ADH1B, the adipocyte-enriched alcohol dehydrogenase, plays an essential, cell-autonomous role in human adipogenesis.

Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its expression in adipose tissue is reduced in obesity and increased by insulin stimulation. Interference with expression has also been reported to impair adipocyte function.

Ivacaftor-Mediated Potentiation of ABCB4 Missense Mutations Affecting Critical Motifs of the NBDs: Repositioning Perspectives for Hepatobiliary Diseases.

ABCB4 (ATP-binding cassette subfamily B member 4) is a hepatocanalicular floppase involved in biliary phosphatidylcholine (PC) secretion. Variations in the ABCB4 gene give rise to several biliary diseases, including progressive familial intrahepatic cholestasis type 3 (PFIC3), an autosomal recessive disease that can be lethal in the absence of liver transplantation. In this study, we investigated the effect and potential rescue of ten ABCB4 missense variations in NBD1:NBD2 homologous positions (Y403H/Y1043H, K435M/K1075M, E558K/E1200A, D564G/D1206G and H589Y/H1231Y) all localized at the conserved and functionally critical motifs of ABC transporters, six of which are mutated in patients.

MRCK-Alpha and Its Effector Myosin II Regulatory Light Chain Bind ABCB4 and Regulate Its Membrane Expression.

ABCB4, is an adenosine triphosphate-binding cassette (ABC) transporter localized at the canalicular membrane of hepatocytes, where it mediates phosphatidylcholine secretion into bile. Gene variations of ABCB4 cause different types of liver diseases, including progressive familial intrahepatic cholestasis type 3 (PFIC3). The molecular mechanisms underlying the trafficking of ABCB4 to and from the canalicular membrane are still unknown.

RAB10 Interacts with ABCB4 and Regulates Its Intracellular Traffic.

ABCB4 (ATP-binding cassette subfamily B member 4) is an ABC transporter expressed at the canalicular membrane of hepatocytes where it ensures phosphatidylcholine secretion into bile. Genetic variations of ABCB4 are associated with several rare cholestatic diseases. The available treatments are not efficient for a significant proportion of patients with ABCB4-related diseases and liver transplantation is often required.

Molecular Regulation of Canalicular ABC Transporters.

The ATP-binding cassette (ABC) transporters expressed at the canalicular membrane of hepatocytes mediate the secretion of several compounds into the bile canaliculi and therefore play a key role in bile secretion. Among these transporters, ABCB11 secretes bile acids, ABCB4 translocates phosphatidylcholine and ABCG5/G8 is responsible for cholesterol secretion, while ABCB1 and ABCC2 transport a variety of drugs and other compounds. The dysfunction of these transporters leads to severe, rare, evolutionary biliary diseases.

Effect of CFTR correctors on the traffic and the function of intracellularly retained ABCB4 variants.

Background & Aim: ABCB4 is expressed at the canalicular membrane of hepatocytes. This ATP-binding cassette (ABC) transporter is responsible for the secretion of phosphatidylcholine into bile canaliculi. Missense genetic variations of ABCB4 are correlated with several rare cholestatic liver diseases, the most severe being progressive familial intrahepatic cholestasis type 3 (PFIC3).

Functional rescue of an ABCB11 mutant by ivacaftor: A new targeted pharmacotherapy approach in bile salt export pump deficiency.

Background & Aim: The canalicular bile salt export pump (BSEP/ABCB11) of hepatocytes is the main adenosine triphosphate (ATP)-binding cassette (ABC) transporter responsible for bile acid secretion. Mutations in ABCB11 cause several cholestatic diseases, including progressive familial intrahepatic cholestasis type 2 (PFIC2) often lethal in absence of liver transplantation. We investigated in vitro the effect and potential rescue of a BSEP mutation by ivacaftor, a clinically approved cystic fibrosis transmembrane conductance regulator (CFTR/ABCC7) potentiator.

Inhibition of receptor-interacting protein kinase 1 improves experimental non-alcoholic fatty liver disease.

Background & Aims: In non-alcoholic fatty liver disease (NAFLD), hepatocytes can undergo necroptosis: a regulated form of necrotic cell death mediated by the receptor-interacting protein kinase (RIPK) 1. Herein, we assessed the potential for RIPK1 and its downstream effector mixed lineage kinase domain-like protein (MLKL) to act as therapeutic targets and markers of activity in NAFLD.

Methods: C57/BL6J-mice were fed a normal chow diet or a high-fat diet (HFD).

Comparison of in silico prediction and experimental assessment of ABCB4 variants identified in patients with biliary diseases.

Genetic variations of the phosphatidylcholine transporter, ABCB4 cause several biliary diseases. The large number of reported variations makes it difficult to foresee a comprehensive study of each variation. To appreciate the reliability of in silico prediction programs, 1) we confronted them with the assessment in cell models of two ABCB4 variations (E528D and P1161S) identified in patients with low phospholipid-associated cholelithiasis (LPAC); 2) we extended the confrontation to 19 variations that we had previously characterized in cellulo.

A PDZ-Like Motif in the Biliary Transporter ABCB4 Interacts with the Scaffold Protein EBP50 and Regulates ABCB4 Cell Surface Expression.

ABCB4/MDR3, a member of the ABC superfamily, is an ATP-dependent phosphatidylcholine translocator expressed at the canalicular membrane of hepatocytes. Defects in the ABCB4 gene are associated with rare biliary diseases. It is essential to understand the mechanisms of its canalicular membrane expression in particular for the development of new therapies.

A functional classification of ABCB4 variations causing progressive familial intrahepatic cholestasis type 3.

