Genetic Analysis of Retinal Cone Photoreceptor Function

光感受器是高度极化的、分隔的细胞。蛋白质合成始于内段 然后，运输沿着顶端方向向外段进行，或基本向突触进行。在……里面 近年来，几位研究人员在运输到心脏的机制(S)方面取得了开创性的发现。 外部管段。相比之下，人们对去往突触或突触的蛋白质的运输知之甚少。 在蛋白质的内部片段中进行分选，目的是送往不同的细胞隔间。肌醇磷脂是 已知是膜运输的关键调节因子，参与信号、规范和蛋白质 在各种各样的细胞类型中招募。细胞内磷脂酰肌醇的关键调节因子是脂质 磷酸酶，Synaptojanin I(SynJ1)，其主要细胞内靶标是PI(4，5)P2。我们已经确立了 斑马鱼作为评价肌醇磷脂信号转导和SynJ1功能的模型系统 视锥细胞光感受器的蛋白质分选过程。我们目前的工作发现SynJ1在内部细分市场中扮演了一个角色。 我们发现SynJ1集中在锥体内段，并且异常地聚集和/或 在缺乏这种蛋白质的NRC突变体中，高尔基体的结构被破坏。我们假设内侧部分 表型反映了去往突触的蛋白质的运输和分类的中断。我们提出了一个 一系列实验验证了这一假设，并准确定义了我们检测到的异常囊泡结构 在NRC内部片段中，它们的内容和派生。我们提案的具体预期结果是一个详细的 多聚肌醇磷脂平衡时NRC检出内节段缺陷的认识 由于关键的PI(4，5)P2磷酸酶SynJ1的丢失，光感受器被破坏。此外，我们的 研究将提供关于多磷肌醇在两种细胞中分布的基本信息 野生型和NRC视锥感受器。最后，我们的研究将确定不同结构的重要性 SynJ1的结构域。我们从本提案中概述的研究中发现的关键、基本信息 将有助于向许多其他重要的调查开放这一领域。

Photoreceptors are highly polarized, compartmentalized cells. Protein synthesis initiates in the inner segment and then transport proceeds in the apical direction toward the outer segment or basally toward the synapse. In recent years, several investigators have made seminal discoveries about the mechanism(s) of transport to the outer segment. In contrast, very little is known about the transport of proteins destined for the synapse or the sorting within the inner segment of proteins destined for different cellular compartments. Phosphoinositides are known to be key regulators in membrane trafficking and are involved in signaling, specification and protein recruitment in a wide variety of cell types. A key regulator of cellular phosphoinositides is the lipid phosphatase, synaptojanin I (SynJ1), whose primary intracellular target is PI(4,5)P2. We have established zebrafish as a model system in which to evaluate both phosphoinositide signaling and SynJ1 function in the process of protein sorting in cone photoreceptors. Our current work finds a role for SynJ1 in the inner segment. We find that SynJ1 concentrates in the cone inner segment and that large vesicles abnormally accrue and/or the Golgi architecture is disrupted in nrc mutants lacking this protein. We hypothesize that the inner segment phenotype reflects a disruption of transport and sorting of proteins destined for the synapse. We propose a series of experiments that test this hypothesis and define precisely the abnormal vesicular structures we detect in nrc inner segments, their content and derivation. The specific expected outcome of our proposal is a detailed understanding of the inner segment defect detected in nrc when the balance of polyphosphoinositides within the photoreceptor is disrupted due to the loss of the critical PI(4,5)P2 phosphatase, SynJ1. In addition, our studies will provide fundamental information about the cellular distribution of polyphosphoinositides in both wild-type and nrc cone photoreceptors. Finally, our studies will define the importance of different structural domains of SynJ1. The critical, fundamental information we discover from the studies outlined in this proposal will help open this field to many additional important investigations.