Weiner Alexis T, Lanz Michael C, Goetschius Daniel J, Hancock William O, Rolls Melissa M
Huck Institutes of the Life Sciences and Biochemistry and Molecular Biology, the Pennsylvania State University, University Park, Pennsylvania.
Biomedical Engineering, the Pennsylvania State University, University Park, Pennsylvania.
Cytoskeleton (Hoboken). 2016 Jan;73(1):35-44. doi: 10.1002/cm.21270.
In Drosophila neurons, kinesin-2, EB1 and Apc are required to maintain minus-end-out dendrite microtubule polarity, and we previously proposed they steer microtubules at branch points. Motor-mediated steering of microtubule plus ends could be accomplished in two ways: 1) by linking a growing microtubule tip to the side of an adjacent microtubule as it navigates the branch point (bundling), or 2) by directing a growing microtubule after a collision with a stable microtubule (collision resolution). Using live imaging to distinguish between these two mechanisms, we found that reduction of kinesin-2 did not alter the number of microtubules that grew along the edge of the branch points where stable microtubules are found. However, reduction of kinesin-2 or Apc did affect the number of microtubules that slowed down or depolymerized as they encountered the side of the branch opposite to the entry point. These results are consistent with kinesin-2 functioning with Apc to resolve collisions. However, they do not pinpoint stable microtubules as the collision partner as stable microtubules are typically very close to the membrane. To determine whether growing microtubules were steered along stable ones after a collision, we analyzed the behavior of growing microtubules at dendrite crossroads where stable microtubules run through the middle of the branch point. In control neurons, microtubules turned in the middle of the crossroads. However, when kinesin-2 was reduced some microtubules grew straight through the branch point and failed to turn. We propose that kinesin-2 functions to steer growing microtubules along stable ones following collisions.
在果蝇神经元中,驱动蛋白-2、EB1和Apc是维持负极向外的树突微管极性所必需的,我们之前提出它们在分支点引导微管。由马达介导的微管正端引导可以通过两种方式实现:1)当生长的微管尖端在穿过分支点时,将其与相邻微管的侧面相连(捆绑);或2)在与稳定微管碰撞后引导生长的微管(碰撞解决)。通过实时成像来区分这两种机制,我们发现驱动蛋白-2的减少并没有改变沿着发现稳定微管的分支点边缘生长的微管数量。然而,驱动蛋白-2或Apc的减少确实影响了那些在遇到与入口点相对的分支侧面时减速或解聚的微管数量。这些结果与驱动蛋白-2与Apc共同作用来解决碰撞的观点一致。然而,由于稳定微管通常非常靠近膜,它们并没有将稳定微管确定为碰撞伙伴。为了确定生长的微管在碰撞后是否沿着稳定微管被引导,我们分析了在稳定微管穿过分支点中间的树突交叉点处生长的微管的行为。在对照神经元中,微管在交叉点中间转弯。然而,当驱动蛋白-2减少时,一些微管直接穿过分支点而没有转弯。我们提出驱动蛋白-2的功能是在碰撞后引导生长的微管沿着稳定微管。