Bacteria have evolved complex strategies to compete and communicate with one another. A new mechanism of interbacterial competition, termed contact-dependent growth inhibition (CDI) was recently discovered in Escherichia coli. CDI systems are found in a wide variety of gram-negative bacteria, including many important human pathogens. CDI is mediated by the CdiB/CdiA two-partner secretion system. CdiB is a predicted outer membrane protein that is required for the export and assembly of the CdiA exoprotein onto the cell surface. The C-terminal region of CdiA (CdiA-CT) contains the growth inhibition activity and is presumably cleaved and translocated into target cells to inhibit growth. CDI systems also encode CdiI immunity proteins, which bind and inactivate their cognate CdiA-CT toxins, thereby protecting CDI+ cells from autoinhibition. Remarkably, the CdiA-CT toxin domains are polymorphic, and accordingly their corresponding CdiI immunity proteins are also highly variable. We have identified at least 30 distinct toxin-immunity families to date. There is typically less than 20% amino acid sequence identity between different CdiA-CT/CdiI families, strongly suggesting that the protein-protein interactions underlying each CdiA-CT/CdiI complex are unique. Moreover, we have recently discovered that some CdiA-CT domains interact with specific target cell proteins termed """"""""permissive factors"""""""", and these binding interactions are required to activate the delivered toxins. Currently, the mechanisms by which CdiI proteins neutralize and permissive factors activate CdiA-CT toxins are not understood. We propose structural, biochemical and genetic analyses to gain insights into the intricate toxin-immunity network encoded by bacterial CDI systems. In this proposal, we challenge the PSI Network center to assist in solving the crystal structures of at least ten distinct CdiA-CT/CdiI complexes. This proposal represents a unique opportunity to elucidate how specific binding is maintained as toxin-immunity pairs diverge through evolution.

Public Health Relevance

Bacteria have evolved complex strategies to compete and communicate with one another. A new mechanism of interbacterial competition, termed contact-dependent growth inhibition (CDI) has recently been discovered in a wide variety of gram-negative bacterial pathogens. This proposal utilizes structural, biochemical and genetic analyses to gain mechanistic insights into the intricate toxin-immunity network encoded by CDI systems. This research will significantly increase our understanding of the evolution and ecology of bacterial pathogens and could inform the development of novel antimicrobial therapies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project--Cooperative Agreements (U01)
Project #
1U01GM102318-01
Application #
8350577
Study Section
Special Emphasis Panel (ZGM1-CBB-0 (BP))
Program Officer
Ainsztein, Alexandra M
Project Start
2012-09-01
Project End
2015-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
1
Fiscal Year
2012
Total Cost
$595,565
Indirect Cost
$105,199
Name
University of California Irvine
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
046705849
City
Irvine
State
CA
Country
United States
Zip Code
92697
Johnson, Parker M; Beck, Christina M; Morse, Robert P et al. (2016) Unraveling the essential role of CysK in CDI toxin activation. Proc Natl Acad Sci U S A 113:9792-7
Johnson, Parker M; Gucinski, Grant C; Garza-Sánchez, Fernando et al. (2016) Functional Diversity of Cytotoxic tRNase/Immunity Protein Complexes from Burkholderia pseudomallei. J Biol Chem 291:19387-400
Willett, Julia L E; Gucinski, Grant C; Fatherree, Jackson P et al. (2015) Contact-dependent growth inhibition toxins exploit multiple independent cell-entry pathways. Proc Natl Acad Sci U S A 112:11341-6
Willett, Julia L E; Ruhe, Zachary C; Goulding, Celia W et al. (2015) Contact-Dependent Growth Inhibition (CDI) and CdiB/CdiA Two-Partner Secretion Proteins. J Mol Biol 427:3754-65
Ruhe, Zachary C; Townsley, Loni; Wallace, Adam B et al. (2015) CdiA promotes receptor-independent intercellular adhesion. Mol Microbiol 98:175-92
Ruhe, Zachary C; Hayes, Christopher S; Low, David A (2015) Measuring Cell-Cell Binding Using Flow-Cytometry. Methods Mol Biol 1329:127-36
Tan, Kemin; Johnson, Parker M; Stols, Lucy et al. (2015) The structure of a contact-dependent growth-inhibition (CDI) immunity protein from Neisseria meningitidis MC58. Acta Crystallogr F Struct Biol Commun 71:702-9
Campanini, Barbara; Benoni, Roberto; Bettati, Stefano et al. (2015) Moonlighting O-acetylserine sulfhydrylase: New functions for an old protein. Biochim Biophys Acta 1854:1184-93
Morse, Robert P; Willett, Julia L E; Johnson, Parker M et al. (2015) Diversification of β-Augmentation Interactions between CDI Toxin/Immunity Proteins. J Mol Biol 427:3766-84
Whitney, John C; Beck, Christina M; Goo, Young Ah et al. (2014) Genetically distinct pathways guide effector export through the type VI secretion system. Mol Microbiol 92:529-42

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