Cross-linking of actin filaments by myosin II is a major contributor to cortical integrity and cell motility in restrictive environments
Cross-linking of actin filaments by myosin II is a major contributor to cortical integrity and cell motility in restrictive environments
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DOI：
10.1242/jcs.00684
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发表时间：
2003-09-15
影响因子：
4
通讯作者：
Knecht, DA
中科院分区：
文献类型：
--
作者：
Laevsky, G;Knecht, DA
细胞经常需要在物理上限制运动的局部环境中移动，例如在外渗或转移性侵袭期间。为了模拟这些事件，我们开发了一种测定法，其中营养体盘基网柄藻在琼脂糖层下经历对叶酸源的趋化性[Laevsky，G.和Knecht，D. A.（2001年）的第10页。Biotechniques 31，1140-1149]。随着琼脂糖浓度从0.5%增加到3%，细胞在琼脂糖下移动的能力受到越来越多的抑制。肌球蛋白II和肌动蛋白交联蛋白在这种限制性环境中对细胞运动的贡献现在已经被研究。缺乏肌球蛋白II重链（mhcA（-））的细胞不能在大于0.5%的琼脂糖覆盖下迁移，即使在该浓度下，它们也只能从波谷移动很短的距离。当试图移动时，细胞由于无法缩回它们的尾足而变得拉伸和破碎。在较高的琼脂糖浓度下，mhcA（-）细胞在琼脂糖下伸出伪足，但不能将细胞体拉到下面。与肌球蛋白II在一般皮质稳定性中的作用一致，GFP-肌球蛋白动态地定位于琼脂糖下移动的细胞的外侧和后部皮质。缺乏肌球蛋白II必需轻链（mlcE（-））的细胞，没有可测量的肌球蛋白II运动活性，但能够在所有琼脂糖浓度下正常运动。缺乏ABP-120或α-辅肌动蛋白的突变体也能够以与野生型细胞相似的速率在琼脂糖下移动。我们假设，肌球蛋白稳定肌动蛋白皮质通过其交联活性，而不是它的运动功能，这种活动是必要的和足够的维持皮质完整性的细胞进行运动在一个限制性的环境。肌动蛋白交联剂α-辅肌动蛋白和ABP-120在提供皮质完整性方面似乎不像肌球蛋白II那样发挥重要作用。
Cells are frequently required to move in a local environment that physically restricts locomotion, such as during extravasation or metastatic invasion. In order to model these events, we have developed an assay in which vegetative Dictyostelium amoebae undergo chemotaxis under a layer of agarose toward a source of folic acid [Laevsky, G. and Knecht, D. A. (2001). Biotechniques 31, 1140-1149]. As the concentration of agarose is increased from 0.5% to 3 % the cells are increasingly inhibited in their ability to move under the agarose. The contribution of myosin II and actin cross-linking proteins to the movement of cells in this restrictive environment has now been examined. Cells lacking myosin II heavy chain (mhcA(-)) are unable to migrate under agarose overlays of greater than 0.5%, and even at this concentration they move only a short distance from the trough. While attempting to move, the cells become stretched and fragmented due to their inability to retract their uropods. At higher agarose concentrations, the mhcA(-) cells protrude pseudopods under the agarose, but are unable to pull the cell body underneath. Consistent with a role for myosin II in general cortical stability, GFP-myosin dynamically localizes to the lateral and posterior cortex of cells moving under agarose. Cells lacking the essential light chain of myosin II (mlcE(-)), have no measurable myosin II motor activity, yet were able to move normally under all agarose concentrations. Mutants lacking either ABP-120 or alpha-actinin were also able to move under agarose at rates similar to wild-type cells. We hypothesize that myosin stabilizes the actin cortex through its cross-linking activity rather than its motor function and this activity is necessary and sufficient for the maintenance of cortical integrity of cells undergoing movement in a restrictive environment. The actin cross-linkers alpha-actinin and ABP-120 do not appear to play as major a role as myosin II in providing this cortical integrity.