Nagasaki Akira, Kanada Masamitsu, Uyeda Taro Qp
Research Institute for Cell Engineering, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8562, Japan.
Cell Res. 2009 Feb;19(2):236-46. doi: 10.1038/cr.2008.318.
To investigate the roles of substrate adhesion in cytokinesis, we established cell lines lacking paxillin (PAXB) or vinculin (VINA), and those expressing the respective GFP fusion proteins in Dictyostelium discoideum. As in mammalian cells, GFP-PAXB and GFP-VINA formed focal adhesion-like complexes on the cell bottom. paxB(-) cells in suspension grew normally, but on substrates, often failed to divide after regression of the furrow. The efficient cytokinesis of paxB(-) cells in suspension is not because of shear forces to assist abscission, as they divided normally in static suspension culture as well. Double knockout strains lacking mhcA, which codes for myosin II, and paxB or vinA displayed more severe cytokinetic defects than each single knockout strain. In mitotic wild-type cells, GFP-PAXB was diffusely distributed on the basal membrane, but was strikingly condensed along the polar edges in mitotic mhcA(-) cells. These results are consistent with our idea that Dictyostelium displays two forms of cytokinesis, one that is contractile ring-dependent and adhesion-independent, and the other that is contractile ring-independent and adhesion-dependent, and that the latter requires PAXB and VINA. Furthermore, that paxB(-) cells fail to divide normally in the presence of substrate adhesion suggests that this adhesion molecule may play additional signaling roles.
为了研究底物黏附在胞质分裂中的作用,我们在盘基网柄菌中建立了缺乏桩蛋白(PAXB)或纽蛋白(VINA)的细胞系,以及表达各自绿色荧光蛋白(GFP)融合蛋白的细胞系。与哺乳动物细胞一样,GFP-PAXB和GFP-VINA在细胞底部形成了黏着斑样复合物。悬浮培养的paxB(-)细胞生长正常,但在底物上,沟回退之后常常无法分裂。paxB(-)细胞在悬浮培养时高效的胞质分裂并非由于剪切力有助于脱离,因为它们在静态悬浮培养中也能正常分裂。缺乏编码肌球蛋白II的mhcA以及paxB或vinA的双敲除菌株表现出比每个单敲除菌株更严重的胞质分裂缺陷。在有丝分裂的野生型细胞中,GFP-PAXB在基底膜上呈弥散分布,但在有丝分裂的mhcA(-)细胞中,它沿着极边缘显著浓缩。这些结果与我们的观点一致,即盘基网柄菌表现出两种胞质分裂形式,一种是依赖收缩环且不依赖黏附的,另一种是不依赖收缩环且依赖黏附的,并且后者需要PAXB和VINA。此外,paxB(-)细胞在存在底物黏附的情况下无法正常分裂,这表明这种黏附分子可能发挥额外的信号作用。