Microbiotas activate a tuft cell-sensory neuron feedback loop to promote esophagogastric junction (EGJ) tumorigenesis

Background Esophagogastric junction (EGJ) adenocarcinoma is rising in developed countries. While diet and reflux contribute, microbial-neuronal mechanisms remain poorly defined. Methods Metabolomic and 16S rRNA sequencing were performed on 40 paired human EGJ tumors and adjacent tissues, and single-cell RNA sequencing on five pairs. Mechanistic studies used the pL2-IL1B mouse model of Barrett's esophagus, Cck2r-CreERT2 mice to label EGJ progenitors, and crosses to p53 R172H ; pL2-IL1B to provide a tumor-prone background. Dclk1-CreERT2 targeted Tuft cells, while Trpv1-Cre, hM3Dq, and Ramp1 flox examined sensory signaling. Additional lines (Cck2r-CreERT2; Ramp1 flox/flox , Dclk1-CreERT2; Cox1 flox/flox ; Cox2 flox/flox , Pou2f3-CreERT2; DTA) dissected the Tuft-neuron-epithelial circuit. P. melaninogenica colonization and deoxycholic acid (DCA) supplementation modeled microbial-metabolic effects. Calcium imaging, phospho-ERK staining, and Tuft-neuron co-cultures assessed functional signaling. Results EGJ tumors were enriched in Firmicutes/Bacteroidota and Prevotella , while adjacent tissues contained Proteobacteria and Helicobacter . Prevotella abundance correlated positively with arachidonic-acid and negatively with tryptophan metabolites. Single-cell data showed epithelial, immune, and metabolic clusters, with bile- and fatty-acid pathways enriched in proliferative cells. P. melaninogenica , linked to bile-acid-rich, cancer-prone environments, was selected for mechanistic study. DCA alone enhanced EGJ dysplasia in pL2-IL1B mice. Oral P. melaninogenica colonized the EGJ, further increased by DCA, forming a bile-acid niche. Colonization amplified CGRP⁺ neuronal signaling, expanded Tuft cells, and aggravated dysplasia. Tumor-prone Cck2r-CreERT2; p53R172H; pL2-IL1B mice showed more Tuft-CGRP⁺ contacts, and pseudotime analysis indicated enhanced Cck2r⁺-to-Tuft differentiation. In ex vivo organoids, activation of Dclk1⁺; hM3Dq⁺ Tuft cells elevated PGE₂/NGF secretion, strengthened Tuft-nerve contacts, and induced calcium influx in co-cultured Trpv1⁺; GCaMP6s⁺ DRG neurons. Trpv1⁺ neurons co-expressed CGRP and EP4. Combined DCA + CNO treatment in Trpv1-Cre; hM3Dq; pL2-IL1B mice further increased dysplasia. Tuft ablation (Pou2f3-CreERT2; DTA), Tuft-specific Cox1/2 loss, or Ramp1 deletion in Cck2r⁺ cells suppressed P. melaninogenica -induced dysplasia, demonstrating that blocking the Tuft-neuron-progenitor axis halts tumor progression. Conclusions We define a microbiota-bile-acid-Tuft-neuron feedback loop driving EGJ tumorigenesis. P. melaninogenica and DCA activate Tuft cells to release PGE₂/NGF, stimulating EP4-CGRP/Ramp1 signaling and promoting epithelial dysplasia. Disrupting this circuit suppresses tumor growth, highlighting a metabolic-neuronal target for EGJ cancer prevention.