PDF(Pubmed)
Abstract：
Fission yeast, a single-cell eukaryotic organism, shares many fundamental cellular processes with higher eukaryotes, including gene transcription and regulation, cell cycle regulation, vesicular transport and membrane trafficking, and cell death resulting from the cellular stress response. As a result, fission yeast has proven to be a versatile model organism for studying human physiology and diseases such as cell cycle dysregulation and cancer, as well as autophagy and neurodegenerative diseases like Alzheimer\'s, Parkinson\'s, and Huntington\'s diseases. Given that viruses are obligate intracellular parasites that rely on host cellular machinery to replicate and produce, fission yeast could serve as a surrogate to identify viral proteins that affect host cellular processes. This approach could facilitate the study of virus-host interactions and help identify potential viral targets for antiviral therapy. Using fission yeast for functional characterization of viral genomes offers several advantages, including a well-characterized and haploid genome, robustness, cost-effectiveness, ease of maintenance, and rapid doubling time. Therefore, fission yeast emerges as a valuable surrogate system for rapid and comprehensive functional characterization of viral proteins, aiding in the identification of therapeutic antiviral targets or viral proteins that impact highly conserved host cellular functions with significant virologic implications. Importantly, this approach has a proven track record of success in studying various human and plant viruses. In this protocol, we present a streamlined and scalable molecular cloning strategy tailored for genome-wide and comprehensive functional characterization of viral proteins in fission yeast.
摘要：
裂变酵母,单细胞真核生物，与高等真核生物共享许多基本的细胞过程，包括基因转录和调控，细胞周期调节，囊泡运输和膜运输，和细胞应激反应导致的细胞死亡。因此,裂变酵母已被证明是研究人体生理和疾病，如细胞周期失调和癌症的多功能模型生物，以及自噬和神经退行性疾病，如阿尔茨海默氏症，帕金森,和亨廷顿病。鉴于病毒是专性的细胞内寄生虫，它们依赖于宿主细胞机制来复制和产生，裂殖酵母可以作为鉴定影响宿主细胞过程的病毒蛋白的替代品。这种方法可以促进病毒与宿主相互作用的研究，并有助于确定抗病毒治疗的潜在病毒靶标。使用裂殖酵母进行病毒基因组的功能表征提供了几个优点，包括一个特征明确的单倍体基因组，鲁棒性，成本效益，易于维护,快速倍增时间。因此,裂殖酵母是一种有价值的替代系统，用于快速和全面的病毒蛋白功能表征，有助于鉴定影响高度保守的宿主细胞功能的治疗性抗病毒靶标或病毒蛋白，具有显著的病毒学意义。重要的是,这种方法在研究各种人类和植物病毒方面具有成功的记录。在这个协议中,我们提出了一种简化和可扩展的分子克隆策略，专门用于裂变酵母中病毒蛋白的全基因组和全面功能表征。
