Mechanisms of photoreceptor disc maturation

这项提案中描述的实验解决了视觉中最令人着迷的未解答问题之一 光感受器外节形态发生的分子机制研究。这个 外节是一个纤毛细胞器，它响应捕捉光而产生电信号。独一无二的 外节的形态特征是，它充满了一堆扁平的膜盘，提供了巨大的 用于光子捕获和信号放大的表面。这种解剖结构的功能意义在于 已经被认识了很长一段时间，但我们对盘是如何在分子水平上构建的理解仍然存在 令人沮丧的简陋。本申请涉及光感受器盘的几个鲜为人知的方面 形态发生，与椎间盘扩张、排列和包绕的过程有关。我们的初步数据显示 新形成的圆盘的边缘包含两种不同类型的细胞外连接：一种将圆盘与 内段质膜与另一个相互连接的圆盘边缘。所描述的实验 在目标1和目标2中，将致力于确定每种连接类型的蛋白质组成并阐明其 在支持光盘伸长和堆叠的高保真度方面具有特定的作用。目标3将专注于DISC的最后一步 从外节质膜上脱落的成熟过程。我们将解决DISC是否 分裂只发生在杆状结构中，并探索参与这一过程的分子成员。实验 在本申请中描述的将使用多功能分子工具与最先进的三种 外段结构的三维电子显微镜断层扫描分析。解决这些问题 机制问题是促进我们对感光细胞生物学的基本理解的关键，以及 阐明遗传性失明的病理生理机制 外节形态发生。

The experiments described in this proposal address one of the most fascinating unanswered questions in vision research regarding the molecular mechanisms responsible for photoreceptor outer segment morphogenesis. The outer segment is a ciliary organelle that produces electrical signals in response to capturing light. A unique morphological feature of the outer segment is that it is filled with a stack of flattened membrane discs providing vast surfaces for photon capture and signal amplification. The functional significance of this anatomical arrangement has been recognized for a very long time, yet our understanding of how discs are built at the molecular level remains frustratingly rudimentary. This application addresses several poorly understood aspects of photoreceptor disc morphogenesis, related to the processes of disc expansion, alignment and enclosure. Our preliminary data show that the edges of newly formed discs contain two distinct types of extracellular links: one connecting discs with the inner segment plasma membrane and another connecting disc edges between themselves. Experiments described in Aims 1 and 2 will be devoted to determining the protein composition of each link type and elucidating their specific roles in supporting the high fidelity of disc elongation and stacking. Aim 3 will focus on the final step in disc maturation consisting of its scission from the outer segment plasma membrane. We will address whether disc scission takes place exclusively in rods and explore molecular players involved in this process. Experiments described in this application will employ versatile molecular tools combined with the state-of-the-art three dimensional electron microscopy tomographic analysis of the outer segment structure. Addressing these mechanistic questions is essential for advancing our basic understanding of photoreceptor cell biology, as well as elucidating the pathophysiological mechanisms underlying inherited blindness frequently associated with defects in outer segment morphogenesis.