LC-MS/MS analysis of HD5-interacting proteins

Human enteric α-defensin 5 (HD5) acts as a “double-edged sword” in host immunity due to its prominent role in promoting pathogen infections, yet the mechanistic basis of its infection-enhancing operation remains unclarified. Here we show that HD5 induces abundant filopodial extensions in epithelial cells which capture Shigella, a major human enteroinvasive pathogen that exploits filopodia for invasion, at high efficiency, suggesting a novel human-specific mechanism of HD5-augmented bacterial invasion. Using multi-omics screening and both gut-on-chip and in vivo models, we identify the HD5 receptor as P2Y11, a purinergic receptor distributed apically on the luminal surface of human colonic epithelium, which to our knowledge is the first HD5 receptor for a specific cellular function. cAMP-PKA signaling was identified to be the main pathway mediating the cytoskeleton-regulating activity of HD5, whose critical residues involved were also determined. In illuminating this mechanism of Shigella invasion, our findings raise the possibility of new intervention strategies against HD5-augmented infections.