Pavalko F M, Schneider G, Burridge K, Lim S S
Department of Physiology and Biophysics, Indiana University School of Medicine, Indianapolis 46202-5120, USA.
Exp Cell Res. 1995 Apr;217(2):534-40. doi: 10.1006/excr.1995.1119.
In this study we demonstrate that alpha-actinin is a prominent component of the focal adhesions of nonmuscle cells but that the alpha-actinin in focal adhesions is largely inaccessible to staining with antibodies against alpha-actinin. Our results explain a controversy that has existed in the literature. Investigators who microinject alpha-actinin into nonmuscle cells have routinely observed significant incorporation of alpha-actinin into focal adhesions as well as stress fibers. Immunofluorescence and immunoelectron microscopy have, however, indicated that alpha-actinin is located farther from the membrane than either talin or vinculin. Immunofluorescence studies of smooth muscle dense plaques and myotendinous junctions have also yielded conflicting results regarding the presence or absence of alpha-actinin at these sites. Here, we confirm that alpha-actinin immunofluorescence of fibroblasts yields weak or absent staining of focal adhesions. We also demonstrate that microinjected alpha-actinin readily incorporates into focal adhesions. However, various antisera against either the cell's endogenous alpha-actinin or against the microinjected chicken gizzard alpha-actinin fail to stain focal adhesions despite the presence of microinjected alpha-actinin at these sites. Furthermore, disassembly of stress fibers induced by dibutyrl cAMP demonstrates that alpha-actinin persists in focal adhesions in the absence of associated stress fibers, suggesting that alpha-actinin's association with focal adhesions is independent of stress fibers.
在本研究中，我们证明α - 辅肌动蛋白是非肌肉细胞粘着斑的主要成分，但粘着斑中的α - 辅肌动蛋白大多无法用抗α - 辅肌动蛋白抗体染色。我们的结果解释了文献中存在的一个争议。将α - 辅肌动蛋白显微注射到非肌肉细胞中的研究人员通常观察到α - 辅肌动蛋白大量掺入粘着斑以及应力纤维中。然而，免疫荧光和免疫电子显微镜表明，α - 辅肌动蛋白比踝蛋白或纽蛋白距离细胞膜更远。关于平滑肌致密斑和肌腱连接点处是否存在α - 辅肌动蛋白，平滑肌致密斑和肌腱连接点的免疫荧光研究也得出了相互矛盾的结果。在这里，我们证实成纤维细胞的α - 辅肌动蛋白免疫荧光产生的粘着斑染色较弱或无染色。我们还证明显微注射的α - 辅肌动蛋白很容易掺入粘着斑中。然而，尽管在这些部位存在显微注射的α - 辅肌动蛋白，但针对细胞内源性α - 辅肌动蛋白或针对显微注射的鸡砂囊α - 辅肌动蛋白的各种抗血清均未能对粘着斑进行染色。此外，二丁酰环磷腺苷诱导的应力纤维解体表明，在没有相关应力纤维的情况下，α - 辅肌动蛋白仍存在于粘着斑中，这表明α - 辅肌动蛋白与粘着斑的结合独立于应力纤维。
