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theses

Communication between stem cells and its niche-supporting cells maintains the homeostasis of adult tissues. Wnt signaling is a critical regulator of stem cell niche, but the mechanism that governs Wnt-ligand delivery in this compartment has not been fully investigated. We identified that Wnt secretion is partly dependent on Rab8a-mediated anterograde transport of Gpr177/Wntless, a Wnt-specific transmembrane transporter. Gpr177 binds to Rab8a, depletion of which compromises Gpr177 traffic thereby weakening the secretion of multiple Wnts. Analyses of generic Wnt/-catenin targets in Rab8a knockout mouse intestinal crypts indicate reduced signaling activities. There, maturation of Paneth cell–a Wnt-dependent cell type is severely affected. Rab8a knockout crypts show an expansion of Lgr5+ and Hopx+ cells in vivo. However, in vitro, the knockout enteroids exhibit significantly weakened growth that can be partly restored by exogenous Wnts or GSK3 inhibitors. Immunogold labeling and surface protein isolation identified decreased plasma membrane localization of Gpr177 in Rab8a knockout Paneth cells and fibroblasts. On stimulation by exogenous Wnts, Rab8a deficient cells show ligand-induced Lrp6 phosphorylation and transcriptional reporter activation. Rab8a thus controls Wnt exocytosis in ligand producing cells and is critical for Paneth cell maturation. Additionally, deletion of Gpr177 alone from epithelium and underlying mucosal layer of intestine showed only subtle changes in stem cell number in vivo. However interestingly, significant deterioration in stem cell activity led to death of all Gpr177 null organoids within 4 days in culture. Taken together our data highlight profound tissue plasticity in vivo in response to stress induced by depletion of a stem cell niche signal.

ETD_6868

Ph.D.

Includes bibliographical references

by Soumyashree Das

doi:10.7282/T3XS5XGR

ETD doctoral

The author owns the copyright to this work.