A Molecular Mechanism of Intrahepatic Cholestasis in Osteo-Oto-Hepato-Enteric Syndrome

Intrahepatic cholestasis is a hallmark of the rare osteo-oto-hepato-enteric syndrome. Osteo-oto-hepato-enteric syndrome is caused by pathogenic variants in uncoordinated (UNC) 45A. UNC45A encodes a cochaperone that coordinates the proper folding and function of target proteins. The molecular basis of cholestasis in osteo-oto-hepato-enteric syndrome has not been demonstrated. Notably, the clinical phenotype overlaps with other familial cholestatic disorders involving impaired localization of the bile salt export pump (ABCB11).

To investigate a mechanistic link between UNC45A dysfunction and bile salt export pump localization, we used immunofluorescence microscopy on liver biopsies from patients, CRISPR-CRISPR-associated protein 9-engineered male hepatocyte cell lines, site-directed mutagenesis, and coimmunoprecipitation assays. We focused on the recurrent c.292C>T/p.(Arg98Trp) variant.

Liver tissue of a patient carrying the UNC45A-c.292C>T/p.(Arg98Trp) variant displayed a zonated canalicular bile salt export pump expression defect. MYO5B, a known regulator of canalicular trafficking, was identified as an UNC45A client. UNC45A-p.(Arg98Trp) caused bile salt export pump to accumulate in MYO5B+/RAB11A+ endosomes that failed to reach the canalicular membrane. Mutant UNC45A showed a weakened interaction with heat shock protein (HSP) 70 chaperones. Of these, HSPA1A showed a zonated expression in the liver, inversely correlating with the zonated bile salt export pump defect in the patient. Paracetamol-mediated increase in heat shock protein A1A abundance restored MYO5B+/RAB11A+ endosome positioning and canalicular bile salt export pump localization.

A weakened UNC45A-p.(Arg98Trp)-HSP70 interaction disrupts bile salt export pump trafficking through impaired MYO5B function and zonated HSPA1A availability. These results connect osteo-oto-hepato-enteric syndrome with MYO5B-associated cholestasis and suggests HSP 70-targeted therapies to restore canalicular bile salt export pump expression.