APOLIPOPROTEIN-B MESSENGER-RNA EDITING IN 12 DIFFERENT MAMMALIAN-SPECIES - HEPATIC EXPRESSION IS REFLECTED IN LOW CONCENTRATIONS OF APO-B-CONTAINING PLASMA-LIPOPROTEINS

Two different isoproteins are encoded by the apolipoprotein (apo) B ge ne, apoB-48 and apoB-100. ApoB-48, core component of intestinally deri ved chylomicrons, has an accelerated plasma turnover as compared with the full-length protein apoB-100. A posttranscriptional modification o f the apoB mRNA by conversion of cytidine into uridine at nucleotide p osition 6666 changes the genomically encoded glutamine codon CAA at am ino acid residue 2153 into a translational stop codon UAA. This mRNA e diting explains the formation of the truncated isoform apoB-48. In the present investigation editing of apoB mRNA in liver and intestine fro m 12 different mammalian species was measured by a quantitative primer extension analysis of reverse-transcribed and polymerase chain reacti on- (PCR) amplified apoB mRNA in order to determine whether i) editing of apoB mRNA is generally restricted to the intestine or may also be found in the liver of other species than rodents, and ii) hepatic expr ession of apoB mRNA editing influences lipoprotein concentrations in p lasma. Intestinal apoB mRNA was edited at high levels in all species, 40% in sheep, 73% in horse, 82% in pig, 84% in dog, 84% in cat, 87% in guinea pig, 88% in rat, 89% in mouse, and > 90% in human, monkey, cow , and rabbit. In liver apoB mRNA was edited to 18% in dog, to 43% in h orse, to 62% in rat, and to 70% in mouse. Low levels of editing below 1% were detected in liver of rabbit and guinea pig. In contrast, hepat ic apoB mRNA from human, monkey, pig, cow, sheep, and cat liver was no t edited. The results of the primer extension analysis were confirmed by cloning and sequencing of the PCR products from dog, horse, cat, gu inea pig, sheep, and cow for all of which the apoB cDNA sequence had n ot been established by previous investigations. Primer extension analy sis of apoB mRNA from dog intestine and dog liver indicated C/U editin g at C655 in addition to C666. Cloning and sequencing of apoB cDNA fro m dog liver and intestine confirmed additional C/U editing at C655 whi ch changes ACA for threonine at amino acid residue 2149 into AUA for i soleucine. Synthesis and secretion of apoB-48-containing lipoproteins from liver was demonstrated by pulse labeling of freshly isolated hors e hepatocytes and immunoprecipitation with apoB-specific antibodies or density gradient ultracentrifugation. The concentrations of VLDL, LDL , and HDL in all species were determined after fractionation by densit y gradient ultracentrifugation. The ratio (VLDL + LDL)/HDL was calcula ted for humans (1.92), pig (1.40), cow (1.04), monkey (0.91), sheep (0 .65), cat (0.47), horse (0.44), rat (0.41), rabbit (0.32), dog (0.26), and mouse (0.25). Therefore, in four (dog, horse, rat, and mouse) out of six species (dog, horse, rat, mouse, cat, rabbit) with low ratios of below 0.5, apoB mRNA was edited in liver. In contrast, none of the species with high amounts of apoB-containing plasma lipoproteins expre ssed apoB mRNA editing in liver. Taken together, these results indicat e that i) editing of apoB mRNA is not intestine-specific but is also f ound in liver of many mammalian species, and ii) editing of apoB mRNA in liver appears to be one important genetic determinant for plasma co ncentrations of apoB-containing lipoproteins.