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A Genome-wide Drosophila RNAi Screen for Regulators of Centrosome Reduction

果蝇全基因组 RNAi 筛选中心体减少的调节因子

基本信息

批准号：
9317290
负责人：
Tomer Avidor-Reiss
金额：
$ 7.38万
依托单位：
UNIVERSITY OF TOLEDO
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-07-18 至 2019-06-30
项目状态：
已结题

项目摘要

Summary During sperm formation, the quantity of many centrosomal proteins declines in a process known as centrosome reduction. However, the role and importance of this reduction in sperm differentiation is poorly understood. In fact, centrosome reduction is often discounted on the assumption that it is inconsequential for male fertility. However, this dogma has not been rigorously tested and our data argues differently. Therefore, the demonstration that centrosome reduction is mediated by specific pathways and is essential for post- fertilization embryo development has the potential to transform the field. Consequently, the long-term goal of the proposed research is to reveal both the biochemical pathways underlying centrosome reduction during sperm maturation and the potential involvement of centrosome reduction in male fertility. The objective of this application is to perform a pilot study in Drosophila melanogaster that will generate tools essential for a detailed follow-up investigation of centrosome reduction. Drosophila is an excellent genetic model to begin these studies because, like in humans and many other animals, specific centrosomal proteins are reduced at distinct steps during sperm differentiation, and these modified sperm centrosomes are critical for zygotic function after fertilization. Indeed, data from the Avidor- Reiss lab indicates that a specific, kinase-dependent pathway is essential for the reduction of one conserved centrosomal protein, but not others, and that blocking this reduction interferes with post- fertilization development. Therefore, our central hypothesis is that decreases in specific centrosomal proteins during centrosome reduction are essential for the distinct role of the zygotic centrosome in post- fertilization embryo development. Our rationale is that the identification of key genes/proteins involved in Drosophila centrosome reduction will allow us to better define the mechanism and function of centrosome reduction in sperm maturation. Our specific aim is to identify genes that regulate the reduction of specific centrosomal proteins during spermiogenesis. To achieve this aim, we will perform a visual RNAi screen targeting kinases and phosphatases that will cause abnormal localization of fluorescently-tagged centrosomal proteins in the spermatozoa. This study is innovative because it is the first to use a genetic approach and genetically-tagged markers to study centrosome reduction. This study is expected to vertically advance the understanding of how and why centrosomal proteins decrease during sperm formation. Ultimately, knowledge gained from this basic research has the potential to inspire research for new causes of male infertility, early stage miscarriages, and developmental diseases.

总结在精子形成过程中，许多中心体蛋白质的数量在一个称为中心体减少然而，这种减少在精子分化中的作用和重要性很差，明白事实上，中心体减少常常被假设是无关紧要的，男性生育能力然而，这一教条并没有经过严格的检验，我们的数据也有不同的说法。因此，我们认为，证明中心体减少是由特定的途径介导的，并且对后受精胚胎发育有可能改变这一领域。因此，长期目标是这项研究的目的是为了揭示在细胞分裂过程中中心体减少的生化途径。精子成熟和中心体减少在男性生育力中的潜在参与。的目的应用程序是在果蝇中进行试点研究，这将产生对对中心体减少进行详细的随访研究。果蝇是开始这些研究的一个极好的遗传模型，因为，像人类和许多其他动物一样，在动物中，在精子分化过程中，特定的中心体蛋白在不同的步骤中减少，修饰的精子中心体对受精后的合子功能至关重要。事实上，Avidor的数据- Reiss实验室指出，一种特异性的激酶依赖性途径对于减少一种保守的中心体蛋白，而不是其他的，并阻止这种减少干扰后，受精发育因此，我们的中心假设是，在中心体减少过程中的蛋白质对于合子中心体在后受精胚胎发育我们的基本原理是，识别参与果蝇中心体的缩小将使我们能够更好地定义中心体的机制和功能减少精子成熟。我们的具体目标是确定基因，调节特定的减少，精子发生过程中的中心体蛋白。为了实现这一目标，我们将进行一个可视化的RNAi屏幕，靶向激酶和磷酸酶，其将导致荧光标记的中心体异常定位精子中的蛋白质这项研究是创新的，因为它是第一个使用遗传方法，基因标记来研究中心体减少。这项研究有望纵向推进了解中心体蛋白在精子形成过程中如何以及为什么减少。最终，知识从这项基础研究中获得的信息有可能激发对男性不育症新原因的研究，分期流产和发育性疾病