AA, Kim K, Cortopassi GA, Ramsey JJ - PLoS ONE (2015)

Glycolytic and other metabolite levels in hindlimb skeletal muscle from WT and ShcKO mice.The levels of several metabolites from fed and fasted ShcKO and WT mice were determined under fed and fasting conditions, as described in the experimental procedures. The glycolytic metabolites determined were G6P (A), F6P (B), F1,6BP (C), the G6P/F1,6BP ratio (D), PYR (E), LAC (F) and LAC/PYR ratio (G). Also determined were the levels of F2,6BP (H), Acetyl-CoA (I), CoA (J), Acetyl-CoA/CoA ratio (K) and citrate (L). The following comparisons were made: within a genotype, fed versus fasted; across genotypes, fed versus fed and fasted versus fasted. Bars that do not share a common symbol differ significantly (P < 0.05). Data presented as mean ± SEM (n = 6). The symbol (~) is used to indicate a trend towards an increase or a decrease (P < 0.10) when the difference is not significant.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400099&req=5

pone.0124204.g004: Glycolytic and other metabolite levels in hindlimb skeletal muscle from WT and ShcKO mice.The levels of several metabolites from fed and fasted ShcKO and WT mice were determined under fed and fasting conditions, as described in the experimental procedures. The glycolytic metabolites determined were G6P (A), F6P (B), F1,6BP (C), the G6P/F1,6BP ratio (D), PYR (E), LAC (F) and LAC/PYR ratio (G). Also determined were the levels of F2,6BP (H), Acetyl-CoA (I), CoA (J), Acetyl-CoA/CoA ratio (K) and citrate (L). The following comparisons were made: within a genotype, fed versus fasted; across genotypes, fed versus fed and fasted versus fasted. Bars that do not share a common symbol differ significantly (P < 0.05). Data presented as mean ± SEM (n = 6). The symbol (~) is used to indicate a trend towards an increase or a decrease (P < 0.10) when the difference is not significant.
Mentions: Several metabolites of the glycolytic pathway from muscle were measured (Fig 4). In the ShcKO mice, G6P, F6P and F1,6BP (Fig 4A–4C) were lower (P < 0.05) than the WT animals under both fed and fasting conditions. However, under fasting conditions, G6P and F6P levels in both WT and ShcKO mice were higher (P < 0.05) while F1,6BP was lower (P < 0.05) when compared with fed conditions. Taking the ratio of G6P/F1,6BP as indicator of PFK-1 inhibition (Fig 4D), the results showed increased ratios (P < 0.05) in both WT and ShcKO mice under fasting compared with fed conditions. However, under fasting conditions, the ratio was higher (P < 0.05) in the ShcKO mice compared to WT mice while under fed conditions a trend toward an increase (P = 0.062) was observed in the ShcKO compared to WT mice. The levels of PYR and LAC were also measured. Under fed conditions, ShcKO mice showed lower (P < 0.05) PYR levels compared to WT animals (Fig 4E), while under fasting conditions the opposite was the case (P < 0.05). When fasted WT and ShcKO mice were compared with their fed counterparts, PYR levels were lower (P < 0.05) in the WT animals and did not differ in the ShcKO mice. In the case of LAC (Fig 4F), under both fed and fasted conditions, the ShcKO mice had higher (P < 0.05) levels of LAC than their WT counterparts. When fed WT and ShcKO mice were compared with their fasted counterparts, the fasted animals had higher (P < 0.05) levels of LAC. The LAC/PYR ratio (Fig 4G), as indicator of the NADH/NAD ratio, was higher (P < 0.05) in the fasted compared to fed WT mice while the fasted ShcKO did not differ from their fed counterparts. Under fed conditions, the ratio was higher (P < 0.05) in the ShcKO compared to WT mice while under fasting there was a trend (P = 0.09) toward a decrease in the ShcKO versus WT animals.

Bottom Line: Shc proteins interact with the insulin receptor, indicating a role in regulating glycolysis.Changes in metabolite levels were consistent with the observed changes in enzyme activities.These studies indicate that decreased levels of Shc proteins in skeletal muscle lead to a decreased glycolytic capacity in both fed and fasted states.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California Davis, Davis, CA 95616, United States of America.

ABSTRACT
Shc proteins interact with the insulin receptor, indicating a role in regulating glycolysis. To investigate this idea, the activities of key glycolytic regulatory enzymes and metabolites levels were measured in skeletal muscle from mice with low levels of Shc proteins (ShcKO) and wild-type (WT) controls. The activities of hexokinase, phosphofructokinase-1 and pyruvate kinase were decreased in ShcKO versus WT mice under both fed and fasted conditions. Increased alanine transaminase and branched-chain amino acid transaminase activities were also observed in ShcKO mice under both fed and fasting conditions. Protein expression of glycolytic enzymes was unchanged in the ShcKO and WT mice, indicating that decreased activities were not due to changes in their transcription. Changes in metabolite levels were consistent with the observed changes in enzyme activities. In particular, the levels of fructose-2,6-bisphosphate, a potent activator of phosphofructokinase-1, were consistently decreased in the ShcKO mice. Furthermore, the levels of lactate (inhibitor of hexokinase and phosphofructokinase-1) and citrate (inhibitor of phosphofructokinase-1 and pyruvate kinase) were increased in fed and fasted ShcKO versus WT mice. Pyruvate dehydrogenase activity was lower in ShcKO versus WT mice under fed conditions, and showed inhibition under fasting conditions in both ShcKO and WT mice, with ShcKO mice showing less inhibition than the WT mice. Pyruvate dehydrogenase kinase 4 levels were unchanged under fed conditions but were lower in the ShcKO mice under fasting conditions. These studies indicate that decreased levels of Shc proteins in skeletal muscle lead to a decreased glycolytic capacity in both fed and fasted states.

No MeSH data available.


Related in: MedlinePlus