在大多数软体动物中,负责壳发育的早期组织,即,shell领域,显示了形态发生过程中内陷的常见过程。此外,证据表明,壳场内陷不是一个独立的事件,而是反映壳场形态发生整体状态的综合输出。然而,这一保守过程的潜在机制在很大程度上仍然未知.我们以前发现,肌动球蛋白网络(定期组织的丝状肌动蛋白(F-肌动蛋白)和肌球蛋白)可能通过揭示F-肌动蛋白在内陷区域的明显聚集并证明非肌肉肌动蛋白II(NMII)在腹足动物Lottiapeitaihoensis(=Lottiagoshimai)中内陷所必需。这里,我们调查了小GTPases的Rho家族(RhoA,Rac1,和Cdc42),以探索肌动球蛋白网络的上游调节剂。使用小分子抑制剂的功能测定表明,Cdc42调节壳场形态发生的关键事件,包括内陷和细胞重排,而RhoA和Rac1的作用可能是非特异性的或可忽略的。进一步的研究表明,Cdc42蛋白集中在壳场细胞的顶端,并与F-肌动蛋白聚集共定位。这两种分子的聚集可以通过用Cdc42抑制剂处理来防止。这些发现表明了壳场形态发生的可能的调节级联,其中Cdc42在壳场细胞的顶端侧招募F-肌动蛋白(肌动球蛋白网络),然后产生合成的机械力,介导正确的壳场形态发生(细胞形状变化,内陷和细胞重排)。我们的结果强调了细胞骨架在早期壳发育中的作用,并为软体动物壳进化提供了新的见解。
In most mollusks (conchiferans), the early tissue responsible for shell development, namely, the shell field, shows a common process of invagination during morphogenesis. Moreover, lines of evidence indicated that shell field invagination is not an independent event, but an integrated output reflecting the overall state of shell field morphogenesis. Nevertheless, the underlying mechanisms of this conserved process remain largely unknown. We previously found that actomyosin networks (regularly organized filamentous actin (F-actin) and myosin) may play essential roles in this process by revealing the evident aggregation of F-actin in the invaginated region and demonstrating that nonmuscle myosin II (NM II) is required for invagination in the gastropod Lottia peitaihoensis (= Lottia goshimai). Here, we investigated the roles of the Rho family of small GTPases (RhoA, Rac1, and Cdc42) to explore the upstream regulators of actomyosin networks. Functional assays using small molecule inhibitors suggested that Cdc42 modulates key events of shell field morphogenesis, including invagination and cell rearrangements, while the roles of RhoA and Rac1 may be nonspecific or negligible. Further investigations revealed that the Cdc42 protein was concentrated on the apical side of shell field cells and colocalized with F-actin aggregation. The aggregation of these two molecules could be prevented by treatment with Cdc42 inhibitors. These findings suggest a possible regulatory cascade of shell field morphogenesis in which Cdc42 recruits F-actin (actomyosin networks) on the apical side of shell field cells, which then generates resultant mechanical forces that mediate correct shell field morphogenesis (cell shape changes, invagination and cell rearrangement). Our results emphasize the roles of the cytoskeleton in early shell development and provide new insights into molluscan shell evolution.