Protein Quality Control is a Risk Factor and Therapeutic Target in Toxin-Induced Biliary Atresia

Extra-hepatic biliary atresia (BA) is an important pediatric liver disease of unknown etiology. The identification of biliatresone, a plant electrophile with selective toxicity for extra-hepatic cholangiocytes (EHC) and linked to outbreaks of epidemic BA in livestock, demonstrated the feasibility of an environmental trigger for this enigmatic condition. Here, we show that susceptibility of zebrafish EHC to biliatresone arises from their inability to restore stress-induced depletion of glutathione, and that heterozygous mutations in glutathione metabolism genes differentially sensitize EHC and the normally resistant intra-hepatic cholangiocytes (IHC) to biliatresone-mediated injury. Collectively these findings argue that genetic variants in stress-response genes could be risk factors for human BA. Supporting this idea, we identified de novo loss-of-function mutations in the STIP1 and REV1 gene, both part of the HSP90 pathway, in two children with BA, respectively, and showed that modulation of their expression sensitized zebrafish larvae and human cholangiocytes to biliatresone. Using an in vivo drug screening assay, we identified activators of cGMP signaling as potent enhancers of the anti-oxidant N-acetylcysteine (NAC) in biliatresone-treated larvae and human cholangiocytes that act independently of glutathione and upstream of chaperone-mediated protein quality control (PQC) pathways. Collectively, these data highlighted novel aspects of cholangiocyte injury responses, identified new genetic risk factors for BA, and suggest combined treatment with NAC and cGMP signaling modulators as a rational therapeutic strategy for BA.