Ad5 Fiber Entry and Trafficking in Lacrimal Acini

Ad5 纤维在泪腺泡中的进入和运输

• 批准号: 7075776
• 负责人: Sarah F Hamm-Alvarez
• 金额: $ 39.14万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-03 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7075776
• 关键词: Adenoviridae; acinar cell; proteins; receptor

DESCRIPTION: A major contribution to ocular morbidity is lacrimal dysfunction, affecting over 10 million Americans. The principal cell of the lacrimal gland and primary contributor of proteins into ocular surface fluid is the lacrimal acinar cell, which is the target of much of the ocular research into the etiology of dry eye diseases including the severe autoimmune disease, Sjogren's syndrome. Ongoing studies are now shedding insights into the precise mechanisms involved in initiation, development and progression of disease, suggesting that identification of prospective therapeutic targets is likely in the not so distant future. However, we are still very limited in our ability to specifically target the next generation of macromolecular drugs, particularly DNA-, protein- or peptide based drugs, to the lacrimal gland, raising the possibility that we may soon identify advanced therapies for treatment of severe dry eye diseases but will be unable to deliver these drugs to the target site. Our focus here is to explore the unusual and possibly unique uptake mechanism utilized for lacrimal acinar internalization of adenovirus serotype 5 (Ad5). We have demonstrated in lacrimal acinar cells that Ad5 utilizes a unique fiber-dependent internalization pathway, in contrast to the penton-dependent internalization described in other systems. We hypothesize that Ad5 may use multiple fiber receptors for binding and entry in lacrimal acini, and further that one or more of these entry pathways is either unusually robust or unique to lacrimal acini, explaining the unusual fiber-dependence of entry in tandem with the high efficiency of viral transduction in lacrimal acini. We propose to characterize the participants in this novel fiber-dependent internalization pathway at the molecular level, with a particular focus on coxsackievirus adenovirus receptor, major histocompatibility complex class 1 and heparin sulfate-glycosaminoglycan receptors, and their different modes of endocytosis. The aims are as follows: Aim #1. Does coxsackievirus adenovirus receptor mediate fiber-dependent Ad5 internalization or free fiber internalization in lacrimal acini? Aim #2. What other receptors participate in fiber-dependent Ad5 internalization or free fiber internalization in lacrimal acini? Aim #3. What intracellular trafficking pathways mediate fiber-dependent Ad5 internalization or free fiber internalization in lacrimal acini? At the conclusion of this work, we will have elucidated the contributions of specific receptors and endocytic internalization pathways responsible for fiber-dependent internalization of Ad5. We will have determined to what extent the recombinant fiber or knob, the terminal region of the fiber protein, can recapitulate these pathways. Finally, we will have tested proof-of-principle experiments to determine whether fiber or knob can facilitate entry of antisense oligonucleotides and proteins into lacrimal acinar cytosol.

描述：对眼部发病率的主要贡献是泪功能障碍，影响了超过1000万美国人。泪腺的主要细胞和蛋白质对眼部表面液的主要因素是泪腺腺泡细胞，这是对干眼症病因的大部分眼科研究的靶标，包括严重的自身免疫性疾病，包括严重的自身免疫性疾病，Sjogren的综合征。现在，正在进行的研究正在洞悉疾病起始，发展和进展所涉及的确切机制，这表明在不久的将来可能会识别前瞻性治疗靶标。但是，我们仍然非常有限地靶向下一代大分子药物，尤其是基于DNA，蛋白质或肽的药物，以靶向泪腺，从而提高了我们很快可能确定严重治疗的晚期治疗的可能性。干眼症，但将无法将这些药物输送到目标部位。我们在这里的重点是探索用于腺病毒血清型5（AD5）的泪腺内在化的异常且可能独特的摄取机制。与其他系统中描述的penton依赖性内部化相反，我们已经在泪腺腺泡细胞中证明了AD5使用了独特的纤维依赖性内部化途径。我们假设AD5可以使用多个纤维受体来结合和进入泪acini，进一步说，这些进入途径中的一个或多个是异常健壮的，或者是富富富富的独特的，这解释了与高的纤维依赖性依赖于高高的纤维依赖性。泪液中病毒转导的效率。我们建议在分子水平的这种新型纤维依赖性内部化途径中表征参与者，特别关注Coxsackievivirus腺病毒受体，主要的组织兼容性复合物1类和硫酸硫酸硫酸盐 - 糖胺聚糖受体，以及它们不同的内细胞增多症模式。目的如下：目标＃1。 Coxsackievivirus腺病毒受体是否介导了纤维依赖的AD5内在化或泪acini中的游离纤维内在化？目标＃2。其他哪些受体参与纤维依赖性的AD5内在化或泪acini中的游离纤维内在化？目标＃3。哪种细胞内运输途径介导纤维依赖的AD5内在化或泪acini中的游离纤维内在化？在这项工作结束时，我们将阐明特定受体和负责AD5纤维依赖性内在化的内吞内部化途径的贡献。我们将确定重组纤维或旋钮（纤维蛋白的末端区域）在多大程度上可以概括这些途径。最后，我们将测试原则实验实验，以确定纤维或旋钮是否可以促进反义寡核苷酸和蛋白质进入富富腺腺泡细胞质。