Bartolini Francesca, Moseley James B, Schmoranzer Jan, Cassimeris Lynne, Goode Bruce L, Gundersen Gregg G
Department of Pathology, Anatomy and Cell Biology, Columbia University, New York, NY 10032, USA.
J Cell Biol. 2008 May 5;181(3):523-36. doi: 10.1083/jcb.200709029.
A critical microtubule (MT) polarization event in cell migration is the Rho/mDia-dependent stabilization of a subset of MTs oriented toward the direction of migration. Although mDia nucleates actin filaments, it is unclear whether this or a separate activity of mDia underlies MT stabilization. We generated two actin mutants (K853A and I704A) in a constitutively active version of mDia2 containing formin homology domains 1 and 2 (FH1FH2) and found that they still induced stable MTs and bound to the MT TIP proteins EB1 and APC, which have also been implicated in MT stabilization. A dimerization-impaired mutant of mDia2 (W630A) also generated stable MTs in cells. We examined whether FH1FH2mDia2 had direct activity on MTs in vitro and found that it bound directly to MTs, stabilized MTs against cold- and dilution-induced disassembly, and reduced the rates of growth and shortening during MT assembly and disassembly, respectively. These results indicate that mDia2 has a novel MT stabilization activity that is separate from its actin nucleation activity.
细胞迁移过程中一个关键的微管（MT）极化事件是Rho/mDia依赖的、朝向迁移方向定向排列的一部分微管的稳定化。尽管mDia可促使肌动蛋白丝成核，但尚不清楚MT稳定化是由mDia的这一功能还是其他独立活性所导致。我们在包含formin同源结构域1和2（FH1FH2）的组成型活性mDia2版本中构建了两个肌动蛋白突变体（K853A和I704A），发现它们仍能诱导产生稳定的微管，并与MT末端蛋白EB1和APC结合，而这两种蛋白也与MT稳定化有关。mDia2的一个二聚化受损突变体（W630A）在细胞中也能产生稳定的微管。我们检测了FH1FH2mDia2在体外对微管是否具有直接活性，发现它能直接与微管结合，稳定微管以抵抗低温和稀释诱导的解聚，并分别降低微管组装和解聚过程中的生长速率和缩短速率。这些结果表明，mDia2具有一种与其肌动蛋白成核活性相分离的新型微管稳定化活性。
