Mechanisms by which Shigella flexneri impairs dendritic cell function: silencing the link between innate and adaptive immunity

Abstract

Shigella flexneri is a Gram-negative bacterial pathogen that causes severe gastroenteritis in humans. Upon infection, Shigella invades the host colonic epithelium using a type III secretion system (T3SS) that is encoded by a large virulence plasmid. This secretion system injects protein virulence determinants, or “effectors” directly into the host cytosol where they interfere with host function. The innate immune response to Shigella infection has been the subject of intense research. By comparison, little is known about how Shigella affects adaptive immunity. Shigella infection fails to initiate long lasting immunity; therefore, no effective shigellosis vaccine has been developed. Antigen presenting cells, such as the dendritic cell, represent the link between innate and adaptive immune responses and I hypothesize that Shigella impairs dendritic cell function as a means of dampening long term immunity. Little is known about Shigella-dendritic cell interactions therefore the goal of this project was to characterize this relationship with a focus on Shigella’s main virulence factors- the T3SS effectors. After performing an initial screen with our library of single gene deletion mutants I found that many genes on the virulence plasmid play a role in killing dendritic cells. I chose four mutants from the deletion collection screen for further investigation: ΔospF and ΔospI to investigate manipulation of immune signaling in dendritic cells, and ΔmxiD and ΔmxiH to investigate programmed cell death mechanisms. I apply a yeast model system to characterize inhibition of immune signaling by outer Shigella proteins (Osps) and show that targeting of MAPK signaling is evolutionarily conserved. I use a chemical inhibitor to observe caspase reliance in cell death and measure activation of caspase-1 and IL1β among mutant and Wild-type infection conditions. Gaining a thorough understanding of the processes undergone by Shigella and related gastroenteric pathogens to interfere with immune responses paves the road for in vivo studies and targets for rational design in vaccine development against bacillary dysentery.