A structural and mechanistic model for BSEP dysfunction in PFIC2 cholestatic disease

Area of Science:

• Hepatology
• Molecular Biology
• Biophysics

Background:

• Bile Salt Export Pump (BSEP/ABCB11) is crucial for bile salt transport in hepatocytes.
• Biallelic BSEP mutations cause Progressive Familial Intrahepatic Cholestasis Type 2 (PFIC2), a severe pediatric liver disease.
• PFIC2 is often linked to missense mutations affecting BSEP expression, maturation, and trafficking.

Purpose of the Study:

• To investigate the impact of PFIC2-associated mutations on BSEP protein thermodynamic stability.
• To elucidate the molecular mechanisms underlying BSEP dysfunction in cholestatic liver disease.

Main Methods:

• In-cell thermal shift assay (CETSA) was used to measure the stability of 13 BSEP variants.
• High-resolution cryo-electron microscopy (cryo-EM) was employed to determine the structure of BSEP.

Main Results:

• CETSA identified a cluster of residues at the NBD2-ICL2 interface critical for BSEP stability.
• These residues showed significant destabilization in PFIC2 variants compared to wild-type BSEP.
• Cryo-EM structure revealed a novel NBD2-localized mechanism by which severe mutations induce cholestasis.

Conclusions:

• Specific BSEP mutations destabilize the protein, leading to impaired function and cholestatic liver disease.
• Findings provide a structural basis for understanding PFIC2 pathogenesis.
• This research may guide the development of small molecule therapies targeting BSEP trafficking defects.