Fukui Y, de Hostos E, Yumura S, Kitanishi-Yumura T
Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Exp Cell Res. 1999 May 25;249(1):33-45. doi: 10.1006/excr.1999.4445.
Eupodia are F-actin-containing cortical structures similar to vertebrate podosomes or invadopodia found in metastatic cells. Eupodia are rich in alpha-actinin and myosin IB/D, but not a Dictyostelium homologue of talin. In the present study, we localized other actin-binding proteins, ABP120, cofilin, coronin, and fimbrin, in the eupodia and examined the three-dimensional organization of their F-actin system by confocal microscopy and transmission electron microscopy. To examine their function, we analyzed the assembly and disassembly dynamics of the F-actin system in eupodia and its relation to lamellipodial protrusion. Actin dynamics was examined by monitoring S65T-GFP-coronin and rhodamine-actin using a real-time confocal unit and a digital microscope system. Fluorescence morphometric analysis demonstrates the presence of a precise spatiotemporal coupling between F-actin assembly in eupodia and lamellipodial protrusion. When a lamellipodium advances to invade a tight space, additional rows of eupodia are sequentially formed at the base of that lamellipodium. These results indicate that mechanical stress at the leading edge modulates the structural integrity of actin and its binding proteins, such that eupodia are formed when anchorage is needed to boost for invasive protrusion of the leading edge.
伪足是含有F-肌动蛋白的皮质结构，类似于在转移细胞中发现的脊椎动物的足体或侵袭性伪足。伪足富含α-辅肌动蛋白和肌球蛋白IB/D，但不含有盘基网柄菌中与踝蛋白同源的蛋白。在本研究中，我们在伪足中定位了其他肌动蛋白结合蛋白，ABP120、丝切蛋白、冠蛋白和丝束蛋白，并通过共聚焦显微镜和透射电子显微镜检查了它们的F-肌动蛋白系统的三维结构。为了研究它们的功能，我们分析了伪足中F-肌动蛋白系统的组装和解聚动力学及其与片状伪足突出的关系。通过使用实时共聚焦装置和数字显微镜系统监测S65T-GFP-冠蛋白和罗丹明-肌动蛋白来检测肌动蛋白动力学。荧光形态计量分析表明，伪足中F-肌动蛋白组装与片状伪足突出之间存在精确的时空耦合。当片状伪足向前推进以侵入狭窄空间时，在该片状伪足的基部会依次形成额外的几排伪足。这些结果表明，前沿的机械应力调节肌动蛋白及其结合蛋白的结构完整性，从而在需要锚定以促进前沿侵入性突出时形成伪足。
