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Mice deficient in ribosomal protein S6 phosphorylation suffer from muscle weakness that reflects a growth defect and energy deficit.

Ruvinsky I, Katz M, Dreazen A, Gielchinsky Y, Saada A, Freedman N, Mishani E, Zimmerman G, Kasir J, Meyuhas O - PLoS ONE (2009)

Bottom Line: Collectively, these experiments have demonstrated that the physical inferiority appears to result from two defects: a) a decrease in total muscle mass that reflects impaired growth, rather than aberrant differentiation of myofibers, as well as a diminished abundance of contractile proteins; and b) a reduced content of ATP and phosphocreatine, two readily available energy sources.However, the apparent energy deficiency in this genotype does not result from a lower mitochondrial mass or compromised activity of enzymes of the oxidative phosphorylation, nor does it reflect a decline in insulin-dependent glucose uptake, or diminution in storage of glycogen or triacylglycerol (TG) in the muscle.Interestingly, a similar role has been assigned for ribosomal protein S6 kinase 1, even though it regulates myoblast growth in an rpS6 phosphorylation-independent fashion.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, IMRIC, Hebrew University-Hadassah Medical School, Jerusalem, Israel.

ABSTRACT

Background: Mice, whose ribosomal protein S6 cannot be phosphorylated due to replacement of all five phosphorylatable serine residues by alanines (rpS6(P-/-)), are viable and fertile. However, phenotypic characterization of these mice and embryo fibroblasts derived from them, has established the role of these modifications in the regulation of the size of several cell types, as well as pancreatic beta-cell function and glucose homeostasis. A relatively passive behavior of these mice has raised the possibility that they suffer from muscle weakness, which has, indeed, been confirmed by a variety of physical performance tests.

Methodology/principal findings: A large variety of experimental methodologies, including morphometric measurements of histological preparations, high throughput proteomic analysis, positron emission tomography (PET) and numerous biochemical assays, were used in an attempt to establish the mechanism underlying the relative weakness of rpS6(P-/-) muscles. Collectively, these experiments have demonstrated that the physical inferiority appears to result from two defects: a) a decrease in total muscle mass that reflects impaired growth, rather than aberrant differentiation of myofibers, as well as a diminished abundance of contractile proteins; and b) a reduced content of ATP and phosphocreatine, two readily available energy sources. The abundance of three mitochondrial proteins has been shown to diminish in the knockin mouse. However, the apparent energy deficiency in this genotype does not result from a lower mitochondrial mass or compromised activity of enzymes of the oxidative phosphorylation, nor does it reflect a decline in insulin-dependent glucose uptake, or diminution in storage of glycogen or triacylglycerol (TG) in the muscle.

Conclusions/significance: This study establishes rpS6 phosphorylation as a determinant of muscle strength through its role in regulation of myofiber growth and energy content. Interestingly, a similar role has been assigned for ribosomal protein S6 kinase 1, even though it regulates myoblast growth in an rpS6 phosphorylation-independent fashion.

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Related in: MedlinePlus

The amount of mitochondria and the activity of complexes of the oxidative phosphorylation system are unchanged in rpS6P−/− muscle.Mitochondria were isolated from hind limb muscle of four age-matched rpS6P+/+ (WT) and rpS6P−/− (−/−) male mice and were assayed for citrate synthase (A), cytochrome C oxidase (B), as well as complexes I (C), II (D), II+III (E) and IV (F) of the oxidative phosphorylation system. The activity of the different complexes was normalized to that of citrate synthase and the results are presented as relative activities. Vertical bars represent SEM (n = 4 age- matched male mice for each genotype).
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pone-0005618-g005: The amount of mitochondria and the activity of complexes of the oxidative phosphorylation system are unchanged in rpS6P−/− muscle.Mitochondria were isolated from hind limb muscle of four age-matched rpS6P+/+ (WT) and rpS6P−/− (−/−) male mice and were assayed for citrate synthase (A), cytochrome C oxidase (B), as well as complexes I (C), II (D), II+III (E) and IV (F) of the oxidative phosphorylation system. The activity of the different complexes was normalized to that of citrate synthase and the results are presented as relative activities. Vertical bars represent SEM (n = 4 age- matched male mice for each genotype).

Mentions: To examine whether the reduced ATP content in rpS6P−/− muscle may result from mitochondrial deficiency, we measured the activity of citrate synthase that is commonly used as an assessment of mitochondrial content [23]. The results show indistinguishable activity of this enzyme in muscles from both genotypes (Figs 5A). Likewise, the activities of cytochrome C oxidase and four complexes of the oxidative phosphorylation system were similar in mitochondria from rpS6P−/− and rpS6P+/+ muscles (Figs 5B to 5F). Finally, measurements of oxygen consumption with glutamate and malate or succinate, as well as respiratory control ratio showed no significant difference between mitochondria from both genotypes (data not shown). These results suggest that the relative ATP deficiency in rpS6P−/− muscle cannot be attributed to either a decline in mitochondrial mass or the activity of the measured parameters. Nevertheless, our proteomic analysis has demonstrated a decline in three mitochondrial proteins in the knockin soleus (Table 1) that might impair the mitochondrial function.


