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The molecular basis of skeletal muscle phosphorylase kinase deficiency
Abstract:
The molecular basis of phosphorylase kinase deficiency was investigated in ICR/IAn mice, which show less than 0.2% of normal activity in skeletal muscle (Cohen, P.T.W. and Cohen, P., 1973). The genetics of the deficiency indicate it is a single gene defect on the X-chromosome (Lyon, J.B., 1970). Phosphorylase kinase was purified from skeletal muscle of a control strain, C3H/He-mg, by three different procedures. (a) Ammonium sulphate precipitation and gel filtration on Sepharose 4B. (b) Hydrophobic chromatography and affinity chromatography on Sepharose 4B to which antibody to rabbit muscle phosphorylase kinase has been linked covalently. (c) Precipitation from muscle extracts with anti-phosphorylase kinase antibody. All three procedures showed C3H/He-mg phosphorylase kinases were similar to the rabbit muscle enzymes, the structures of the two isoenzymes being (alphabetagamma)4 and (alpha'betagamma)4 respectively. The proportion of the (alpha'betagamma)4 isoenzyme relative to the (alphabetagamma)4 isoenzyme was however about 1:1 in murine muscle compared to about 1:10 in rabbit muscle. Since the alpha and alpha' subunits appear to be distinct gene products, the defect in ICR/IAn mice cannot be caused by a mutation in the genes coding for either the alpha or alpha'chains, or 50% of normal activity would be observed. All three procedures for C3H/He-mg mice failed to detect any of the four subunits alpha, alpha', beta and gamma in ICR/IAn mice, suggesting that all four chains are absent in the deficiency. An allele for the beta-subunit was identified in rabbits, and the inheritance of the allele showed that it was determined by an autosomal gene. Assuming conservation of X-linkage between mammals, the defect in ICR/IAn mice cannot be caused by a mutation in a beta-subunit gene. It is proposed that ICR/IAn mice are defective in a control gene located on the X-chromosome which is required for the expression of structural genes, at least one of which, the gene for the beta-subunit, is located on an autosome. The results imply that interchromosomal transfer of information takes place during the synthesis of phosphorylase kinase.
Insights
Phosphorylase kinase deficiency in ICR/IAn mice, a single X-chromosome gene defect, results in the absence of all four subunits. This suggests a regulatory gene defect impacting structural gene expression, not a mutation in the structural genes themselves.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Phosphorylase kinase deficiency in ICR/IAn mice exhibits <0.2% normal skeletal muscle activity.
- Genetic analysis indicates a single gene defect localized to the X-chromosome.
Purpose of the Study:
- Investigate the molecular basis of phosphorylase kinase deficiency in ICR/IAn mice.
- Characterize the structure and subunits of phosphorylase kinase in control and deficient mice.
Main Methods:
- Purification of phosphorylase kinase from C3H/He-mg mouse skeletal muscle using ammonium sulfate precipitation, gel filtration, hydrophobic, and affinity chromatography.
- Analysis of enzyme subunits using anti-phosphorylase kinase antibodies.
- Comparison of murine and rabbit muscle phosphorylase kinase isoenzymes.
Main Results:
- Murine phosphorylase kinase isoenzymes, (αβγ)4 and (α'βγ)4, were identified, with a 1:1 ratio in mice versus 1:10 in rabbits.
- Absence of all four subunits (α, α', β, γ) was detected in ICR/IAn mice.
- The β-subunit gene in rabbits is autosomal, ruling out X-linked mutation for this subunit in mice.
Conclusions:
- The deficiency in ICR/IAn mice is likely due to a defect in an X-chromosome regulatory gene controlling structural gene expression.
- This regulatory defect affects multiple structural genes, including the autosomal β-subunit gene.
- Interchromosomal information transfer is implied in phosphorylase kinase synthesis.