Unlabelled: Progressive familial intrahepatic cholestasis type 3 is caused by biallelic variations of ABCB4, most often (≥70%) missense. In this study, we examined the effects of 12 missense variations identified in progressive familial intrahepatic cholestasis type 3 patients. We classified these variations on the basis of the defects thus identified and explored potential rescue of trafficking-defective mutants by pharmacological means.

Phosphorylation of ABCB4 impacts its function: insights from disease-causing mutations.

Unlabelled: The ABCB4 transporter mediates phosphatidylcholine (PC) secretion at the canalicular membrane of hepatocytes and its genetic defects cause biliary diseases. Whereas ABCB4 shares high sequence identity with the multidrug transporter, ABCB1, its N-terminal domain is poorly conserved, leading us to hypothesize a functional specificity of this domain. A database of ABCB4 genotyping in a large series of patients was screened for variations altering residues of the N-terminal domain.

Oligomerization is required for normal endocytosis/transcytosis of a GPI-anchored protein in polarized hepatic cells.

Targeting of glycosyl-phosphatidylinositol (GPI)-anchored proteins (GPI-APs) in polarized epithelial cells depends on their association with detergent-resistant membrane microdomains called rafts. In MDCK cells, GPI-APs associate with rafts in the trans-Golgi network and are directly delivered to the apical membrane. It has been shown that oligomerization is required for their stabilization in rafts and their apical targeting.

Effects of cellular, chemical, and pharmacological chaperones on the rescue of a trafficking-defective mutant of the ATP-binding cassette transporter proteins ABCB1/ABCB4.

The ATP-binding cassette transporter ABCB4 is a phosphatidylcholine translocator specifically expressed at the bile canalicular membrane in hepatocytes, highly homologous to the multidrug transporter ABCB1. Variations in the ABCB4 gene sequence cause progressive familial intrahepatic cholestasis type 3. We have shown previously that the I541F mutation, when reproduced either in ABCB1 or in ABCB4, led to retention in the endoplasmic reticulum (ER)/Golgi.

Basolateral internalization of GPI-anchored proteins occurs via a clathrin-independent flotillin-dependent pathway in polarized hepatic cells.

In polarized hepatocytes, the predominant route for apical resident proteins to reach the apical bile canalicular membrane is transcytosis. Apical proteins are first sorted to the basolateral membrane from which they are internalized and transported to the opposite surface. We have noted previously that transmembrane proteins and GPI-anchored proteins reach the apical bile canaliculi at very different rates.

A missense mutation in ABCB4 gene involved in progressive familial intrahepatic cholestasis type 3 leads to a folding defect that can be rescued by low temperature.

Unlabelled: Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a rare liver disease characterized by early onset of cholestasis that leads to cirrhosis and liver failure before adulthood. PFIC3 may be improved by chronic administration of ursodeoxycholic acid, although in many cases liver transplantation is the only therapy. The disease is caused by mutations of the adenosine triphosphate (ATP)-binding cassette, sub-family B, member 4 (ABCB4) [multidrug resistance 3 (MDR3)] gene encoding a specific hepatocellular canalicular transporter involved in biliary phosphatidylcholine secretion.

[FRAP, FLIP, FRET, BRET, FLIM, PRIM...new techniques for a colourful life].

Cell and tissue imaging provides scientists with wonderful tools, thanks to a fruitful dialog between chemistry, optical, mechanical, computational sciences and biology. Confocal microscopy, videomicroscopy together with a new generation of fluorochromes (especially those derived from green fluorescent protein, GFP) and image analysis software allow to visualize life in all its dimensions (space and time). Cell imaging also allows to quantify biological processes at the cellular level, to analyse both stoechiometry and dynamics of molecular interactions involved in cell and tissue regulations.

[Mechanisms of sorting and transport of proteins to tw membrane domains of epithelial cells].

Polarity is a fundamental characteristic of most eukaryotic cells. The plasma membrane of such cells consists in two structurally and functionally different domains, i.e.

1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside blocks the apical biosynthetic pathway in polarized HT-29 cells.

In previous work we reported that long term treatment of polarized HT-29 cells by 1-benzyl-2-acetamido-2-deoxy-alpha-d-galactopyranoside (GalNAcalpha-O-bn) induced undersialylation and intracellular distribution of apical glycoproteins such as dipeptidyl peptidase IV (DPP-IV), and we suggested therefore that sialylation could act as an apical targeting signal. In this work, the apical direct biosynthetic route was studied after transfection of polarized enterocyte-like HT-29 5M12 cloned cells with a murine cDNA coding for a soluble form of DPP-IV, which was secreted into the apical medium. A 24-h treatment of transfected cells by GalNAcalpha-O-bn markedly inhibited the apical secretion and the sialylation of this soluble murine DPP-IV, which became blocked inside the cell.

Membrane dynamics and cell polarity: the role of sphingolipids.

In recent years, glycosphingolipids (GSLs) have attracted widespread attention due to the appreciation that this class of lipids has a major impact on biological life. Inhibition of the synthesis of glucosylceramide, which serves as a precursor for the generation of complex glycosphinglipids, is embryonic lethal. GSLs play a major role in growth and development.

Raft-mediated trafficking of apical resident proteins occurs in both direct and transcytotic pathways in polarized hepatic cells: role of distinct lipid microdomains.

In polarized hepatic cells, pathways and molecular principles mediating the flow of resident apical bile canalicular proteins have not yet been resolved. Herein, we have investigated apical trafficking of a glycosylphosphatidylinositol-linked and two single transmembrane domain proteins on the one hand, and two polytopic proteins on the other in polarized HepG2 cells. We demonstrate that the former arrive at the bile canalicular membrane via the indirect transcytotic pathway, whereas the polytopic proteins reach the apical membrane directly, after Golgi exit.