AP2 adaptor complex mediates bile salt export pump internalization and modulates its hepatocanalicular expression and...

Area of Science:

• Hepatology
• Molecular Biology
• Cellular Physiology

Background:

• The bile salt export pump (BSEP) is vital for excreting bile salts into bile, and its impaired function leads to intrahepatic cholestasis.
• Reduced BSEP levels at the canalicular membrane (CM), often due to increased internalization, contribute to cholestatic liver diseases.
• The molecular mechanisms driving BSEP internalization from the CM are not fully understood.

Purpose of the Study:

• To investigate the role of the AP2 complex in the internalization of BSEP from the hepatocyte canalicular membrane.
• To explore the potential of targeting the AP2-BSEP interaction for therapeutic interventions in cholestatic liver diseases.

Main Methods:

• Utilized liver specimens from rats and OTCD patients, HeLa cells expressing BSEP, and primary human hepatocytes.
• Employed RNA interference to impair AP2 function, targeting α-adaptin.
• Conducted immunostaining, coimmunoprecipitation, glutathione S-transferase pulldown assays, and time-lapse imaging.

Main Results:

• 4-phenylbutyrate (4PBA) treatment reduced α- and μ2-adaptin expression in OTCD liver samples, suggesting AP2 involvement.
• Impairing AP2 function via RNA interference significantly inhibited BSEP internalization and increased cell-surface BSEP expression and function.
• Demonstrated direct interaction between AP2 and BSEP at the CM via a tyrosine motif in BSEP's carboxyl terminus, mediating internalization.

Conclusions:

• The AP2 complex directly interacts with BSEP at the canalicular membrane, mediating its internalization and subsequent degradation.
• This AP2-mediated internalization regulates BSEP's canalicular expression and transport function.
• Understanding this mechanism offers insights into the pathogenesis of severe liver diseases characterized by intrahepatic cholestasis.