Myosin remodelling in the contracting A7r5 smooth muscle cell.
OBJECTIVE
Using confocal microscopy and standard immunohistochemical techniques, changes in the structure of alpha-actin and beta-actin as well as the distribution of myosin II were studied in the contracting A7r5 smooth muscle cell.


RESULTS
Prior to stimulation, each of the three proteins were incorporated into filamentous structures extending the length of the cell body. At 30 min after stimulation by phorbol 12, 13 dibutyrate (PDBu), the system of densely packed beta-actin fibres was retained. By comparison, alpha-actin and myosin were observed to undergo significant remodelling, characterized by loss in fibre structure and the formation of brightly fluorescing peripheral bodies. Co-immunoprecipitation of alpha-actin and myosin II suggested an association between the proteins. Consistent with this observation, dual immunostaining for alpha-actin and myosin revealed strong co-localization of the two proteins prior to stimulation. Following PDBu stimulation myosin II was observed to partially disassociate from alpha-actin structure but showed significant co-localization with alpha-actin filaments and peripheral bodies throughout the interval of contraction. The use of cytochalasin B to block actin polymerization or the selective dissolution of alpha-actin cable structure by thapsigargin produced similar patterns of change in alpha-actin structure and the localization of myosin II.


CONCLUSIONS
The results support the concept of myosin liability and potential for remodelling. The results suggest that myosin undergoes extensive relocalization in association with alpha-actin remodelling which may be an important determinant of contractile function in the A7r5 smooth muscle cell.
Fultz
M E
ME
Department of Physiology, The Joan Edwards School of Medicine, Marshall University, Huntington, WV 25704, USA.
Wright
G L
GL
D000199:Actins / Q000032:analysis / Q000502:physiology; D000818:Animals; D002478:Cells, Cultured; D003571:Cytochalasin B / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007150:Immunohistochemistry / Q000379:methods; D018613:Microscopy, Confocal / Q000379:methods; D009119:Muscle Contraction / Q000502:physiology*; D009130:Muscle, Smooth / Q000187:drug effects / Q000502:physiology*; D024462:Myosin Type II / Q000032:analysis / Q000502:physiology*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology
D000199:Actins; D004791:Enzyme Inhibitors; D015240:Phorbol 12,13-Dibutyrate; D003571:Cytochalasin B; D019284:Thapsigargin; D024462:Myosin Type II