Transcriptional regulation of human angiotensin receptor

描述（由申请人提供）：八肽，血管紧张素-II（Ang II）是已知的最有效的血管活性物质之一，并且调节多种生理反应，包括流体稳态、醛固酮产生、肾功能和血管平滑肌（VSM）的收缩。血管紧张素受体-1（AT 1 R）基因的过度表达可导致高血压，尤其是在雌性转基因小鼠中。这些研究表明，增加的hAT 1 R基因的转录可能会导致高血压。因此，我们分析了hAT 1 R基因5'侧翼区的单核苷酸多态性（SNP）在高血压中的作用。我们发现hAT 1 R基因启动子至少有5个单倍型区，包括T/A at - 777，T/G at-680，A/C at-214，G/C at-213和A/G at-119。我们的研究表明，变体-777 T、-680 T、-214 A、-213 G和-119 A总是一起出现（分别产生含有TTAGA的单倍型-1和含有AGCCG的单倍型-11）。我们的研究表明，hAT 1 R基因的单体型-1与高加索妇女的高血压有关，并且与单体型-11相比，瞬时转染含有hAT 1 R基因的单体型-1的报告构建体在肾上腺皮质细胞和VSMC中具有增加的启动子活性。我们的凝胶迁移试验表明：（a）转录因子C/EBP更强地结合含有-119A的寡核苷酸，（单体型-1）与-119 G相比（单倍型-II）和（B）转录因子USF（其与E-box基序CANNTG结合）与在-214和-213处含有核苷A和G的寡核苷酸强烈结合（单倍型-I）与在-213和-214处含有核苷C的相同寡核苷酸（单倍型-II）相比。因此，我们将在使用肾上腺皮质和血管平滑肌细胞的体外系统中分析单倍型-I和II对hAT 1 R基因转录调节的影响，使用雄性和雌性转基因小鼠检测单倍型-I和II对hAT 1 R mRNA水平和对血压的影响，并检测IL-6、糖皮质激素、和雌激素对hAT 1 R基因单倍型I和II的转基因小鼠中hAT 1 R mRNA水平的影响，并将转基因小鼠中mRNA水平的潜在变化与血压相关联。

DESCRIPTION (provided by applicant): The octapeptide, angiotensin-ll (Ang II) is one of the most potent vasoactive substances known and regulates a variety of physiological responses, including fluid homeostasis, aldosterone production, renal function and contraction of vascular smooth muscle (VSM). Over-expression of the angiotensin receptor-1 (AT1R) gene produces hypertension especially in female transgenic mice. These studies have suggested that increased transcription of the hAT1 R gene may lead to hypertension. We have therefore analyzed the role of single nucleotide polymorphisms (SNPs) in the 5' flanking region of the hAT1R gene in hypertension. We have found that hAT1 R gene promoter has a haplotype block of at least five SNPs consisting of T/A at - 777, T/G at -680, A/C at -214, G/C at -213, and A/G at -119. Our studies have shown that variants -777T, - 680T, -214A, -213G, and -119A always occur together (creating haplotype-l containing TTAGA and haplotype-ll containing AGCCG respectively). Our studies have shown that haplotype-l of the hAT1R gene is associated with hypertension in Caucasian women and transient transfection of reporter construct containing haplotype-l of the hAT1R gene has increased promoter activity in adrenal cortical cells and VSMC as compared to haplotype-ll. Our gel shift assays have shown that: (a) transcription factor C/EBP binds more strongly to an oligonucleotide containing -119A (haplotype-l) as compared to -119G (haplotype- ll) and (b) transcription factor USF (which binds to an E-box motif CANNTG) binds strongly to an oligonucleotide containing nucleoside A and G at -214 and -213 (haplotype-l) as compared to the same oligonucleotide containing nucleoside C at -213 and -214 (haplotype-ll). We will therefore analyze the effect of haplotypes -I and II on transcriptional regulation of the hAT1R gene in an in-vitro system using adrenocortical and vascular smooth muscle cells, examine the effect of haplotypes-l and II on hAT1 R mRNA level and on blood pressure using male and female transgenic mice, and examine the effect of IL-6, glucocorticoids, and estrogens on hAT1R mRNA level in transgenic mice containing haplotype I and II of the hAT1 R gene, and to correlate potential changes in mRNA levels with blood pressure in transgenic mice.