Non-sarcomeric mode of myosin II organization in the fibroblast lamellum.
The organization of myosin in the fibroblast lamellum was studied by correlative fluorescence and electron microscopy after a novel procedure to reveal its underlying morphology. An X-rhodamine analog of conventional smooth muscle myosin (myosin II) that colocalized after microinjection with endogenous myosin was used to trace myosin distribution in living fibroblasts. Then, the same cells were examined by EM of platinum replicas. To visualize the structural arrangement of myosin, other cytoskeletal fibrillar structures had to be removed: microtubules were depolymerized by nocodazole treatment of the living cells before injection of myosin; continued nocodazole treatment also induced the intermediate filaments to concentrate near the nucleus, thus removing them from the lamellar region; actin filaments were removed after lysis of the cells by incubation of the cytoskeletons with recombinant gelsolin. Possible changes in myosin organization caused by this treatment were examined by fluorescence microscopy. No significant differences in myosin distribution patterns between nocodazole-treated and control cells were observed. Cell lysis and depletion of actin also did not induce reorganization of myosin as was shown by direct comparison of myosin distribution in the same cells in the living state and after gelsolin treatment. EM of the well-spread, peripheral regions of actin-depleted cytoskeletons revealed a network of bipolar myosin mini-filaments, contracting each other at their terminal, globular regions. The morphology of this network corresponded well to the myosin distribution observed by fluorescence microscopy. A novel mechanism of cell contraction by folding of the myosin filament network is proposed.
Verkhovsky
A B
AB
A. N. Belozersky Laboratory of Molecular Biology, Moscow State University, Russia.
Borisy
G G
GG
D016475:3T3 Cells; D008841:Actin Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D000199:Actins / Q000302:isolation & purification / Q000378:metabolism; D000818:Animals; D003599:Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure*; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism / Q000648:ultrastructure; D005895:Gizzard, Avian; D051379:Mice; D008845:Microinjections; D008854:Microscopy, Electron; D009218:Myosins / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D012235:Rhodamines; D012518:Sarcomeres / Q000502:physiology; D014422:Turkeys
D000199:Actins; D012235:Rhodamines; D009218:Myosins