Yamada-Mouri N, Hirata S, Kato J, Hoshi K
Department of Obstetrics and Gynecology, Yamanashi Medical University, Tamaho, Nakakoma, Japan.
J Steroid Biochem Mol Biol. 1997 Mar;60(5-6):325-9. doi: 10.1016/s0960-0760(96)00222-1.
Our previous study showed that a relatively high level of the aromatase mRNA existed in the cerebral cortex (CC) of the rat, where the aromatase activity was reported to be little or absent. To elucidate the identity of the aromatase mRNA in the CC of the rat, we investigated the 5'-region of the aromatase mRNA in the rat CC. When the sequence of the 5'-region of the cortical message was analysed by the 5'-rapid amplification of cDNA ends (5'-RACE) with the antisense primer for exon II using the RNA extracted from the CC, no clone could be isolated. However, the upstream sequence from the 5'-end of exon IV of the aromatase clones, isolated from the CC by RACE with the antisense primers for exon V, was different from that on the aromatase mRNA encoding the full translated region. The new sequence of the cortical type message, called the cortical type aromatase mRNA variant, was located on the intron upstream of exon IV in the genomic cDNA. Distribution of the brain aromatase message with exons III-V and the cortical type aromatase mRNA variant were analysed by the reverse transcription-polymerase chain reaction (RT-PCR) using total RNAs extracted from the hypothalamus-preoptic area (HPOA), amygdala (AMY) and CC. The PCR products with primers for exons III-V were generated from the HPOA and AMY, but not from the CC. On the other hand, the PCR products with primers for exon IIIv (cortical type aromatase mRNA variant specific)-V were detected in significant amounts in the CC as well as the HPOA and AMY. These results indicate the existence of the aromatase mRNA variant lacking exons I-III in the adult rat brain. This cortical type mRNA variant seemed to be widely distributed in the tissues.
我们之前的研究表明,大鼠大脑皮层(CC)中存在相对高水平的芳香化酶mRNA,而据报道该区域的芳香化酶活性很低或不存在。为了阐明大鼠CC中芳香化酶mRNA的特性,我们研究了大鼠CC中芳香化酶mRNA的5'区域。当使用从CC提取的RNA,通过外显子II的反义引物进行5'-cDNA末端快速扩增(5'-RACE)分析皮层信息的5'区域序列时,未分离到任何克隆。然而,通过外显子V的反义引物进行RACE从CC中分离出的芳香化酶克隆的外显子IV 5'端上游序列,与编码完整翻译区域的芳香化酶mRNA上的序列不同。这种被称为皮层型芳香化酶mRNA变体的皮层型信息的新序列,位于基因组cDNA中外显子IV上游的内含子上。使用从下丘脑-视前区(HPOA)、杏仁核(AMY)和CC提取的总RNA,通过逆转录-聚合酶链反应(RT-PCR)分析具有外显子III-V的脑芳香化酶信息和皮层型芳香化酶mRNA变体的分布。用外显子III-V引物产生的PCR产物来自HPOA和AMY,但不是来自CC。另一方面,用外显子IIIv(皮层型芳香化酶mRNA变体特异性)-V引物产生的PCR产物在CC以及HPOA和AMY中均大量检测到。这些结果表明成年大鼠脑中存在缺乏外显子I-III的芳香化酶mRNA变体。这种皮层型mRNA变体似乎在组织中广泛分布。