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Published on: November 18, 2013
Reduced expression and increased CpG dinucleotide methylation of the rat APOBEC-1 promoter in transgenic rabbits
Frank Apostel1, Reinhard Dammann, Gerd P Pfeifer
1Klinik und Poliklinik für Innere Medizin, Universitätsklinikum Hamburg-Eppendorf, Martinistrasse 52, Germany.
Abstract:
Editing of apolipoprotein (apo) B mRNA in liver limits the plasma LDL levels in horses, dogs, rats or mice. Species such as man or rabbit do not edit the hepatic apo B mRNA and are therefore susceptible to atherosclerosis and coronary artery disease due to elevated plasma LDL levels. The catalytic subunit APOBEC-1 is the only missing component of the apo B mRNA editing enzyme complex in the human or rabbit liver. Here we describe the generation of transgenic rabbits in which APOBEC-1 expression is mediated by the proximal promoter of the rat APOBEC-1 gene. These transgenic rabbits are healthy and fertile, and rat APOBEC-1 mRNA is expressed in liver, intestine, kidney, lung, brain and muscle. The transgenic APOBEC-1 expression is low and not sufficient to induce editing in rabbit liver. In rat, the proximal APOBEC-1 promoter demonstrates a progressive loss of CpG dinucleotide methylation towards the core promoter region that is entirely unmethylated. In the transgenic rabbits, this distinct pattern of CpG methylation is lost, and throughout the entire rat APOBEC-1 promoter, >90% of the CpGs are methylated. Thus, the weak proximal rat APOBEC-1 promoter appears to be down-regulated in the rabbit and may be species-specific.
Insights
Transgenic rabbits expressing rat APOBEC-1 showed no liver mRNA editing. The rat APOBEC-1 promoter was heavily methylated in rabbits, suggesting species-specific regulation limits its activity.
Area of Science:
- Molecular Biology
- Genetics
- Cardiovascular Disease Research
Background:
- Apolipoprotein (apo) B mRNA editing in the liver regulates plasma low-density lipoprotein (LDL) levels.
- Humans and rabbits lack hepatic apo B mRNA editing, contributing to elevated LDL and cardiovascular disease risk.
- APOBEC-1, the catalytic subunit, is the missing component for hepatic apo B mRNA editing in humans and rabbits.
Purpose of the Study:
- To investigate the potential of APOBEC-1 expression in transgenic rabbits to induce hepatic apo B mRNA editing.
- To explore the role of the rat APOBEC-1 promoter's methylation pattern in regulating gene expression in rabbits.
Main Methods:
- Generation of transgenic rabbits using the proximal promoter of the rat APOBEC-1 gene to drive APOBEC-1 expression.
- Analysis of rat APOBEC-1 mRNA expression in various tissues of the transgenic rabbits.
- Assessment of apo B mRNA editing activity in the liver of transgenic rabbits.
- Methylation analysis of the rat APOBEC-1 promoter in both rat and transgenic rabbit livers.
Main Results:
- Transgenic rabbits expressed rat APOBEC-1 mRNA in multiple tissues, including the liver, but expression levels were insufficient for apo B mRNA editing.
- The rat APOBEC-1 promoter exhibited a distinct, progressive loss of CpG methylation in rats, which was absent in transgenic rabbits.
- In transgenic rabbits, the rat APOBEC-1 promoter was heavily methylated (>90% CpG methylation), indicating promoter silencing.
Conclusions:
- The rat APOBEC-1 promoter is significantly down-regulated in rabbits due to extensive CpG methylation.
- This promoter activity appears to be species-specific, hindering the induction of hepatic apo B mRNA editing in rabbits.
- Further research is needed to identify suitable promoters for achieving therapeutic apo B mRNA editing in rabbits.
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