酵母是被归类为真菌的单细胞真核生物,主要在子囊门。在大约1500个命名物种中,酿酒酵母,也被称为面包酵母,在烹饪和酿造的背景下被人类驯化,是探索Wolbachia和原核生物新型效应蛋白功能的深刻遗传工具。Wolbachia是一种革兰氏阴性α-变形杆菌,可将高达75%的所有昆虫感染为专性细胞内微生物(JeyaprakashA,HoyMA,昆虫摩尔生物9:393-405,2000)。Wolbachia的生活方式为研究人员提出了独特的挑战。Wolbachia不能进行轴突培养,也从未进行过基因操纵。此外,许多Wolbachia基因在其他基因组中没有已知的功能或注释良好的直系同源物。然而考虑到Wolbachia对宿主表型的影响,在害虫防治方面有相当大的实际应用,它们无疑涉及与宿主基因产物相互作用的分泌效应蛋白。研究这些效应是Wolbachia目前遗传限制的挑战。然而,通过在酿酒酵母中表达候选蛋白可以克服与Wolbachia合作的一些限制。这种方法利用了酵母的小基因组(~6500个基因),典型的真核细胞组织,以及可用于其在文化中操纵的复杂遗传工具套件。因此,酵母可以作为强大的模拟真核宿主背景来研究Wolbachia效应子功能。具体来说,酵母用于重组蛋白表达,药物发现,蛋白质定位研究,蛋白质相互作用作图(酵母双杂交系统),模拟染色体进化,并在难以处理的原核系统中检查负责复杂表型的蛋白质之间的相互作用。作为一个例子,负责Wolbachia介导的细胞质不相容性(CI)的配对基因编码与其他已知蛋白质同源性有限的新型蛋白质,没有明显的功能。本文详细介绍了如何将酿酒酵母用作初始分期地,以探索Wolbachia商标表型(CI)之一的分子基础。
Yeasts are single-celled eukaryotic organisms classified as fungi, mostly in the phylum Ascomycota. Of about 1500 named species, Saccharomyces cerevisiae, also known as baker\'s yeast, domesticated by humans in the context of cooking and brewing, is a profound genetic tool for exploring functions of novel effector proteins from Wolbachia and prokaryotes in general. Wolbachia is a Gram-negative alpha-proteobacterium that infects up to ~75% of all insects as an obligate intracellular microbe (Jeyaprakash A, Hoy MA, Insect Mol Biol 9:393-405, 2000). Wolbachia\'s lifestyle presents unique challenges for researchers. Wolbachia cannot be axenically cultured and has never been genetically manipulated. Furthermore, many Wolbachia genes have no known function or well-annotated orthologs in other genomes. Yet given the effects of Wolbachia on host phenotypes, which have considerable practical applications for pest control, they undoubtedly involve secreted effector proteins that interact with host gene products. Studying these effectors is challenging with Wolbachia\'s current genetic limitations. However, some of the constraints to working with Wolbachia can be overcome by expressing candidate proteins in S. cerevisiae. This approach capitalizes on yeast\'s small genome (~6500 genes), typical eukaryotic cellular organization, and the sophisticated suite of genetic tools available for its manipulation in culture. Thus, yeast can serve as a powerful mock eukaryotic host background to study Wolbachia effector function. Specifically, yeast is used for recombinant protein expression, drug discovery, protein localization studies, protein interaction mapping (yeast two-hybrid system), modeling chromosomal evolution, and examining interactions between proteins responsible for complex phenotypes in less tractable prokaryotic systems. As an example, the paired genes responsible for Wolbachia-mediated cytoplasmic incompatibility (CI) encode novel proteins with limited homology to other known proteins, and no obvious function. This article details how S. cerevisiae was used as an initial staging ground to explore the molecular basis of one of Wolbachia\'s trademark phenotypes (CI).