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Role of Ca2+ and Ca2+-activated protease in myoblast fusion
Abstract:
In this report, we have examined the effects of a calcium chelator, EGTA, and a calcium ionophore, A23187, on fusion of a cloned muscle cell line, L6. Our results confirm that EGTA essentially blocks all myoblast fusion because the lateral alignment of presumptive myoblasts cannot occur in the absence of extracellular calcium. A23187, however, promotes the precocious fusion of myoblasts, apparently by facilitating Ca2+ transport into myoblasts. We have also demonstrated that a Ca2+-activated protease, CAF (mM), appears to relocate in response to the Ca2+ flux, changing from a random, dispersed distribution in proliferative myoblasts to a predominantly peripheral distribution in prefusion myoblasts. Coincident with the mM CAF relocation is an altered distribution of a surface glycoprotein, fibronectin. Extracellular fibronectin is seen in abundance in proliferating myoblasts, but is essentially absent from the surface of fusing myoblasts. We suggest that mM CAF when activated by Ca2+ influx may act to promote the release of fibronectin from the myoblast cell surface, thus providing a mechanism by which the membrane of the fusing myoblast may be rearranged to accommodate fusion.
Insights
Extracellular calcium is essential for myoblast fusion. Calcium influx triggers a protease (CAF) to relocate, potentially releasing fibronectin and enabling muscle cell fusion.
Area of Science:
- Cell Biology
- Muscle Development
- Biochemistry
Background:
- Myoblast fusion is critical for skeletal muscle formation.
- The role of calcium ions (Ca2+) in regulating myoblast fusion is not fully understood.
- Specific molecular events mediating fusion remain to be elucidated.
Purpose of the Study:
- To investigate the effects of calcium modulation on L6 myoblast fusion.
- To identify molecular changes associated with calcium-induced fusion.
- To explore the role of calcium-activated protease (CAF) and fibronectin in myoblast fusion.
Main Methods:
- Utilized L6 muscle cell line for fusion studies.
- Applied EGTA (calcium chelator) and A23187 (calcium ionophore) to manipulate extracellular and intracellular calcium levels.
- Examined cellular localization of CAF and fibronectin using microscopy.
Main Results:
- EGTA blocked myoblast fusion by preventing lateral alignment due to lack of extracellular calcium.
- A23187 promoted premature myoblast fusion, suggesting facilitated Ca2+ transport.
- Calcium influx induced relocation of Ca2+-activated protease (CAF) from a dispersed to a peripheral distribution.
- Fibronectin, abundant in proliferating myoblasts, was absent on the surface of fusing myoblasts.
Conclusions:
- Extracellular calcium is indispensable for myoblast alignment and subsequent fusion.
- Calcium influx appears to trigger CAF relocation and subsequent fibronectin release from the cell surface.
- This mechanism involving CAF and fibronectin may facilitate membrane rearrangement necessary for myoblast fusion.