Failure to make normal α ryanodine receptor is an early event associated with the Crooked Neck Dwarf (cn) mutation in chicken

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Abstract
We have investigated the molecular basis of the Crooked Neck Dwarf (cn) mutation in embryonic chickens. Using biochemical and pharmacological techniques we are unable to detect normal α ryanodine receptor (RyR) protein in intact cn/cn skeletal muscle. Extremely low levels of αRyR immunoreactivity can be observed in mutant muscles, but the distribution of this staining differs from that in normal muscle and colocalizes with the rough endoplasmic reticulum immunoglobulin binding protein, BiP. This suggests the existence of an abnormal αRyR protein in mutant muscle. In day E12 cn/cn muscle the levels of RyR mRNA are reduced by ∼80%, while the levels of other muscle proteins, including the α1 subunit of the dihydropyridine receptor, the SRCa2+-ATPase, calsequestrin, and glyceraldehyde-3-phosphate dehydrogenase, and their associated mRNAs are essentially normal in cn/cn muscle. There is also a failure to express αRyR in cn/cn cerebellar Purkinje neurons. Expression of the βRyR, a second RyR isoform, is not initiated in normal skeletal muscle until day E18. In cn/cn skeletal muscle significant muscle degeneration has occurred by this time and the βRyR is found at low levels in only a subset of fibers suggesting the reduced levels of this isoform are a secondary consequence of the mutation. The cardiac RyR isoform is found in cn/cn cardiac muscle, which contracts in a vigorous manner. In summary, a failure to make normal αRyR receptor appears to be an event closely associated with the cn mutation and one which may be largely responsible for development of the cn/cn phenotype in embryonic skeletal muscle.