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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

Protein levels of transcription factors in hindlimb skeletal muscle from WT and ShcKO mice.Skeletal muscle samples from WT and ShcKO mice, under fed and fasting conditions, were resolved by SDS-PAGE electrophoresis and western blotting performed as described in the experimental procedures. Representative immunoblots of the expression levels and normalization of the transcription factors AMPK (A), FOXO1 (B) and pAkt (C) are shown. Normalized values are expressed in arbitrary units. 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)
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pone.0124204.g006: Protein levels of transcription factors in hindlimb skeletal muscle from WT and ShcKO mice.Skeletal muscle samples from WT and ShcKO mice, under fed and fasting conditions, were resolved by SDS-PAGE electrophoresis and western blotting performed as described in the experimental procedures. Representative immunoblots of the expression levels and normalization of the transcription factors AMPK (A), FOXO1 (B) and pAkt (C) are shown. Normalized values are expressed in arbitrary units. 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)

Mentions: Samples were resolved by SDS-PAGE and transferred to membranes by western blotting. Results indicated that pAMPK levels (Fig 6A) were not different between WT and ShcKO mice under fed or fasting conditions. However, fasted WT and ShcKO mice showed higher levels of pAMPK (P < 0.05) when compared with their fed counterparts. FOXO1 levels (Fig 6B) were lower (P < 0.05) in the ShcKO when compared with WT under both fed and fasting conditions, and, under fasting conditions, both groups showed increased (P < 0.05) FOXO1 levels when compared with their fed counterparts. In the case of pAkt (Fig 6C), fasting decreased pAkt levels (P < 0.05) in both WT and ShcKO mice when compared with fed mice. In the fed state, the ShcKO mice had lower (P < 0.05) levels of pAkt than WT mice while there were no differences between genotypes in the fasted state.


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)

Protein levels of transcription factors in hindlimb skeletal muscle from WT and ShcKO mice.Skeletal muscle samples from WT and ShcKO mice, under fed and fasting conditions, were resolved by SDS-PAGE electrophoresis and western blotting performed as described in the experimental procedures. Representative immunoblots of the expression levels and normalization of the transcription factors AMPK (A), FOXO1 (B) and pAkt (C) are shown. Normalized values are expressed in arbitrary units. 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)
© Copyright Policy
Related In: Results  -  Collection

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

pone.0124204.g006: Protein levels of transcription factors in hindlimb skeletal muscle from WT and ShcKO mice.Skeletal muscle samples from WT and ShcKO mice, under fed and fasting conditions, were resolved by SDS-PAGE electrophoresis and western blotting performed as described in the experimental procedures. Representative immunoblots of the expression levels and normalization of the transcription factors AMPK (A), FOXO1 (B) and pAkt (C) are shown. Normalized values are expressed in arbitrary units. 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)
Mentions: Samples were resolved by SDS-PAGE and transferred to membranes by western blotting. Results indicated that pAMPK levels (Fig 6A) were not different between WT and ShcKO mice under fed or fasting conditions. However, fasted WT and ShcKO mice showed higher levels of pAMPK (P < 0.05) when compared with their fed counterparts. FOXO1 levels (Fig 6B) were lower (P < 0.05) in the ShcKO when compared with WT under both fed and fasting conditions, and, under fasting conditions, both groups showed increased (P < 0.05) FOXO1 levels when compared with their fed counterparts. In the case of pAkt (Fig 6C), fasting decreased pAkt levels (P < 0.05) in both WT and ShcKO mice when compared with fed mice. In the fed state, the ShcKO mice had lower (P < 0.05) levels of pAkt than WT mice while there were no differences between genotypes in the fasted state.

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