Tuft cell-derived acetylcholine promotes epithelial chloride secretion and intestinal helminth clearance.

Epithelial cells secrete chloride to regulate water release at mucosal barriers, supporting both homeostatic hydration and the "weep" response that is critical for type 2 immune defense against parasitic worms (helminths). Epithelial tuft cells in the small intestine sense helminths and release cytokines and lipids to activate type 2 immune cells, but whether they regulate epithelial secretion is unknown. Here, we found that tuft cell activation rapidly induced epithelial chloride secretion in the small intestine. This response required tuft cell sensory functions and tuft cell-derived acetylcholine (ACh), which acted directly on neighboring epithelial cells to stimulate chloride secretion, independent of neurons. Maximal tuft cell-induced chloride secretion coincided with immune restriction of helminths, and clearance was delayed in mice lacking tuft cell-derived ACh, despite normal type 2 inflammation. Thus, we have uncovered an epithelium-intrinsic response unit that uses ACh to couple tuft cell sensing to the secretory defenses of neighboring epithelial cells.

Genetic mapping reveals Pou2af2/OCA-T1-dependent tuning of tuft cell differentiation and intestinal type 2 immunity.

Chemosensory epithelial tuft cells contribute to innate immunity at barrier surfaces, but their differentiation from epithelial progenitors is not well understood. Here, we exploited differences between inbred mouse strains to identify an epithelium-intrinsic mechanism that regulates tuft cell differentiation and tunes innate type 2 immunity in the small intestine. Balb/cJ (Balb) mice had fewer intestinal tuft cells than C57BL/6J (B6) mice and failed to respond to the tuft cell ligand succinate. Most of this differential succinate response was determined by the 50- to 67-Mb interval of chromosome 9 (Chr9), such that congenic Balb mice carrying the B6 Chr9 interval had elevated baseline numbers of tuft cells and responded to succinate. The Chr9 locus includes Pou2af2, which encodes the protein OCA-T1, a transcriptional cofactor essential for tuft cell development. Epithelial crypts expressed a previously unannotated short isoform of Pou2af2 predicted to use a distinct transcriptional start site and encode a nonfunctional protein. Low tuft cell numbers and the resulting lack of succinate response in Balb mice were explained by a preferential expression of the short isoform and could be rescued by expression of full-length Pou2af2. Physiologically, Pou2af2 isoform usage tuned innate type 2 immunity in the small intestine. Balb mice maintained responsiveness to helminth pathogens while ignoring commensal Tritrichomonas protists and reducing norovirus burdens.

Tuft cell-derived acetylcholine regulates epithelial fluid secretion.

Tuft cells are solitary chemosensory epithelial cells that can sense lumenal stimuli at mucosal barriers and secrete effector molecules to regulate the physiology and immune state of their surrounding tissue. In the small intestine, tuft cells detect parasitic worms (helminths) and microbe-derived succinate, and signal to immune cells to trigger a Type 2 immune response that leads to extensive epithelial remodeling spanning several days. Acetylcholine (ACh) from airway tuft cells has been shown to stimulate acute changes in breathing and mucocilliary clearance, but its function in the intestine is unknown. Here we show that tuft cell chemosensing in the intestine leads to release of ACh, but that this does not contribute to immune cell activation or associated tissue remodeling. Instead, tuft cell-derived ACh triggers immediate fluid secretion from neighboring epithelial cells into the intestinal lumen. This tuft cell-regulated fluid secretion is amplified during Type 2 inflammation, and helminth clearance is delayed in mice lacking tuft cell ACh. The coupling of the chemosensory function of tuft cells with fluid secretion creates an epithelium-intrinsic response unit that effects a physiological change within seconds of activation. This response mechanism is shared by tuft cells across tissues, and serves to regulate the epithelial secretion that is both a hallmark of Type 2 immunity and an essential component of homeostatic maintenance at mucosal barriers.