Endogenous cardiac Ca2+ channels do not overcome the E-C coupling defect in immortalized dysgenic muscle cells: evidence for a missing link
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1Institute of Molecular Pharmacology and Biophysics, University of Cincinnati, OH 45267-0828, USA.
Abstract:
The expression of subunit genes of the Ca2+ channel complex was studied in differentiating, immortalized mouse mdg cells. These cells expressed alpha 1 and alpha 2/delta transcripts of the skeletal muscle Ca2+ channel genes, a cardiac Ca2+ channel alpha 1 subunit gene and several known transcript variants of skeletal, cardiac and brain beta genes. The mdg mutation is retained in the 129DA3 cell line and occurs exclusively at nucleotide position 4010 in the skeletal alpha 1 transcript in which a cytosine residue is deleted. In early stages of differentiation and fusion, Ba2+ currents were detected in dysgenic myotubes the same as the cardiac L-type Ca2+ channel. These data provide specific structural evidence [Chaudhari, N. (1992) J. Biol. Chem. 267, 25636-25639] for the major genetic defect in mouse muscular dysgenesis and show a change in the expression levels of alpha 1S and alpha 1C. The upregulation of the expression of alpha 1C results in functional Ca2+ channel activity, however, presumably not sufficient for excitation-contraction coupling.
Insights
Mouse muscular dysgenesis involves a genetic defect in skeletal muscle calcium channels. Upregulation of cardiac calcium channels in these cells leads to some functional activity, but not enough for excitation-contraction coupling.
Area of Science:
- Molecular Biology
- Neuroscience
- Genetics
Background:
- The study investigates calcium (Ca2+) channel gene expression in differentiating mouse mdg cells, which carry a mutation associated with muscular dysgenesis.
- These cells express various transcripts for skeletal muscle, cardiac, and brain calcium channel subunits.
Purpose of the Study:
- To identify the specific genetic defect in mouse muscular dysgenesis.
- To analyze the expression of calcium channel subunit genes during cell differentiation.
- To understand the functional consequences of the mutation on calcium channel activity.
Main Methods:
- Analysis of Ca2+ channel subunit gene expression in immortalized mouse mdg cells (129DA3 cell line).
- Identification of the specific mutation site in the skeletal alpha 1 transcript.
- Electrophysiological recordings (Ba2+ currents) in differentiating myotubes to assess Ca2+ channel function.
Main Results:
- The mdg mutation is a deletion of a cytosine residue at nucleotide position 4010 in the skeletal alpha 1 transcript.
- Dysgenic myotubes exhibit Ba2+ currents similar to cardiac L-type Ca2+ channels during early differentiation.
- Expression levels of alpha 1S and alpha 1C subunits change, with alpha 1C upregulation leading to functional Ca2+ channel activity.
Conclusions:
- The identified mutation is the major genetic defect in mouse muscular dysgenesis.
- Upregulation of the alpha 1C subunit results in functional Ca2+ channel activity, though insufficient for excitation-contraction coupling.
- These findings provide structural evidence for the genetic defect and its impact on calcium channel function.