Mice deficient in ribosomal protein S6 phosphorylation suffer from muscle weakness that reflects a growth defect and energy deficit.

Ruvinsky I, Katz M, Dreazen A, Gielchinsky Y, Saada A, Freedman N, Mishani E, Zimmerman G, Kasir J, Meyuhas O - PLoS ONE (2009)

The amount of mitochondria and the activity of complexes of the oxidative phosphorylation system are unchanged in rpS6P−/− muscle.Mitochondria were isolated from hind limb muscle of four age-matched rpS6P+/+ (WT) and rpS6P−/− (−/−) male mice and were assayed for citrate synthase (A), cytochrome C oxidase (B), as well as complexes I (C), II (D), II+III (E) and IV (F) of the oxidative phosphorylation system. The activity of the different complexes was normalized to that of citrate synthase and the results are presented as relative activities. Vertical bars represent SEM (n = 4 age- matched male mice for each genotype).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2682700&req=5

pone-0005618-g005: The amount of mitochondria and the activity of complexes of the oxidative phosphorylation system are unchanged in rpS6P−/− muscle.Mitochondria were isolated from hind limb muscle of four age-matched rpS6P+/+ (WT) and rpS6P−/− (−/−) male mice and were assayed for citrate synthase (A), cytochrome C oxidase (B), as well as complexes I (C), II (D), II+III (E) and IV (F) of the oxidative phosphorylation system. The activity of the different complexes was normalized to that of citrate synthase and the results are presented as relative activities. Vertical bars represent SEM (n = 4 age- matched male mice for each genotype).
Mentions: To examine whether the reduced ATP content in rpS6P−/− muscle may result from mitochondrial deficiency, we measured the activity of citrate synthase that is commonly used as an assessment of mitochondrial content [23]. The results show indistinguishable activity of this enzyme in muscles from both genotypes (Figs 5A). Likewise, the activities of cytochrome C oxidase and four complexes of the oxidative phosphorylation system were similar in mitochondria from rpS6P−/− and rpS6P+/+ muscles (Figs 5B to 5F). Finally, measurements of oxygen consumption with glutamate and malate or succinate, as well as respiratory control ratio showed no significant difference between mitochondria from both genotypes (data not shown). These results suggest that the relative ATP deficiency in rpS6P−/− muscle cannot be attributed to either a decline in mitochondrial mass or the activity of the measured parameters. Nevertheless, our proteomic analysis has demonstrated a decline in three mitochondrial proteins in the knockin soleus (Table 1) that might impair the mitochondrial function.

Bottom Line: Collectively, these experiments have demonstrated that the physical inferiority appears to result from two defects: a) a decrease in total muscle mass that reflects impaired growth, rather than aberrant differentiation of myofibers, as well as a diminished abundance of contractile proteins; and b) a reduced content of ATP and phosphocreatine, two readily available energy sources.However, the apparent energy deficiency in this genotype does not result from a lower mitochondrial mass or compromised activity of enzymes of the oxidative phosphorylation, nor does it reflect a decline in insulin-dependent glucose uptake, or diminution in storage of glycogen or triacylglycerol (TG) in the muscle.Interestingly, a similar role has been assigned for ribosomal protein S6 kinase 1, even though it regulates myoblast growth in an rpS6 phosphorylation-independent fashion.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, IMRIC, Hebrew University-Hadassah Medical School, Jerusalem, Israel.

ABSTRACT

Background: Mice, whose ribosomal protein S6 cannot be phosphorylated due to replacement of all five phosphorylatable serine residues by alanines (rpS6(P-/-)), are viable and fertile. However, phenotypic characterization of these mice and embryo fibroblasts derived from them, has established the role of these modifications in the regulation of the size of several cell types, as well as pancreatic beta-cell function and glucose homeostasis. A relatively passive behavior of these mice has raised the possibility that they suffer from muscle weakness, which has, indeed, been confirmed by a variety of physical performance tests.

Methodology/principal findings: A large variety of experimental methodologies, including morphometric measurements of histological preparations, high throughput proteomic analysis, positron emission tomography (PET) and numerous biochemical assays, were used in an attempt to establish the mechanism underlying the relative weakness of rpS6(P-/-) muscles. Collectively, these experiments have demonstrated that the physical inferiority appears to result from two defects: a) a decrease in total muscle mass that reflects impaired growth, rather than aberrant differentiation of myofibers, as well as a diminished abundance of contractile proteins; and b) a reduced content of ATP and phosphocreatine, two readily available energy sources. The abundance of three mitochondrial proteins has been shown to diminish in the knockin mouse. However, the apparent energy deficiency in this genotype does not result from a lower mitochondrial mass or compromised activity of enzymes of the oxidative phosphorylation, nor does it reflect a decline in insulin-dependent glucose uptake, or diminution in storage of glycogen or triacylglycerol (TG) in the muscle.

Conclusions/significance: This study establishes rpS6 phosphorylation as a determinant of muscle strength through its role in regulation of myofiber growth and energy content. Interestingly, a similar role has been assigned for ribosomal protein S6 kinase 1, even though it regulates myoblast growth in an rpS6 phosphorylation-independent fashion.

Show MeSH
Related in: MedlinePlus