Bacterial Stimulation of Intestinal Proliferation via the Wnt Pathway in Zebrafish
Neal, James Thomas
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Neal, James Thomas
This dissertation describes research into microbial influences on host signaling in the zebrafish intestine. Vertebrate organisms are consistently exposed to microbes, especially on epithelial tissues that are exposed to the environment, such as the skin and the gastrointestinal tract. The close association between these tissues and microbes over time has resulted in a symbiotic state, whereby microorganisms have gained the ability to utilize vertebrate epithelia as a niche for replication and the acquisition of nutrients. These associations run the gamut from beneficial to exceedingly pathogenic and often involve complex bidirectional signaling between microbe and host. Microbial signals can interact with host cell pathways involved in a wide range of cellular processes. Here, we describe our investigations into one such pathway, the Wnt signaling pathway, and how microbial activation of Wnt signaling can translate into alterations in cell proliferation and homeostasis in the intestinal epithelium of the teleost fish Danio rerio. We report that epithelial cell proliferation in the developing zebrafish intestine is stimulated both by the presence of the resident microbiota and by activation of Wnt signaling and demonstrate that resident intestinal bacteria enhance the stability of β-catenin in intestinal epithelial cells, promoting cell proliferation in the developing vertebrate intestine. We also describe how transgenic expression of the bacterial effector protein CagA from the human gastric pathogen Helicobacter pylori is capable of causing significant overproliferation of the intestinal epithelium and adult intestinal hyperplasia, as well as significant upregulation of the Wnt target genes cyclinD1 and the zebrafish c-myc ortholog myca. We show that co-expression of CagA with a mutant allele of the β-catenin destruction complex protein Axin1 resulted in a further increase in intestinal proliferation, while co-expression of CagA with a null allele of the essential β-catenin transcriptional cofactor Tcf4 restored intestinal proliferation to wild-type levels. These results suggest that CagA activates canonical Wnt signaling downstream of the β-catenin destruction complex and upstream of Tcf4. Our studies provide in vivo evidence of Wnt pathway activation by CagA and implicate this activation in CagA-induced epithelial overproliferation, an early step in gastrointestinal cancer development. This dissertation contains both my previously published and unpublished co-authored material.