GAMETE MEMBRANE ADHESINO AND FUSION DURING FERTILIZATION

描述（由申请人提供）：我实验室的长期目标是了解受精过程中配子相互作用和细胞-细胞融合的细胞和分子机制。我们使用单细胞，双鞭毛衣原体为我们的研究，一个有机体，其受精是非常适合细胞生物学，生物化学，分子生物学和遗传学的方法。重要的是，在衣原体受精过程中，相互作用的雄性和雌性配子经历了几乎所有动物受精的细胞生物学事件。了解配子融合是必要的，以告知未来的研究对人类生殖。此外，配子融合对于昆虫传播由寄生原生动物引起的几种毁灭性人类疾病至关重要，包括引起疟疾和昏睡病的生物体。了解寄生原生动物中配子融合的分子机制将是减少它们引起的疾病影响的重要进展。不幸的是，尽管有许多不同的受精系统正在研究中，但我们对任何生物体中配子融合的分子理解仍然是基本的。该提案的目的是表征衣原体配子融合的细胞和分子机制，重点是两种配子融合蛋白，Fus 1，一种雌性配子特异性蛋白，和一种新的雄性配子特异性蛋白，我们在此资助期间发现，FusM。两者都位于融合部位，并且是融合所必需的。与迄今为止在任何生物体中鉴定的所有其他配子融合蛋白不同，FusM是广泛保守的。FusM家族成员虽然在两侧对称动物中没有发现，但存在于简单动物、高等植物和病原性原生动物中。在合作努力中使用靶向基因破坏伯氏疟原虫，啮齿动物疟疾寄生虫，我们发现，疟原虫FusM是配子融合所必需的。因此，我们对衣原体的研究揭示了配子融合的保守机制。现在，我们在每个衣原体配子上都有一种对配子融合至关重要的蛋白质，我们处于一个独特的位置，可以对配子融合的基本细胞和分子机制进行新的见解。我们建议确定Fus 1和FusM的结合伙伴，以确定配子膜粘附和融合的蛋白质的分子功能，调查其本地化的分子机制，并表征其分子特性之前，期间和之后的融合。

DESCRIPTION (provided by applicant): The long-term goals of my laboratory are to understand the cellular and molecular mechanisms that underlie gamete interactions and cell-cell fusion during fertilization. We use the unicellular, biflagellated alga Chlamydomonas for our studies, an organism whose fertilization is highly amenable to cell biological, biochemical, molecular biological and genetic approaches. Importantly, during Chlamydomonas fertilization, the interacting male and female gametes undergo the cell biological events that characterize fertilization in almost all animals. Understanding gamete fusion is needed to inform future studies on human reproduction. In addition, gamete fusion is essential for insect transmission of several devastating human diseases caused by parasitic protozoa, including the organisms that cause malaria and sleeping sickness. Knowing the molecular mechanisms of gamete fusion in parasitic protozoa would be an important advance in efforts to reduce the impact of the diseases they cause. Unfortunately, in spite of the many different fertilization systems being studied, our molecular understanding of gamete fusion in any organism remains rudimentary. The objectives of this proposal are to characterize the cellular and molecular mechanisms of gamete fusion in Chlamydomonas, focusing on two gamete fusion proteins, Fus1 a female gamete-specific protein, and a new male gamete-specific protein we discovered in this funding period, FusM. Both are localized at the sites of fusion and essential for fusion. Unlike all other gamete fusion proteins identified to date in any organism, FusM is widely conserved. Although not found in bilaterian animals, FusM family members are present in simple animals, higher plants, and pathogenic protozoa. In collaborative efforts using targeted gene disruption in Plasmodium berghei, a rodent malaria parasite, we discovered that the Plasmodium FusM is essential for gamete fusion. Thus, our studies on Chlamydomonas have revealed a conserved mechanism for gamete fusion. Now that we have a protein on each Chlamydomonas gamete that is essential for gamete fusion, we are in a unique position to make new insights into fundamental cellular and molecular mechanisms of gamete fusion. We propose to identify the binding partners of Fus1 and FusM, to determine the molecular functions of the proteins in gamete membrane adhesion and fusion, to investigate the molecular mechanisms of their localization, and to characterize their molecular properties before, during, and after fusion.