Differentiation defective mutants of skeletal myoblasts altered in a gelatin-binding glycoprotein

Area of Science:

• Biochemistry
• Cell Biology
• Developmental Biology

Background:

• A 46,000 molecular mass cell surface glycoprotein (gp46) is linked to myoblast differentiation.
• gp46 exhibits specific binding to gelatin-Sepharose, similar to a previously identified 47,000 molecular mass glycoprotein.

Purpose of the Study:

• To investigate the role of gp46 in myoblast differentiation.
• To analyze gp46 expression in wild-type L6 myoblasts and differentiation-negative mutants (D-1, C-8).

Main Methods:

• Utilized gelatin-Sepharose binding assays to assess gp46 properties.
• Examined gp46 expression using Western blotting with a monoclonal antibody against gp46.
• Analyzed gp46 localization in plasma membrane and endoplasmic reticulum fractions.

Main Results:

• gp46 binds specifically to gelatin-Sepharose.
• In mutant D-1 (dolichol mannosyl transferase defect), mannose incorporation and ConA binding to gp46 were reduced, but gelatin adhesion was unaffected.
• gp46 expression was normal in D-1 but reduced in mutant C-8, affecting both plasma membrane and endoplasmic reticulum.
• gp46 expression remained constant during replication and fusion but decreased post-fusion in L6 myoblasts.
• gp46 expression was invariant in non-fusing mutants and in L6 cells prevented from fusing.

Conclusions:

• Collagen interactions mediated by gp46 are crucial for normal skeletal muscle differentiation in rats.
• gp46 expression dynamics correlate with myoblast fusion and differentiation stages.