Pape M E, Schultz P A, Rea T J, DeMattos R B, Kieft K, Bisgaier C L, Newton R S, Krause B R
Division of Therapeutics, Parke-Davis Pharmaceutical Research Division, Ann Arbor, MI 48105, USA.
J Lipid Res. 1995 Apr;36(4):823-38.
A human cDNA clone (K1) was recently isolated that encodes functional acyl-CoA:cholesterol acyltransferase (ACAT) protein (Chang et al. J. Biol. Chem. 1993. 268: 20747-20755). We used the K1 clone to screen a rabbit liver cDNA library and isolated a 919 base pair partial rabbit cDNA (ACAT14b) that was greater than 90% homologous with the human nucleotide sequence. Northern blotting using the rabbit ACAT cDNA14b revealed the existence of at least six related mRNA species (ranging from 6.2 to 1.7 kb) in various rabbit tissues. Using an RNAse protection assay, ACAT mRNA14b was detected in twelve separate rabbit organs. Adrenal gland contained the highest concentrations of ACAT mRNA14b (per microgram of total RNA) being 20-, 30-, and 50-fold higher than small intestine, aorta, and liver, respectively. Additional studies with isolated liver cell populations revealed that rabbit hepatic nonparenchymal cells contained 30-fold more ACAT mRNA14b (per microgram of total RNA) than parenchymal cells. To determine whether ACAT mRNA14b levels are regulated in vivo, rabbits were fed for 4 weeks a high fat/high cholesterol diet (HFHC; 0.5% cholesterol, 3% coconut oil, 3% peanut oil) at which point they were either kept for an additional 4 weeks on the HFHC-diet or switched to the HFHC-diet plus CI-976 (50 mg/kg), a potent and specific ACAT inhibitor; another group of rabbits was fed a chow diet for the entire 8 weeks. The HFHC-diet caused a 2- and 3-fold increase in hepatic and aortic ACAT mRNA14b levels, respectively, in comparison to chow-fed animals; there was no change in adrenal or small intestine levels. CI-976 treatment lowered ACAT mRNA14b levels by 60% and 40% in liver and aorta, respectively, in comparison to the HFHC controls; again there was no change in adrenal or small intestine levels. These data indicate that ACAT mRNA14b levels increase in a tissue specific manner in response to dietary fat and cholesterol.
最近分离出一个人类cDNA克隆（K1），它编码功能性酰基辅酶A：胆固醇酰基转移酶（ACAT）蛋白（Chang等人，《生物化学杂志》，1993年。268：20747 - 20755）。我们用K1克隆筛选兔肝cDNA文库，分离出一个919个碱基对的兔cDNA片段（ACAT14b），它与人类核苷酸序列的同源性超过90%。用兔ACAT cDNA14b进行Northern印迹分析，发现在兔的各种组织中存在至少六种相关的mRNA种类（范围从6.2到1.7 kb）。使用RNA酶保护试验，在兔的十二个不同器官中检测到ACAT mRNA14b。肾上腺中ACAT mRNA14b的浓度最高（每微克总RNA），分别比小肠、主动脉和肝脏高20倍、30倍和50倍。对分离的肝细胞群体进行的进一步研究表明，兔肝非实质细胞中ACAT mRNA14b的含量（每微克总RNA）比实质细胞多30倍。为了确定ACAT mRNA14b水平在体内是否受到调节，给兔子喂食高脂肪/高胆固醇饮食（HFHC；0.5%胆固醇，3%椰子油，3%花生油）4周，此时将它们要么再继续喂食HFHC饮食4周，要么换成HFHC饮食加CI - 976（50毫克/千克），一种强效且特异性的ACAT抑制剂；另一组兔子在整个8周内喂食普通饲料。与喂食普通饲料的动物相比，HFHC饮食使肝脏和主动脉中ACAT mRNA14b水平分别增加了2倍和3倍；肾上腺或小肠中的水平没有变化。与HFHC对照组相比，CI - 976处理使肝脏和主动脉中ACAT mRNA14b水平分别降低了60%和40%；肾上腺或小肠中的水平再次没有变化。这些数据表明，ACAT mRNA14b水平会以组织特异性的方式响应饮食中的脂肪和胆固醇而增加。
