Molecular Basis of Developmental Cell Interactions

长期以来，人们已经认识到，发育领域中的细胞通过根据位置信息来组织生长和发育命运的变化，从而对位置的变化做出反应，位置信息是通过与邻近细胞的相互作用得出和解释的。了解这些相互作用的分子基础对于理解组织通过修复和再生复杂结构来应对无数侮辱的令人印象深刻的能力至关重要。优雅而有序的再生过程为图案形成的机制提供了线索，但也提供了希望，有一天这个过程可能会被操纵来替换受损的身体部分。为了操纵这一过程，有必要了解这一过程的遗传基础。在昆虫腿的例子中，我们正在接近这样的理解水平，所学到的许多经验都与脊椎动物系统有关。基因调控网络的动态网络似乎创建了一个强大的自组织系统，该系统既极其复杂，又可能简单，因为它依赖于几个关键的信号通路和几个简单的过程，如自激活和交叉抑制。这一应用的重点是在分子和组织水平上了解这些过程的机制和逻辑。本项目最后阶段的研究表明，WG和DPP这两种关键的形态因子通过抑制彼此的表达以及可能通过自动激活自己的表达而形成一个自组织系统。我们发现Wg对DPP表达的抑制是由dTcf直接介导的，并在WG调节区发现了介导其自身激活和DPP抑制的位点。在这里，我们建议继续我们的研究，以确定自激活和交叉抑制的机制是直接的还是间接的。我们建议确定不同dTcf亚型的生化特异性和功能，以更好地了解该蛋白在这些调节事件中的作用。我们将寻求确定一个假定的共同抑制者。初步实验表明，对沃尔伯特提到的位置对抗的反应达到了一个新的复杂水平。也就是说，不能对Wg产生反应的细胞仍然能够采用传统上认为的Wg依赖的细胞命运，尽管这只有在异位形态原的产生被阻止的情况下才被揭示出来。这增加了关于决定细胞命运和再生反应的替代机制的可能性，这些机制将被测试。最后，我们已经确定了一组影响视盘生长模式的突变体，这些突变体是参与这些活动的新候选基因的丰富集合。

It has long been recognized that cells in a developmental field respond to changes in position by organizing growth and changes in developmental fate according to positional information which is derived and interpreted via interactions with their neighbors". Understanding the molecular basis of those interactions is central to understanding the impressive ability of tissues to respond to a myriad of insults by repairing and regenerating complex structures. The elegant and orderly process of regeneration provides clues to the mechanisms of pattern formation but also offers the hope that the process might one day be manipulated to replace damaged body parts. To manipulate the process, it will be necessary to understand the genetic basis of the process. In the case of the insect leg, we are coming close to such a level of understanding and many of the lessons learned are relevant to vertebrate systems. A dynamic web of gene regulatory networks appears to create a robust self organizing system that is at once extremely intricate but also perhaps simple in its reliance on a few key signaling pathways and a few simple processes e.g. autoactivation and cross inhibition. The focus of this application is to understand the mechanisms and the logic of these processes at the molecular as well as tissue levels. Studies during the last period of this project reveal that two key morphogens, wg and dpp, form a self organizing system by suppressing each other's expression and perhaps by autoactivating their own expression. We have found that the repression of dpp expression by Wg is direct and mediated by dTcf and have identified sites in the wg regulatory region that mediate its autoactivation and repression by dpp. Here we propose to continue our studies to determine whether the mechanisms of autoactivation and cross inhibition are direct or indirect. We propose to determine the biochemical specificity and function of different dTcf isoforms to better understand the role of this protein in these regulatory events. We will seek to identify a putative co-repressor. Preliminary experiments suggest a new level of complexity in the response to positional confrontations alluded to by Wolpert. Namely, cells that are unable to respond to Wg are still able to adopt what are traditionally thought of as Wg dependent cell fates although this is revealed only if production of ectopic morphogens is blocked. This raises possibilities with respect to alternative mechanisms of cell fate determination and regeneration responses that will be tested. Finally, we have identified a collection of mutants affecting disc growth patterns that serve as an enriched collection of new candidate genes to participate in these events.