Differences in elasticity of vinculin-deficient F9 cells measured by magnetometry and atomic force microscopy
Differences in elasticity of vinculin-deficient F9 cells measured by magnetometry and atomic force microscopy
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DOI：
10.1006/excr.1997.3915
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发表时间：
1998-03-15
影响因子：
3.7
通讯作者：
Ezzell, RM
中科院分区：
文献类型：
--
作者：
Goldmann, WH;Galneder, R;Ezzell, RM
我们已经研究了小鼠F9胚胎癌细胞系，其中两个黏着斑蛋白基因都通过同源重组失活，其表现出有缺陷的粘附和扩散[Cell ct al.（1995）Proc.Natl. Acad. Sci. USA 92，9161-9165]。使用磁力计和RGD涂层的磁性微珠，我们测量了局部的损失和更换粘着斑蛋白的机械力传递的影响。与野生型细胞相比，曲马多蛋白缺陷型F9 Vin（-/-）细胞显示出21%的相对刚度差异。在转染和组成型表达增加量的黏着斑蛋白到F9 Vin（-/-）细胞中后，这恢复到接近野生型水平。相比之下，在F9 Vin（-/-）细胞中转染缺乏氨基酸1-288（含有talin和α-辅肌动蛋白结合位点）或用酪氨酸取代苯丙氨酸（磷酸化位点，氨基酸822）的黏着斑蛋白构建体导致刚度的部分恢复。使用原子力显微镜通过128 x 128（n = 16，384）力扫描绘制整个F9细胞的相对弹性，我们观察到与磁力计测量的相关性。这些研究结果表明，黏着斑蛋白可能通过稳定粘着斑和转移驱动细胞骨架重塑的机械应力来促进细胞粘附和扩散，从而影响细胞的弹性特性。(C)北京：科学出版社.
We have investigated a mouse F9 embryonic carcinoma cell line, in which both vinculin genes were inactivated by homologous recombination, that exhibits defective adhesion and spreading [Cell ct al. (1995) Proc. Natl. Acad. Sci. USA 92, 9161-9165]. Using a magnetometer and RGD-coated magnetic microbeads, we measured the local effect of loss and replacement of vinculin on mechanical force transfer across integrins. Vinculin-deficient F9Vin(-/-) cells showed a 21% difference in relative stiffness compared to wildtype cells. This was restored to near wild-type levels after transfection and constitutive expression of increasing amounts of vinculin into F9Vin(-/-) cells. In contrast, the transfection of vinculin constructs deficient in amino acids 1-288 (containing the talin-and alpha-actinin-binding site) or substituting tyrosine for phenylalanine (phosphorylation site, amino acid 822) in F9Vin(-/-) cells resulted in partial restoration of stiffness. Using atomic force microscopy to map the relative elasticity of entire F9 cells by 128 x 128 (n = 16,384) force scans, we observed a correlation with magnetometer measurements. These findings suggest that vinculin may promote cell adhesion and spreading by stabilizing focal adhesions and transferring mechanical stresses that drive cytoskeletal remodeling, thereby affecting the elastic properties of the cell. (C) 1998 Academic Press.