Analysis on switching of alternative exons 1 causing aberrant expression of estrogen synthetase

我们已经证明，乳腺癌组织近端区域芳香化酶表达的升高与这些区域人类芳香化酶基因的组织特异性外显子1的切换显著相关。为了进一步研究人类乳腺癌组织中芳香化酶基因转录因子中芳香化酶mRNA升高和从外显子1b切换到外显子1c的原因，我们在培养的乳腺组织脂肪基质细胞中研究了各种因素对表达水平和选择偏好1的影响。在犊牛血清存在下培养的基质细胞中，从外显子1b转录出芳香化酶mRNA。然而，剥夺血清诱导芳香化酶mRNA的快速升高和芳香化酶转录本切换到细胞外显子1c。HepG2细胞和非恶性乳腺组织中的芳香酶mRNA也从外显子1b转录。然后，使用包含4个主要外显子1和启动子的新报告子，通过荧光启动子试验检测了HepG2细胞中负责外显子1b特异性利用的启动子区域。结果表明，决定细胞对1b外显子优先利用的转录元件位于1b外显子-255 ~ -1145的启动子区域。我们还观察到骨和血管组织中显著水平的免疫反应性芳香化酶蛋白和芳香化酶mRNA。由于这些组织中雌激素水平的下降与骨组织骨质疏松症和血管组织动脉硬化的原因有关，并且这些组织中的芳香化酶mRNA是从1b外显子转录的，芳香化酶基因外显子1的切换可能是这些疾病的部分原因。

We have shown that elevation of aromatase expression in the proximal regions to breast cancer tissues was significantly correlated with switching of tissue-specific exons 1 of human aromatase gene in these regions. To further investigate what causes elevation of the aromatase mRNA and switching from exon 1b to exon 1c in the transcription factors of the aromatase gene in human breast cancer tissues, the effects of various on the exoression levels and preference of alternative expns 1 were examined in cultured adipose stromal cells from breast tissues. The aromatase mRNA was transcribed from exon 1b in the stromal cells cultured in the presence of calf serum. However, deprivation of the serum induced rapid elevation of aromatase mRNA and switching of aromatase transcripts to exon 1c the cells. Aromatase mRNA in HepG2 cells as well as in non-malignant breast tissues was also transcribed from exon 1b.Then, the promoter region responsible for the exon 1b-specific utilization in HepG2 cells was examined by fluorometric promoter assay using a new reporter containing 4 major alternative exons 1 and promoters. The results suggested that transcriptional elements determining preferential utilization of exon 1b in the cells was located on the promoter region of exon 1b form -255 to -1145. We have also observed significant levels of immunoreactive aromatase proteins and aromatase mRNA in the bone and vascular tissues. As decreased levels of estrogen in these tissues are paid attention in connecion with causes of osteoporosis in bone tissues and arterial sclerosis in vascular tissues and aromatase mRNA in these tissues is transcribed from exon 1b, switching of alternative exons 1 of the aromatase gene might contribute to a part of causes in these diseases.