The mitochondrial BCKD complex interacts with hepatic apolipoprotein E in cultured cells in vitro and mouse livers in vivo

Abstract

Background and aims Apolipoprotein E (APOE) is known for its role in lipid metabolism and its association with age-related disease pathology. The aim of the present work was to identify previously unknown functions of APOE based on the detection of novel APOE protein–protein interaction candidates. Approach and results APOE targeted replacement mice and transfected cultured hepatocytes expressing the human isoforms APOE3 and APOE4 were used. For 7 months, APOE3 and APOE4 mice were fed a high-fat and high-sugar diet to induce obesity, while a subgroup was subjected to 30% dietary restriction. Proteomic analysis of coimmunoprecipitation products from APOE mouse liver extracts revealed 28 APOE-interacting candidate proteins, including branched-chain alpha-keto acid dehydrogenase (BCKD) complex subunit alpha (BCKDHA) and voltage-dependent anion-selective channel 1 (VDAC1). The binding of APOE and BCKDHA was verified in situ by proximity ligation assay in cultured cells. The activity of the BCKD enzyme complex was significantly higher in obese APOE4 mice than in APOE3 mice, while the plasma levels of branched-chain amino acids and mTOR signalling proteins were not different. However, the protein–protein interaction with VDAC1 was strongly induced in APOE3 and APOE4 mice upon dietary restriction, suggesting a prominent role of APOE in mitochondrial function. Conclusions The protein–protein interactions of APOE with BCKDHA and VDAC1 appear to be of physiological relevance and are modulated upon dietary restriction. Because these are mitochondrial proteins, it may be suggested that APOE is involved in mitochondria-related processes and adaptation to hepatic energy demands.

Human APOE4 Protects High-Fat and High-Sucrose Diet Fed Targeted Replacement Mice against Fatty Liver Disease Compared to APOE3

“…( E ), the circulating APOE concentration was determined in the plasma of 4 h fasted mice after termination of the feeding trial using an ELISA kit. Hepatic APOE protein levels were analyzed by Western blotting and the data shown are according to Rueter et al [ 57 ]. Relative mRNA levels of human APOE were analyzed in the liver (squares) and visceral adipose tissue (circles) by qRT-PCR.…”

“…The study was performed according to the German regulations of animal welfare and approved by the local animal ethical committee. The mouse experiment has been described also elsewhere [ 55 - 57 ]. In brief, twelve male APOE3 and APOE4 targeted replacement (six per APOE genotype group) and six unmodified C57Bl/ 6JBOM mice were purchased from Taconic Europe A/S (Ry, Denmark).…”

“…Target proteins were normalized by the total protein load per lane, measured as membrane fluorescence and as described previously [ 55 ]. The determination of hepatic APOE has been performed in a previously published study and the values are according to Rueter et al [ 57 ].…”

Developing a novel immune infiltration-associated mitophagy prediction model for amyotrophic lateral sclerosis using bioinformatics strategies

“…BCKDHA and VDAC1 are both mitochondrial proteins that jointly participate in lipid metabolism processes. Research has indicated that BCKDHA and VDAC1 can undergo immunoprecipitation with APOE in mouse liver extracts ( 32 ). This interaction plays a crucial role in the energy production processes of mitochondria and helps the liver to adapt to its energy demands.…”