Control of epithelial cell layer spreading in zebrafish

上皮细胞层铺展是各种发育和疾病相关事件中常见的和基本的过程，如果蝇的背部闭合和伤口愈合。在之前的资助期间，我们已经剖析了斑马鱼原肠发育过程中驱动上皮细胞层扩散的力量产生机制。具体地说，我们发现卵黄细胞上的包膜细胞层(EVL)是由肌动球蛋白皮质的收缩触发的，在卵黄合体细胞层(YSL)内形成了一个带状结构，YSL是卵黄细胞表面靠近EVL边缘的一个薄的细胞质层。我们可以进一步证明，该肌动球蛋白条带通过两种不同的运动活动--缆索收缩运动和流动摩擦运动--拉动EVL的边缘。缆索收缩马达通过肌动球蛋白带的周向收缩来运作，肌球蛋白带与卵黄囊的球形几何形状相连，拉动EVL边缘，就像钱包绳一样。流动-摩擦马达通过肌动球蛋白在YSL内逆行流动来工作。它们通过与邻近结构的摩擦来抵抗，从而直接将EVL边缘拉向植物极点。虽然这些发现为YSL肌动球蛋白带驱动EVL外胚层运动的机制提供了新的见解，但重要的问题仍然存在，如YSL内肌动球蛋白带的形成以及EVL和YSL之间的机械联系允许YSL拉动EVL。为了解决这些问题，我们已经开始分析EVL在其边缘如何与潜在的YSL建立联系，以及这种联系形成如何触发肌动球蛋白在YSL内的带状积累。根据我们的初步观察，我们假设YSL内的微管(MT)促进了YSL和EVL之间紧密连接(TJ)的形成，这些连接的正确形成反过来指导了EVL外胚层运动所需的YSL内的肌动球蛋白带组装。我们将使用来自发育生物学、细胞生物学、生物物理学和理论建模的方法和工具，通过跨学科的方法来解决这一假设。我们的目的是阐明在EVL细胞层扩散过程中MTS、TJS和肌动球蛋白网络之间的相互作用。我们期望这一方法将对上皮细胞层扩散过程中的力产生、传递和机制补偿机制提供深入的了解。

Epithelial cell layer spreading is a common and fundamental process in various developmental and disease-related events, such as dorsal closure in Drosophila and wound healing. In the previous funding period, we have dissected the force-generating mechanisms driving epithelial cell layer spreading during zebrafish gastrulation. Specifically, we showed that spreading of the enveloping cell layer (EVL) over the yolk cell during epiboly is triggered by contraction of the actomyosin cortex forming a band-like structure within the yolk syncytial layer (YSL), a thin cytoplasmic layer on the yolk cell surface close to the margin of the EVL. We could further show that this actomyosin band pulls on the margin of the EVL by two distinct motor activities - a cable-constriction motor and a flow-friction motor. The cable-constriction motor operates via circumferential contraction of the actomyosin band, which couples to the spherical geometry of the yolk sac to pull on the EVL margin, much like a purse string. The flow-friction motor operates via retrograde actomyosin flows within the YSL. These are resisted by friction to adjacent structures and thus directly pull the EVL margin towards the vegetal pole. While these findings provided novel insight into the mechanisms by which the YSL actomyosin band drives EVL epiboly movements, important questions remain as to the initiation of actomyosin band formation within the YSL and the mechanical linkage between EVL and YSL allowing the YSL to pull on the EVL. To address these questions, we have begun to analyze how the EVL at its margin establishes contacts with the underlying YSL, and how this contact formation triggers the band-like accumulation of actomyosin within the YSL. Based on our preliminary observations, we hypothesize that microtubules (MTs) within the YSL promote tight junction (TJ) formation between the YSL and EVL, and that proper formation of these junctions in turn directs actomyosin band assembly within the YSL required for EVL epiboly movements. We will address this hypothesis by an interdisciplinary approach employing methods and tools from developmental biology, cell biology, biophysics and theoretical modeling. We aim to elucidate the interplay between MTs, TJs and the actomyosin network during EVL cell layer spreading. We expect that this approach will provide insight into the mechanisms of force-generation, transduction and mechansensation during epithelial cell layer spreading.