Novel zebrafish mutants reveal new roles for Apolipoprotein B during embryonic development and pathological conditions

ABSTRACTApolipoprotein B (ApoB) is the primary protein of chylomicrons, VLDLs and LDLs and is essential for their assembly. Defects in ApoB synthesis and secretion result in several human diseases, including abetalipoproteinemia and familial hypobetalipoproteinemia. Conversely, high levels of APOB in plasma are associated with increased risk for coronary heart disease and atherosclerosis.The involvement of APOB in lipid metabolism and atherogenesis prompted the generation of several mutant mice. However, as APOB is required for supplying nutrients to the developing embryo, ApoB null mice are embryonic lethal, thereby precluding the study of the roles of this protein during development.Here, we established novel zebrafish mutants for two apoB genes: apoBa and apoBb.1. Double-mutant embryos display clear hallmarks of human hypolipidemia-related diseases, including intestinal defects and fatty liver, as well as profound vascular defects. We further use these models to identify the domains within ApoB responsible for its functions. By assessing the ability of different truncated forms of human APOB to rescue the mutant phenotypes, we demonstrate the benefits of this model for prospective therapeutic screens. Overall, our novel zebrafish models uncover new functions of ApoB in organ development and morphogenesis and shed new light on the mechanisms underlying hypolipidemia-related diseases.

Apolipoprotein B100 biogenesis: a complex array of intracellular mechanisms regulating folding, stability, and lipoprotein assemblyThis paper is one of a selection of papers published in this special issue entitled “Canadian Society of Biochemistry, Molecular & Cellular Biology 52nd Annual Meeting — Protein Folding: Principles and Diseases” and has undergone the Journal's usual peer review process.

Apolipoprotein B100 (apoB) is a large amphipathic lipid-binding protein that is synthesized by hepatocytes and used to assemble and stabilize very low density lipoproteins (VLDL). It may have been derived through evolution from other lipid-associating proteins such as microsomal triglyceride transfer protein or vitellogenin. The correct folding of apoB requires assistance from chaperone proteins in co-translational lipidation, disulfide bond formation, and glycosylation. Any impairment in these processes results in co-translational targeting of the misfolded apoB molecule for proteasomal degradation. In fact, most of the regulation of apoB production is mediated by intracellular degradation. ApoB that misfolds post-translationally, perhaps as a result of oxidative stress, may be eliminated through autophagy. This review focuses on the proposed pentapartite domain structure of apoB, the role that each domain plays in the binding of lipid species and regulation of apoB synthesis, and the process of VLDL assembly. The factors involved in the recognition, ubiquitination, and proteasomal delivery of defective apoB molecules are also discussed.

Effects of dietary coconut oil on apolipoprotein B synthesis and VLDL secretion by calf liver slices

Incorporation of coconut oil (CO) rich in lauric acid into the milk diet induces a lipid infiltration of the liver (steatosis) in 1-month-old calves. Among possible steps involved in diet-induced liver steatosis, the ability of the calf liver to synthesize apolipoprotein (Apo) B and to secrete it as part of VLDL particles was investigated. Liver samples were taken from calves fed for 17 d on a conventional milk replacer containing CO (n5) and beef tallow (BT,n4) as reference. Samples were cut into slices 0·5 mm thick and subsequently incubated for 12 h in a medium containing a [35S]methionine–[35S]cysteine mix and 0·8 mM-sodium laurate or oleate, the major fatty acids of CO and BT diets respectively. Concentrations of total [35S]proteins, [35S]albumin and [35S]ApoB in liver cells were 2-fold lower (P=0·08, 0·0004 and 0·03 respectively) in CO- than in BT-fed calves. Although the total amount of proteins secreted (including albumin) was similar in both groups of calves, the amount of VLDL-[35S]Apo secreted was 2-fold lower (P=0·004) in CO- than in BT-fed calves. These results suggest that a CO-enriched milk diet induces in preruminant calves a lipid infiltration of the liver by decreasing ApoB synthesis, leading to a reduction in secretion of VLDL particles.