Cell type-variation of cytokinesis

在动物细胞中，胞质分裂是由肌动球蛋白收缩环的收缩驱动的， 其由来自纺锤体微管的信号定位和控制。长期以来，人们一直认为， 动物细胞的分裂也有类似的分子机制。然而，细胞质分裂的变异，或机制的多样性， 和调节途径，对于多细胞生物体中的不同细胞类型变得越来越清楚。 这种细胞类型特异性的胞质分裂调控的机制仍然知之甚少。在 初步研究，我们建立了C.线虫4-细胞胚胎作为研究细胞类型特异性的系统 胞质分裂的调节。在4细胞阶段，每个细胞都有一个独特的细胞命运，由 保守的细胞命运信号传导途径（例如Src、Wnt/frizzled、Notch/Delta）和依赖于直接的细胞-细胞 胚胎内特定细胞对之间的接触。使用化学（LatrunculinA）或遗传（ 温度敏感突变体）扰动，以削弱F-肌动蛋白收缩环，我们确定了细胞类型- 4细胞胚胎中胞质分裂的特异性调节。我们发现四个细胞中有两个受到的保护 在F-actin细胞骨架被破坏的情况下， 变弱了胚胎显微解剖和细胞配对实验表明，在这两个保护的一个， 细胞（P2），细胞动力学保护是细胞内在的，并依赖于种系命运的规范，而在 其他受保护细胞（EMS），细胞动力学保护是细胞外调节的，需要直接细胞-细胞 与其邻近细胞（P2）接触和Src依赖性细胞命运信号传导。利用我们收集的ts突变体， 还确定了细胞类型特异性保护，防止纺锤体信号受损后胞质分裂失败， 在4-和8-细胞胚胎中的机器。本提案中的实验将确定分子 细胞命运特化以三种方式调节胞质分裂中细胞类型特异性变异的机制： 1)我们将确定细胞命运依赖性细胞动力学的细胞和分子机制 保护后肌动蛋白为基础的损害; 2）我们将比较细胞类型特异性保护细胞质分裂 在4 - 16细胞胚胎中，纺锤体信号损伤后的失败与肌动蛋白损伤后的失败相比较， 保守的和细胞类型特异性的机制，保护免受不同的细胞动力学压力;和3）我们将 确定细胞命运的规范是否也可以保护胞质分裂免受多细胞环境中的压力。 组织，以及测试是否适用于早期胚胎的原则也适用于哺乳动物细胞， 文化拟议的实验将确定细胞命运信号在促进基因组变异中的作用。 胞质分裂，确定特定的细胞命运途径，并测试其作用机制，并确定 胞质分裂中细胞类型特异性变异的普遍性。因为胞质分裂失败和细胞周期调节失调 细胞命运信号传导作为人类疾病如癌症的生物标志物而出现，并且由 进化上保守的分子机制，我们的工作将与人类健康有关。

PROJECT SUMMARY: In animal cells, cytokinesis is driven by constriction of an actomyosin contractile ring, which is positioned and controlled by signaling from spindle microtubules. It has long been assumed that all animal cells divide by a similar molecular mechanism. Yet variation in cytokinesis, or diversity in mechanistic and regulatory pathways, is becoming increasingly clear for different cell types within multicellular organisms. The mechanisms underlying this cell type-specific regulation of cytokinesis remain poorly understood. In preliminary studies, we established the C. elegans 4-cell embryo as a system to study cell type-specific regulation of cytokinesis. At the 4-cell stage, each individual cell already has a unique cell fate, specified by conserved cell fate signaling pathways (e.g. Src, Wnt/frizzled, Notch/Delta) and dependent on direct cell-cell contact between specific cell pairs within the embryo. Using either chemical (LatrunculinA) or genetic (formin temperature sensitive mutant) perturbations to weaken the F-actin contractile ring, we identified cell type- specific regulation of cytokinesis in the 4-cell embryo. We found that two of the four cells are more protected against cytokinetic stress than the others and can divide successfully when the F-actin cytoskeleton is weakened. Embryo micro-dissection and cell pairing experiments revealed that in one of these two protected cells (P2), cytokinetic protection is cell-intrinsic and dependent on germline fate specification, whereas in the other protected cell (EMS), cytokinetic protection is cell-extrinsically regulated and requires direct cell-cell contact with its neighbor cell (P2) and Src-dependent cell-fate signaling. Using our collection of ts mutants, we also identified cell type-specific protection against cytokinesis failure upon damage to the spindle signaling machinery in both the 4- and 8-cell embryo. The experiments in this proposal will determine the molecular mechanisms by which cell fate specification regulates cell type-specific variation in cytokinesis in three ways: 1) we will identify the cellular and molecular mechanisms that underlie cell fate-dependent cytokinetic protection after actin-based damage; 2) we will compare the cell type-specific protection against cytokinesis failure after spindle signaling-damage to that after actin-damage in the 4- thru 16-cell embryo and identify conserved and cell type-specific mechanisms that protect against different cytokinetic stresses; and 3) we will determine if cell fate specification can also protect cytokinesis from stress in the context of a multicellular tissue, as well as test whether the principles that apply to the early embryo also apply to mammalian cells in culture. The proposed experiments will define the role of cell fate signaling in promoting variation in cytokinesis, identify specific cell fate pathways and test their mechanisms of action, and determine the universality of cell type-specific variation in cytokinesis. Because both cytokinesis failure and dysregulation of cell fate signaling are both emerging as biomarkers for human diseases such as cancer and are regulated by evolutionarily conserved molecular mechanisms, our work will have relevance for human health.