No mechanical role for vinculin in strain transduction in primary bovine osteoblasts.
Vinculin is thought to play a central role in linking the actin cytoskeleton to the integrin adhesion proteins of focal contacts and also in cell-cell adhesion sites. We have investigated aspects of attachment and the transduction of mechanical stimulation on the function of vinculin and its assembly in primary bovine osteoblasts. The dynamic attachment process was concomitant with the reassembly of vinculin from a soluble cytoplasmic pool into Triton-insoluble focal adhesions. Immunoblotting experiments demonstrated a slight increase in the amount of vinculin concentrated in focal contacts that was paralleled by a discrete decrease in the Triton-soluble fraction of cytoplasmic vinculin. Sixty cycles of 1 Hz deformations at 0.02% (hypophysiological, no change in cell division rate), 0.2% (within the normal physiological range), and 1% (hyperphysiological) resulted in a rapid and reversible disassembly of vinculin from focal contacts to a homogeneous cytoplasmic localization, although alterations in the shape and morphology of cells could not be detected. Although all mechanical loading protocols dramatically depleted vinculin from focal contacts, its initial distribution in adhesion plaques was fully restored within the next 60 min, demonstrating the highly dynamic and reversible shift of this protein from a membrane-associated pool to the cytoplasm. By first depleting vinculin from the focal adhesion sites by applying 0.02% strain, waiting 60 s, and then applying 0.2% strain, we show that the transduction of mechanical deformation was identical with controls that had not been pretreated. This indicates that vinculin does not play a role in mechanical transduction, that significant mechanical forces are not transmitted through vinculin, and also that loss of vinculin at the focal adhesion site does not have significant effects, in the short term, on cell adhesion.
Meyer
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U
Department of Maxillofacial Surgery, University of Münster, Germany.
Meyer
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Jones
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D000818:Animals; D001696:Biomechanical Phenomena; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002448:Cell Adhesion; D002455:Cell Division; D002478:Cells, Cultured; D004492:Edetic Acid / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D008856:Microscopy, Fluorescence; D010006:Osteoblasts / Q000502:physiology*; D016596:Vinculin / Q000502:physiology*
D016596:Vinculin; D004492:Edetic Acid; D002118:Calcium