In enterohemorrhagic E. coli (EHEC) infection, Shiga toxin (Stx) is a major factor in bacterial virulence, but must work in tandem with other bacterial virulence factors to produce severe disease. Although the bacterial protein intimin, expressed by the type III secretion system of E. coli, has been proposed as one of the other major virulence factors, recent evidence indicates that intimin is not sufficient for virulence. The 2011 outbreak of sprout-related diarrhea/hemolytic uremic syndrome (HUS) was caused by an intimin-negative strain of E. coli. This Shiga toxin-producing E. coli (STEC) strain affected >3800 patients, of which >20% developed HUS. Though STEC infections are often more severe in children or the elderly, this outbreak caused severe disease in healthy adults as well. Therefore, there must be a re-evaluation of the importance of different pathogenic EHEC proteins and their interaction with the host to cause severe disease. Research into EHEC infection in humans has been hindered by the lack of a physiologically relevant model system. The development of the human intestinal enteroids as mimicking both normal physiology and pathophysiology offers a powerful research tool to study EHEC proteins that cause disease and lead to preventative or curative therapy. This proposal will use enteroids to characterize one of EHEC's major pathogenic proteins, EspP, as stimulating both Shiga toxin translocation and intestinal stem cell cytoprotection. The proposal is divided up into two major aims:
Aim 1 - Characterize the serine protease activity of EspP and its role in induction of macropinocytosis and Shiga toxin uptake. The two subaims will focus on functionally characterizing the serine protease activity and endocytosis of EspP in order to cause changes to host proteins.
Aim 2 - Characterize the effects of EspP on intestinal stem cells.
This aim will determine the role EspP plays in disrupting the intestinal stem cell niche and how it contributes to the cytoprotection of the non-EHEC-infected intestinal cells. This proposal will contribute to the understanding of EHEC infections in human disease as well as develop a novel model for intestinal host-pathogen interactions. The candidate is committed to understanding gastrointestinal (GI) physiology and pathophysiology and has been at the forefront of developing a human model for these studies at her institution. The results obtained from these proposed studies will form the basis of the candidate's independent research as she focuses on a future academic research career. Her mentoring committee comprised of renowned GI and microbial researchers, as well as the research environment in the Division of Gastroenterology at Johns Hopkins, are ideal for fostering her development and road to independence as she focuses on characterizing the pathogenic proteins of EHEC and develops a career in functional understanding of enteric pathogens.

Public Health Relevance

Enterohemorrhagic Escherichia coli (EHEC) is a major life-threatening food borne pathogen for which is no currently available preventative or curative therapy. The EHEC-secreted protein, EspP, is involved in the uptake of Shiga toxins across the intestinal cells that can lead to systemic disease and renal failure and may be sufficient to cause EHEC-related infection, Shiga toxin virulence, as well as stimulating a cytoprotective effect of intestinal stem cells. Defining the functional characteristics of EspP, which shares >60% similarity with a family of pathogenic bacterial proteins, will provide valuable insight into: 1) te pathogenic functions of these proteins that are secreted by other major enteric bacteria (e.g. enteroaggregative E. coli, Shigella sp., Salmonella sp.), and 2) how intestinal stem cells are stimulated to regenerate and replace EHEC infected cells; a vital component in protecting us against enteric infections

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Scientist Development Award - Research & Training (K01)
Project #
5K01DK106323-03
Application #
9413485
Study Section
Kidney, Urologic and Hematologic Diseases D Subcommittee (DDK)
Program Officer
Saslowsky, David E
Project Start
2016-04-01
Project End
2021-01-31
Budget Start
2018-02-01
Budget End
2019-01-31
Support Year
3
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Johns Hopkins University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21205
In, Julie G; Foulke-Abel, Jennifer; Estes, Mary K et al. (2016) Human mini-guts: new insights into intestinal physiology and host-pathogen interactions. Nat Rev Gastroenterol Hepatol 13:633-642