Thesis: Alix Bruneau – Regulation of membrane expression of the bile phospholipid transporter ABCB4 : Effect of mutations
October 02 - 2019
directed by Tounsia Aït Slimane
ABCB4 is an ABC transporter expressed at the level of the hepatocyte ducticular membrane. Its function is to secrete phosphatidylcholine, a major component of bile. Since biliary diseases could be associated with a molecular defect of ABCB4, more than 500 variations have been identified. The most severe pathology is progressive familial intrahepatic cholestasis type 3 (PFIC3). It develops early in childhood and progresses rapidly to cirrhosis and liver failure before adulthood. The only effective therapy is liver transplantation, but it has limitations: graft availability and post-transplant mortality. The development of alternative therapies is therefore a key issue. This thesis focuses on the effect of five mutations described in patients located in ABCB4 ATP binding sites. Thanks to an innovative approach combining 3D modelling and in vitro studies, we have shown that these mutations are responsible for a lack of activity of the carrier ABCB4. We have shown that VX-770, a clinically approved drug to treat cystic fibrosis patients, can restore the activity of these five mutants. This work opens up prospects for the repositioning of VX-770 for the personalized treatment of patients with biliary pathologies related to ABCB4. The objective of the second part of this thesis is to study the role of the interaction of the N-terminal domain of ABCB4 with the MRCK-alpha kinase. Inhibition or extinction of this kinase shows that MRCK-alpha plays a role in regulating ABCB4 membrane expression by controlling its internalization from the plasma membrane. We then wanted to determine if the effect of MRCK-alpha on ABCB4 expression passes through its MLC2 effector, which is a partner in the ABCB4 linker domain. By inhibiting the MLC2 protein, we obtained the same effects on ABCB4 as when MRCK-alpha is inhibited. Our work clearly shows a common role of these two partners in regulating the internalization of ABCB4.