Cell surface membrane proteins perform diverse and critical functions and are spatially and temporally regulated by membrane trafficking pathways. Although perturbations in these pathways underlie many pathologies, our understanding of these pathways at a mechanistic level remains incomplete. Using yeast as a model, we have developed an assay that reports on the surface activity of the uracil permease Fur4 in uracil auxotroph strains grown in the presence of limited uracil. This assay was used to screen a library of haploid deletion strains and identified mutants with both diminished and enhanced comparative growth in restricted uracil media. Factors identified, including various multisubunit complexes, were enriched for membrane trafficking and transcriptional functions, in addition to various uncharacterized genes. Bioinformatic analysis of expression profiles from many strains lacking transcription factors required for efficient uracil-scavenging validated particular hits from the screen, in addition to implicating essential genes not tested in the screen. Finally, we performed a secondary mating factor secretion screen to functionally categorize factors implicated in uracil-scavenging.

中文翻译：

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Fur4 介导的尿嘧啶清除以筛选表面蛋白调节剂

细胞表面膜蛋白执行多种关键功能，并在空间和时间上受膜运输途径的调节。尽管这些途径中的扰动是许多病理的基础，但我们在机械水平上对这些途径的理解仍然不完整。使用酵母作为模型，我们开发了一种检测方法，报告在有限尿嘧啶存在的情况下生长的尿嘧啶营养缺陷型菌株中尿嘧啶通透酶 Fur4 的表面活性。该测定法用于筛选单倍体缺失菌株库，并鉴定在限制性尿嘧啶培养基中相对生长减少和增强的突变体。除了各种未鉴定的基因外，已鉴定的因子，包括各种多亚基复合物，都富含膜运输和转录功能。对缺乏有效清除尿嘧啶所需转录因子的许多菌株的表达谱进行生物信息学分析验证了筛选中的特定命中，此外还暗示了筛选中未测试的必需基因。最后，我们进行了二次交配因子分泌筛选，以对与尿嘧啶清除有关的因子进行功能分类。