Control of organ formation and morphogenesis during development

我的研究计划的长期目标是了解细胞群如何发育成专门的成人器官。这一过程需要调节生长、形态发生和分化。我的团队主要集中在果蝇的眼睛上，研究驱动组织规范的细胞信号的许多方面。现在我们把注意力转向细胞信号传导和基因表达调控发生后发生的事件。具体来说，我们对导致最终三维器官形成的机械和结构变化感兴趣。我们获得的见解可能广泛适用于理解未分化上皮如何发育成专门的成人器官。我的实验室使用特征良好、基因易于处理的果蝇作为模型生物，以了解眼睛形成过程中细胞事件是如何被调节的。成体组织在幼虫阶段在称为象盘的上皮细胞的离散囊中形成图案。所述成人眼由所述眼盘产生并包括有序的单位眼阵列。眼的发育开始于眼盘的后缘，并以一种称为形态发生沟（MF）的分化的虚拟“波”向前移动。MF被看作是一个狭窄的细胞带，顶端和顶基部都收缩，这使得它们在上皮中表现为物理沟槽。MF前面的细胞处于未分化的增殖状态，而MF后面的细胞则处于形态发生和分化状态。因此，MF代表细胞分裂和随后的视网膜分化之间的过渡点。MF的速度受到严格调节，以确保这两种相反的细胞状态之间的适当平衡。迄今为止，很少有研究涉及在MF进展过程中发生的深刻的细胞形状变化的作用，并且对形态发生如何影响分化知之甚少，尽管它们被认为是紧密相关的。我们将在未来5年通过研究以下短期目标来解决这些问题：确定Ste20激酶家族成员Misshapen （Msn）在调节MF进展中的作用和机制。最近我们发现了Ste20激酶Msn作为MF进展的负调节因子的新作用。msn的缺失导致细胞形态的改变。Ste20激酶具有不同的作用，我们认为Msn作为整合素的调节剂的假定作用是其调节MF的能力的核心。我们将通过免疫荧光、遗传相互作用研究和生化分析来确定Msn如何调节整合素的稳定性、定位和活性。此外，我们将使用类似的多方面方法研究Msn对微管、肌凝蛋白和肌动蛋白的调节。目的2：表征幼虫眼发育过程中细胞-细胞外基质（ECM）粘附相关成分的表达、定位和功能。迄今为止，我们对眼内MF进展的理解主要局限于顶细胞表面和细胞间粘附。我们的目的是确定细胞骨架参与者在基底细胞表面的作用，以及在这一动态过程中细胞- ecm粘附的相对贡献。鉴于在其他形态发生背景下粘附和细胞骨架重塑的关键作用，我们假设在MF进展中有类似但尚未被认识的贡献。我们将对眼形态发生过程中ECM和细胞骨架成分的表达和定位进行深入分析。我们将使用遗传分析来确定整合素亚基在募集ECM因子以及促进细胞分化方面的相关功能要求。

The long-term goal of my research program is to understand how groups of cells develop into specialized adult organs. Such a process requires regulation of growth, morphogenesis and differentiation. My group has focused primarily on the fruit fly eye to investigate numerous aspects of cell signaling that drive tissue specification. We now turn our attention to the events that occur after cell signaling and regulation of gene expression has occurred. Specifically, we are interested in the mechanical and structural changes that lead to the formation of the final three-dimensional organ. The insights we gain are likely to be broadly applicable to understanding how an undifferentiated epithelium develops into a specialized adult organ.My lab uses the well-characterized, genetically-tractable fruit fly Drosophila as a model organism to understand how cellular events are regulated during eye formation. Adult tissues are patterned during larval stages in discrete sacs of epithelial cells called imaginal discs. The adult eye arises from the eye disc and comprises an ordered array of unit eyes. Eye development initiates at the posterior edge of the disc and moves anteriorly in a virtual "wave" of differentiation known as the morphogenetic furrow (MF). The MF is seen as a narrow band of cells constricting both apically and apico-basally which causes them to appear as a physical groove in the epithelium. Cells anterior to the MF exist in an undifferentiated proliferative state, while those behind the MF undergo morphogenesis and differentiation. Thus the MF represents the transition point between cell division and subsequent retinal differentiation. The pace of the MF is tightly regulated to ensure the proper balance between these two opposing cellular states. To date, few studies have addressed the role of the profound cellular shape changes that occur during MF progression, and little is known about how morphogenesis influences differentiation, although they have been proposed to be tightly linked. We will address these issues through researching the following short-term objectives over the next 5 years:Objective 1. Determine the role and mechanism of the Ste20 kinase family member Misshapen (Msn) in regulating MF progression. Recently we found a novel role for the Ste20 kinase Msn as a negative regulator of MF progression. Loss of msn leads to altered cell morphology. Ste20 kinases have diverse roles, and we propose that Msn’s putative role as a regulator of integrins is central to its ability to regulate the MF. We will determine how Msn regulates integrin stability, localization and activity through immunofluorescence, genetic interactions studies and biochemical assays. Further, we will investigate regulation of microtubules, myosin and actin by Msn, using similar multi-facetted approaches.Objective 2: Characterize the expression, localization and function of cell-extracellular matrix (ECM) adhesion associated components during larval eye development. To date, our understanding of MF progression in the eye has been largely restricted to apical cell surfaces and to cell-cell adhesion. We aim to determine the role of cytoskeletal players at the basal cell surface and the relative contribution of cell-ECM adhesion during this dynamic process. Given the critical roles identified for adhesion and cytoskeletal remodeling in other morphogenetic contexts, we hypothesize a similar but as yet unrecognized contribution to MF progression. We will perform an in depth analysis of the expression and localization of ECM and cytoskeletal components during eye morphogenesis. We will use genetic analyses to determine the relative functional requirements for integrin subunits in recruiting ECM factors, as well as in promoting cell differentiation.