Department of Biology

Program

Microbiology

Research Area

Microbial Interactions and Pathogenesis

Education

Ph.D., University of Texas at Dallas, 2002
Postdoctoral Fellow, University of Chicago, 2003-2006

Research Description

My research focuses on understanding host-microbe interactions, specifically as related to plague pathogenesis.  Yersinia pestis, the causative agent of plague, employs multiple virulence strategies to establish an infection.  One well-studied mechanism is the type III secretion system, effectively a molecular syringe that allows the bacterium to inject cytotoxic proteins (known as Yops) directly into the cytoplasm of target host cells.  Type III secretion is essential for virulence, and null mutants display a seven-log increase in the lethal dose for a mouse model of infection.  Our research is therefore aimed at understanding the molecular mechanisms that promote type III secretion and regulate this pathway during infection.
           
Regulation of the Type III Secretion System:
Y. pestis encounters both mammalian hosts and insect vectors during its life cycle.  Each host presents a different set of environmental conditions, and the expression of a large number of genes having basic metabolic as well as pathogenic functions is regulated accordingly.  Signals such as temperature, calcium and serum protein and amino acids serve to regulate Yop delivery until contact with an appropriate target cell has been made.  Several regulatory proteins that respond to these signals have been identified (such as LcrF, YopN, LcrV, LcrG, LcrQ, YopB, and YopD, and there respective chaperones).  However the gene products responsible for sensing the environmental cues and then relaying that information to the known regulatory components of the secretion machinery remain a mystery.  We therefore hope to identify the sensors and signal transduction pathways involved in controlling type III secretion.  Our work has revealed a two component system, which in other organisms such as Salmonella, Pseudomonas, and E. coli, functions as part of an elaborate signal transduction network.  Several uncharacterized genes have also been linked to the type III pathway.  Future studies will address the mechanism by which these gene products contribute to the processing of environmental cues and regulation of type III secretion.

Target Cell Selection:
Bacteria are faced with a wide array of cells when entering a mammalian host; however most bacteria will only colonize specific tissues.  Plague bacteria and other Yersinia species share a common tropism for lymphoid tissues (liver, spleen, lymph nodes).  In vitro, however, many cell types including epithelial, hepatic, and macrophage cell lines can all be targets for injection by the type III secretion system.  How then is tissue tropism determined?  To identify the cells targeted by yersiniae, a fluorescent reporter assay was developed to visualized injected cells.  In this assay, a Yop protein is fused to beta-lactamase to generate a hybrid protein which can be injected by the type III pathway.  Host cells that have been injected can be distinguished from uninjected cells based on a shift from green to blue fluorescence upon addition of a fluorescent beta-lactamase substrate.  With this technology, we showed that plague bacilli are able to specifically target cells of the innate immune system (neutrophils, macrophages, dendritic cells) in the spleens of infected mice.  While this helps to explain tissue tropism, it also opens the door to other questions: 1) Are the same type of cells targeted in different organs?  2) Do different routes of infection influence the selection process?  3) Are all Yop proteins delivered with the same efficiency into all targeted cell types?   4) How is the selection process mediated; i.e., what are the bacterial and host genes involved in target cell identification?

Regulatory Mechanism of YopQ:
YopQ, a substrate of the type III secretion system, has long been a mystery.  While yopQ mutants inject greater quantities of other Yops into tissue culture cells, they display a severe virulence defect in mice.  Furthermore, the location of YopQ during infection was unknown.  Using the beta-lactamase reporter assay described above, we found YopQ injected into host cells.  Bioinformatic analysis reveals only a putative CaaX box, which in eukaryotes is a signal for prenyl modification.  We will determine whether this CaaX box is functional and if host prenylation contributes to YopQ function.  Various biochemical approaches will also be employed to investigate the subcellular location of YopQ and to identify interacting proteins.

Select Publications

DeBord, K.L., D.M. Anderson, M.M. Marketon, K.A. Overheim, R.W. DePaolo, N. Ciletti, B. Jabri, and O. Schneewind.  2006.  Immunogenicity and protective immunity against bubonic plague and pneumonic plague by immunization of mice with the recombinant V10 antigen, a variant of LcrV.  Infect Immun. 74: 4910-4.

Marketon, M.M., E. Kwiatkowski, E.M. Glass, K.L. DeBord, O. Schneewind. Identification of Chromosomal Yersinia pestis Genes Required for Type III Secretion and Delivery of Yops into Target Cells. Manuscript in preparation.

Sorg, J.A, N.C. Miller, M.M. Marketon, and O. Schneewind.  2005.  Rejection of Impassable Substrates by Yersinia Type III Secretion Machines.  J. Bacteriol. 187: 7090-7102.

Marketon, M.M., R.W. DePaolo, K.L. DeBord, B. Jabri, and O. Schneewind.  2005.  Plague Bacteria Target Immune Cells during Infection.  Science 309: 1739-1741.

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