McNeely Katrina C, Cupp Timothy D, Little Jessica Neville, Janisch Kerstin M, Shrestha Ayushma, Dwyer Noelle D
Department of Cell Biology, University of Virginia, Charlottesville, VA, 22908, USA.
Neural Dev. 2017 Mar 31;12(1):5. doi: 10.1186/s13064-017-0082-5.
How neurons change their cytoskeleton to adopt their complex polarized morphology is still not understood. Growing evidence suggests that proteins that help build microtubule structures during cell division are also involved in building and remodeling the complex cytoskeletons of neurons. Kif20b (previously called MPP1 or Mphosph1) is the most divergent member of the Kinesin-6 family of "mitotic" kinesins that also includes Kif23/MKLP1 and Kif20a/MKLP2. We previously isolated a loss-of-function mouse mutant of Kif20b and showed that it had a thalamocortical axon guidance defect and microcephaly.
We demonstrate here, using the mouse mutant, that Kif20b is required for neuron morphogenesis in the embryonic neocortex. In vivo and in vitro cortical neurons were labeled and imaged to analyze various aspects of morphogenesis.
Loss of Kif20b disrupts polarization as well as neurite outgrowth, branching and caliber. In vivo, mutant cortical neurons show defects in orientation, and have shorter thinner apical dendrites that branch closer to the cell body. In vitro, without external polarity cues, Kif20b mutant neurons show a strong polarization defect. This may be due in part to loss of the polarity protein Shootin1 from the axonal growth cone. Those mutant neurons that do succeed in polarizing have shorter axons with more branches, and longer minor neurites. These changes in shape are not due to alterations in cell fate or neuron layer type. Surprisingly, both axons and minor neurites of mutant neurons have increased widths and longer growth cone filopodia, which correlate with abnormal microtubule organization. Live analysis of axon extension shows that Kif20b mutant axons display more variable growth with increased retraction.
These results demonstrate that Kif20b is required cell-autonomously for proper morphogenesis of cortical pyramidal neurons. Kif20b regulates neuron polarization, and axon and dendrite branching, outgrowth, and caliber. Kif20b protein may act by bundling microtubules into tight arrays and by localizing effectors such as Shootin1. Thus it may help shape neurites, sustain consistent axon growth, and inhibit branching. This work advances our understanding of how neurons regulate their cytoskeleton to build their elaborate shapes. Finally, it suggests that neuronal connectivity defects may be present in some types of microcephaly.
神经元如何改变其细胞骨架以形成复杂的极化形态仍不清楚。越来越多的证据表明，在细胞分裂过程中帮助构建微管结构的蛋白质也参与构建和重塑神经元复杂的细胞骨架。Kif20b（以前称为MPP1或Mphosph1）是驱动蛋白-6家族中“有丝分裂”驱动蛋白中差异最大的成员，该家族还包括Kif23/MKLP1和Kif20a/MKLP2。我们之前分离出了Kif20b的功能缺失小鼠突变体，并表明它存在丘脑皮质轴突导向缺陷和小头畸形。
我们在此利用该小鼠突变体证明，Kif20b是胚胎新皮质中神经元形态发生所必需的。对体内和体外的皮质神经元进行标记和成像，以分析形态发生的各个方面。
Kif20b的缺失会破坏极化以及神经突的生长、分支和直径。在体内，突变的皮质神经元表现出定向缺陷，其顶端树突更短更细，且分支更靠近细胞体。在体外，在没有外部极性线索的情况下，Kif20b突变神经元表现出强烈的极化缺陷。这可能部分归因于轴突生长锥中极性蛋白Shootin1的缺失。那些成功极化的突变神经元轴突更短且分支更多，次要神经突更长。这些形状变化并非由于细胞命运或神经元层类型的改变。令人惊讶的是，突变神经元的轴突和次要神经突宽度均增加，生长锥丝状伪足更长，这与微管组织异常相关。对轴突延伸的实时分析表明，Kif20b突变轴突表现出更多变的生长且回缩增加。
这些结果表明，Kif20b是皮质锥体神经元正常形态发生所必需的细胞自主因子。Kif20b调节神经元极化以及轴突和树突的分支、生长和直径。Kif20b蛋白可能通过将微管捆绑成紧密阵列并定位诸如Shootin1等效应物来发挥作用。因此，它可能有助于塑造神经突、维持一致的轴突生长并抑制分支。这项工作推进了我们对神经元如何调节其细胞骨架以构建其精细形状的理解。最后，这表明在某些类型的小头畸形中可能存在神经元连接缺陷。
