a functional genomic yeast screen to identify pathogenic bacterial proteins功能基因酵母屏幕识别病原细菌的蛋白质.pdfVIP

A Functional Genomic Yeast Screen to Identify Pathogenic Bacterial Proteins 1 1 1 2 1* Naomi L. Slagowski , Roger W. Kramer , Monica F. Morrison , Joshua LaBaer , Cammie F. Lesser 1 Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Harvard Medical School, Cambridge, Massachusetts, United States of America, 2 Harvard Institute of Proteomics, Harvard Medical School, Cambridge, Massachusetts, United States of America, Many bacterial pathogens promote infection and cause disease by directly injecting into host cells proteins that manipulate eukaryotic cellular processes. Identification of these translocated proteins is essential to understanding pathogenesis. Yet, their identification remains limited. This, in part, is due to their general sequence uniqueness, which confounds homology-based identification by comparative genomic methods. In addition, their absence often does not result in phenotypes in virulence assays limiting functional genetic screens. Translocated proteins have been observed to confer toxic phenotypes when expressed in the yeast Saccharomyces cerevisiae. This observation suggests that yeast growth inhibition can be used as an indicator of protein translocation in functional genomic screens. However, limited information is available regarding the behavior of non-translocated proteins in yeast. We developed a semi-automated quantitative assay to monitor the growth of hundreds of yeast strains in parallel. We observed that expression of half of the 19 Shigella translocated proteins tested but almost none of the 20 non-translocated Shigella proteins nor ;1,000 Francisella tularensis proteins significantly inhibited yeast growth. Not only does this study establish that yeast growth inhibition is a sensitive and specific indicator of translocated proteins, b