Immunolocalization of muscle and nonmuscle isoforms of actin in myogenic cells and adult skeletal muscle.
In vertebrate skeletal muscle, the proliferating myoblasts synthesize nonmuscle isoforms of actin, and the cells begin to express muscle-specific actin isoforms during their myogenic differentiation. To study the distributions of the actin isoforms in myogenic cells and fully differentiated skeletal muscle, we prepared a peptide antibody specific for the skeletal alpha isoform of actin and used this antibody along with an antibody specifically reactive with nonmuscle gamma actin to stain cultured myotubes and adult skeletal myofibrils by double-indirect immunofluorescence. At this level of resolution, no differences in isoform localization were seen: Both muscle and nonmuscle actins were detected in the myotubes and in the striations of mature myofibrils. Myotubes were also double-stained using immunogold electron microscopy, and the isoform distributions were determined quantitatively by counting the two sizes of gold particles that corresponded to labeling with each antibody. A quantitative analysis of immunoreactivity revealed that, although both forms were present in all actin-containing structures, nonmuscle actin was relatively more prevalent along the edges (cortical microfilaments) of the myotubes, whereas the muscle isoform predominated in the interior regions (containing forming myofibrils). Thus, we have found evidence of a heterogeneous distribution of muscle and nonmuscle actin isoforms in differentiating myogenic cells, and we have demonstrated that a nonmuscle actin isoform is a component of the muscle contractile apparatus.
Otey
C A
CA
Department of Biology, University of California, Los Angeles.
Kalnoski
M H
MH
Bulinski
J C
JC
D000199:Actins / Q000032:analysis*; D000818:Animals; D002455:Cell Division; D002478:Cells, Cultured; D005260:Female; D005455:Fluorescent Antibody Technique; D006168:Guinea Pigs; D007525:Isoelectric Focusing; D008854:Microscopy, Electron; D009132:Muscles / Q000378:metabolism* / Q000648:ultrastructure; D009210:Myofibrils / Q000648:ultrastructure; D011817:Rabbits
D000199:Actins