The genomes of pathogenic bacteria are being rapidly sequenced. A major challenge is to devise efficient, sensitive, and specific assays to screen bacterial genomes to identify pathogenic proteins and determine their specific roles in pathogenesis. Although yeast cannot serve as a physiologic model of human infection, our laboratory and others have recently established Saccharomyces cerevisiae as a powerful model system to study bacterial proteins that target potentially conserved eukaryotic host cell processes. Preliminary evidence presented in this proposal demonstrates that toxic yeast phenotypes conferred by Shigella proteins are a sensitive and specific screen for proteins that target host cell processes. Given its relatively small genome, genetic tractability, well- developed post-genomic tools, conservation of many basic cellular processes, and the wealth of available systematic data S. cerevisiae is an ideal model organism for multidisciplinary systems- biology studies. In response to PA-02-011, """"""""Bioengineering Research Grants,"""""""" we propose to develop and validate a multidisciplinary, integrative, systems approach in yeast involving genomics, proteomics, cell biology and novel bioinformatics software development to identify host cell processes targeted by three Shigella proteins, IpgB, OspCI and OspF. Evidence suggests that each of these proteins is delivered directly into host cells during infection, but little is known about their functions within. We hypothesize that the genome-wide screens described in this proposal will result in the characterization of the molecular roles in pathogenesis of each of these proteins. Experiments in this proposal focus on proteins from the genetically manipulable Shigella so that we can relatively easily test hypotheses in physiologic models of disease. However, once optimized, this multidisciplinary approach should be applicable to study any microbial pathogen that targets intracellular host cell processes, especially pathogens that are dangerous to grow or difficult to genetically manipulate like Mycobacterium and Chlamydia. This work is important is relevant to public health issues since by investigations mechanisms that bacterial pathogens used to cause disease, we will gain information that will help develop new antimicrobials to treat these infections. Furthermore, we are interested in developing a new and efficient way to study bacterial pathogens that are dangerous and difficult to study in other ways.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI064285-05
Application #
7771772
Study Section
Host Interactions with Bacterial Pathogens Study Section (HIBP)
Program Officer
Mills, Melody
Project Start
2006-03-01
Project End
2012-02-28
Budget Start
2010-03-01
Budget End
2012-02-28
Support Year
5
Fiscal Year
2010
Total Cost
$371,317
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Ernst, Nadja Heinz; Reeves, Analise Z; Ramseyer, Julia E et al. (2018) High-Throughput Screening of Type III Secretion Determinants Reveals a Major Chaperone-Independent Pathway. MBio 9:
Mou, Xiangyu; Souter, Skye; Du, Juan et al. (2018) Synthetic bottom-up approach reveals the complex interplay of Shigella effectors in regulation of epithelial cell death. Proc Natl Acad Sci U S A 115:6452-6457
Lynch, Jason P; Lesser, Cammie F (2018) Host-Pathogen Interactions: What the EHEC Are We Learning from Host Genome-Wide Screens? MBio 9:
González-Prieto, Coral; Lesser, Cammie F (2018) Rationale redesign of type III secretion systems: toward the development of non-pathogenic E. coli for in vivo delivery of therapeutic payloads. Curr Opin Microbiol 41:1-7
Janakiraman, Anuradha; Lesser, Cammie F (2017) How to manage stress: Lessons from an intracellular pathogen. Virulence 8:359-361
Piro, Anthony S; Hernandez, Dulcemaria; Luoma, Sarah et al. (2017) Detection of Cytosolic Shigella flexneri via a C-Terminal Triple-Arginine Motif of GBP1 Inhibits Actin-Based Motility. MBio 8:
Du, Juan; Reeves, Analise Z; Klein, Jessica A et al. (2016) The type III secretion system apparatus determines the intracellular niche of bacterial pathogens. Proc Natl Acad Sci U S A 113:4794-9
Sheehan, Kathy B; Martin, MaryAnn; Lesser, Cammie F et al. (2016) Identification and Characterization of a Candidate Wolbachia pipientis Type IV Effector That Interacts with the Actin Cytoskeleton. MBio 7:
Reeves, Analise Z; Lesser, Cammie F (2016) Transfer of Large Contiguous DNA Fragments onto a Low Copy Plasmid or into the Bacterial Chromosome. Bio Protoc 6:
Reeves, Analise Z; Spears, William E; Du, Juan et al. (2015) Engineering Escherichia coli into a protein delivery system for mammalian cells. ACS Synth Biol 4:644-54

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