Abstract
The nuclear receptor constitutive androstane receptor (CAR) is a key regulator for drug metabolism in liver. Human CAR (hCAR) transcripts are subjected to alternative splicing. Some hCAR splicing variants (SVs) have been shown to encode functional proteins by reporter assays. However, in vivo research on the activity of these hCAR SVs has been impeded by the absence of a valid model. This study engineered an hCAR-BAC-transgenic (hCAR-TG) mouse model by integrating the 8.5-kbp hCAR gene as well as 73-kbp upstream and 91-kbp downstream human genomic DNA into the genome of CAR-null mice. A series of experiments demonstrate that (1) the expression of major hCAR mRNA SVs, SV0-4, in livers of hCAR-TG mice is comparable to that in human livers; (2) the hCAR SVs are predominantly expressed in liver, which resembles the tissue distribution of CAR in humans, but diverges from that in mice; and (3) major hCAR mRNA SVs increase markedly in postnatal livers of hCAR-TG mice, which mimics the ontogeny of CAR mRNA in humans. Thus, the transgene likely contains all the functional regulatory elements controlling proper spatial and temporal expression of the hCAR gene. Moreover, hCAR-TG mice respond to the hCAR-specific agonist 6-(4-chlorophenyl)imidazo[2,1-b] [1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime instead of the mouse CAR agonist 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene, as well as the common CAR activator, phenobarbital, suggesting that hCAR is fully functional in livers of transgenic mice. In summary, the hCAR-TG mice developed by this study represent a valid model for studying in vivo function and regulation of hCAR and its splicing variants.