Complete Phenotypic Characterization of apobec-1 Knockout Mice with a Wild-type Genetic Background and a Human Apolipoprotein B Transgenic Background, and Restoration of Apolipoprotein B mRNA Editing by Somatic Gene Transfer of Apobec-1

Abstract

We have produced gene knockout mice by targeted disruption of the apobec-1 gene. As recently reported by Hirano et al. (Hirano, K.-I., Young, S. G., Farese, R. V., Jr., Ng, J., Sande, E., Warburton, C., Powell-Braxton, L. M., and Davidson, N. O. (1996) J. Biol. Chem. 271, 9887-9890), these animals do not edit apolipoprotein (apo) B mRNA or produce apoB-48. In this study we have performed a detailed analysis of the lipoprotein phenotypic effects of apobec-1 gene disruption that were not examined in the previous study. We first analyzed the plasma lipoproteins in knockout animals with a wild-type genetic background. Although there was no difference in plasma cholesterol between apobec-1(-/-), +/-, or +/+ mice, there was a marked (176%) increase in plasma apoB-100, from 1.8 +/- 1.2 mg/dl in apobec-1(+/+) mice to 2.7 +/- 0.6 mg/dl in apobec-1(+/-) and 5.0 +/- 1.4 mg/dl in apobec-1(-/-) mice. Plasma apoE was similar in these animals. By fast protein liquid chromatography (FPLC) analysis, there was a significant decrease in plasma high density lipoprotein (HDL) cholesterol in apobec-1(-/-) mice. We further fractionated the plasma lipoproteins into d < 1.006, 1.006-1.02, 1.02-1.05, 1.05-1.08, 1.08-1.10, and 1.10-1.21 g/ml classes, and found a marked (30-40%) reduction in the cholesterol and protein content in the (d 1.08-1.10 and 1.10-1.21) HDL fractions, corroborating the FPLC data. SDS-gel analysis revealed an absence of apoB-48, an increase in apoB-100 in the very low density lipoprotein (VLDL) and low density lipoprotein (LDL) fractions, and a small decrease in apoA-I in the HDL fractions in the apobec-1(-/-) samples. We next raised the basal plasma apoB levels in the apobec-1(-/-) animals by cross-breeding them with human apoB transgenic (TgB) mice. The plasma apoB-100 was 3-fold higher in apobec-1(-/-)/TgB+/- mice (26.6 +/- 18.3 mg/dl) than in apobec-1(+/+)/TgB+/- mice (9.8 +/- 3.9 mg/dl, p < 0.05). The apobec-1(-/-)/TgB+/- mice had a plasma cholesterol levels of 170 +/- 28 mg/dl and triglyceride levels of 106 +/- 31 mg/dl, which are 80% and 58% higher, respectively, than the corresponding values of 94 +/- 21 mg/dl and 67 +/- 11 mg/dl in apobec+/+/TgB+/- mice. By FPLC, the apobec-1(-/-)/TgB+/- animals developed markedly elevated plasma LDL cholesterol (518.5 +/- 329.5 microg/ml) that is 373% that of apobec1(+/+)/TgB+/- mice (139.0 +/- 87.0 microg/ml) (p < 0.05). The elevated plasma triglyceride was accounted for mainly by a 97% increase in VLDL triglyceride in the apobec1(-/-)/TgB+/- mice. We conclude that apobec-1(-/-) animals have a distinctive lipoprotein phenotype characterized by significant hyperapoB-100 and HDL deficiency in mice with a wild-type genetic background. Furthermore, the abolition of apoB mRNA editing elevates plasma total cholesterol and LDL cholesterol in apobec-1(-/-) animals with a TgB background. Finally, to exclude the possibility that absence of apoB mRNA editing was a secondary effect of chronic Apobec-1 deficiency, we treated apobec-1(-/-) mice with a replication-defective mouse Apobec-...

Liver-specific Inactivation of the Abetalipoproteinemia Gene Completely Abrogates Very Low Density Lipoprotein/Low Density Lipoprotein Production in a Viable Conditional Knockout Mouse

“…The cells were expanded to passage 14 and used to generate knockout mice as described previously (17). Three positive ES cell clones were injected into blastocysts of C57BL/6J, and chimeric mice were obtained.…”

APOBEC1 complementation factor (A1CF) is dispensable for C-to-U RNA editing in vivo

“…We analyzed phenotypic data generated by the JAX KOMP2 pipeline and available from the International Mouse Phenotyping Consortium (IMPC; www.mousephenotype.org/) and found significant impacts on the immune and hematopoietic system as well as altered homeostasis and metabolism. While no specific role for APOBEC1 has been reported in either the immune or hematopoietic system, knockouts do have altered cholesterol metabolism (Nakamuta et al 1996).…”

“…However, while global ablation of Apobec1 in a mouse model resulted in a complete loss of ApoB editing, there were no gross impacts on fecundity or fertility (Hirano et al 1996) and only moderate impacts on serum lipoprotein profiles (Nakamuta et al 1996). In contrast, while A1CF and APOBEC1 have very similar expression profiles in adult tissues, there was strong evidence that A1CF may have roles outside of C-to-U editing (Sowden et al 2002;Chester et al 2003).…”

“…Both types of editing influence RNA function and its regulation. Deficiency in RNA editing can lead to substantial physiological defects including embryonic or postnatal lethality in the case of A-to-I editing (Higuchi et al 2000;Hartner et al 2004) and metabolic disorders in the case of C-to-U editing (Nakamuta et al 1996). In mammals, the most common form of RNA editing is A-to-I, which is both relatively widespread throughout the body and impacts a broad range of targets.…”

Phylogenetic Analysis of the Apolipoprotein B mRNA-editing Region

“…Interestingly, the apoB RNAs from 29 of the 30 species were good substrates for APOBEC-1 and were edited between 33 and 59% by the rabbit APOBEC-1. Previously, it has been shown that rat and mouse apoB mRNA are good substrates for APOBEC-1, in vitro and in vivo, respectively (2,28,(32)(33)(34). However, the ApoB mRNA from one of those species, the guinea pig, was edited poorly (3%).…”