Functional rescue of trafficking-impaired ABCB4 mutants by chemical chaperones

Abstract

Multidrug resistance protein 3 (MDR3, ABCB4) is a hepatocellular membrane protein that mediates biliary ecretion of phosphatidylcholine. Null mutations in ABCB4 gene give rise to severe early-onset cholestatic liver disease. We ave reviously shown that the diseaseassociated mutations p.G68R, p.G228R, p.D459H, and p.A934T resulted in retention of ABCB4 in the endoplasmic reticulum, thus failing to target the plasma membrane. In the present study, we tested the ability of wo compounds with chaperone-like activity, 4-phenylbutyrate and curcumin, to rescue these ABCB4 mutants by assessing their ffects on subcellular localization, protein maturation, and phospholipid efflux capability. Incubation of transfected cells at a educed temperature (30°C) or exposure to pharmacological doses of either 4-PBA or curcumin restored cell surface expression f mutants G228R and A934T. The delivery of these mutants to the plasma membrane was accompanied by a switch in the ratio f mature to inmature protein forms, leading to a predominant expression of the mature protein. This effect was due to an mprovement in the maturation rate and not to the stabilization of the mature forms. Both mutants were also functionally rescued, isplaying bile salt-dependent phospholipid efflux activity after addition of 4-PBA or curcumin. Druginduced rescue was mutant pecific, given neither 4-PBA nor curcumin had an effect on the ABCB4 mutants G68R and A934T. Collectively, these data ndicate that the functionality of selected trafficking-defective ABCB4 mutants can be recovered by chemical chaperones through restoration of membrane localization, suggesting a potential treatment for patients carrying such mutations.