Filamin depletion blocks endoplasmic spreading and destabilizes force-bearing adhesions.
Filamin depletion blocks endoplasmic spreading and destabilizes force-bearing adhesions.
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DOI：
10.1091/mbc.e10-08-0661
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发表时间：
2011-04-15
影响因子：
3.3
通讯作者：
Sheetz MP
中科院分区：
文献类型：
--
作者：
Lynch CD;Gauthier NC;Biais N;Lazar AM;Roca-Cusachs P;Yu CH;Sheetz MP
观察到严重缺乏细丝的细胞具有许多运动相关的缺陷，包括内质网扩散缺陷;更小、更动态的局灶性粘连;以及无法维持高水平的牵引力。作为一个单独的机械单位，通过拉力传播的内质也进行了讨论。细胞运动是一个重要的过程，它依赖于一个连贯的，交联的肌动蛋白细胞骨架，物理协调许多结构和信号分子的行动。肌动蛋白交联蛋白，细丝蛋白（Fln），已被牵连在三维皮质肌动蛋白网络的支持，既能够保持细胞的完整性和承受大的力量。尽管许多研究已经检查了缺乏多种Fln同种型之一的细胞，但补偿机制可以掩盖仅通过进一步Fln耗尽观察到的新表型。事实上，在FlnB-/-小鼠胚胎成纤维细胞（MEF）中，shRNA介导的FlnA敲低导致如通过内质网标记物检测到的新的内质网铺展缺陷。微管（MT）的延伸率也下降，但不是由周边肌动蛋白流，因为这也是减少在Fln耗尽系统。此外，Fln耗尽的MEF表现出降低的粘附稳定性，其表现为细胞边缘的褶皱增加、粘附尺寸减小、瞬时牵引力和应力纤维减少。FlnA-/- MEFs，而不是FlnB-/- MEFs，也显示出中等缺陷的内质网扩展，其特征在于初始延伸，然后突然收缩和应力纤维断裂。FlnA定位于胞质、粘连和应力纤维周围的肌动蛋白连接。因此，我们认为，Flns有一个主要的作用，在维护肌动蛋白为基础的机械连接，使内质网的蔓延和MT的延伸，以及持续的牵引力和成熟的局灶性粘连。
Cells severely depleted of filamins were observed to have numerous motility-related defects, including a defect in endoplasmic spreading; smaller, more dynamic focal adhesions; and an inability to sustain high levels of traction force. The endoplasm as a separate mechanical unit spread by pulling forces is also discussed. Cell motility is an essential process that depends on a coherent, cross-linked actin cytoskeleton that physically coordinates the actions of numerous structural and signaling molecules. The actin cross-linking protein, filamin (Fln), has been implicated in the support of three-dimensional cortical actin networks capable of both maintaining cellular integrity and withstanding large forces. Although numerous studies have examined cells lacking one of the multiple Fln isoforms, compensatory mechanisms can mask novel phenotypes only observable by further Fln depletion. Indeed, shRNA-mediated knockdown of FlnA in FlnB–/– mouse embryonic fibroblasts (MEFs) causes a novel endoplasmic spreading deficiency as detected by endoplasmic reticulum markers. Microtubule (MT) extension rates are also decreased but not by peripheral actin flow, because this is also decreased in the Fln-depleted system. Additionally, Fln-depleted MEFs exhibit decreased adhesion stability that appears in increased ruffling of the cell edge, reduced adhesion size, transient traction forces, and decreased stress fibers. FlnA–/– MEFs, but not FlnB–/– MEFs, also show a moderate defect in endoplasm spreading, characterized by initial extension followed by abrupt retractions and stress fiber fracture. FlnA localizes to actin linkages surrounding the endoplasm, adhesions, and stress fibers. Thus we suggest that Flns have a major role in the maintenance of actin-based mechanical linkages that enable endoplasmic spreading and MT extension as well as sustained traction forces and mature focal adhesions.