Iñiguez M A, Morte B, Rodriguez-Peña A, Muñoz A, Gerendasy D, Sutcliffe J G, Bernal J
Instituto de Investigaciones Biomédicas, CSIC, Facultad de Medicina, UAM, Madrid, Spain.
Brain Res Mol Brain Res. 1994 Dec;27(2):205-14. doi: 10.1016/0169-328x(94)90002-7.
RC3 encodes a thyroid hormone-dependent, calmodulin-binding, protein kinase C substrate (neurogranin, p17) present in the dendritic spines of discrete neuronal populations in the forebrain. Its physiological role could be related to synaptic plasticity, memory, and other processes. In the present work we have isolated and sequenced 2.4 kbp of genomic DNA upstream from the origin of transcription and determined its nucleotide sequence. The major features of the RC3 promoter are the absence of TATA and CAAT boxes and the presence of an Initiator sequence surrounding the cap site. By sequence analysis we identified several cis-acting regulatory elements, among them response elements for retinoic acid and steroid (glucocorticoids/progesterone) hormone receptors. An oligonucleotide containing the retinoic acid responsive element bound to retinoic acid receptors specifically in vitro and conferred retinoic acid regulation to a heterologous promoter after transfection in COS-7 cells. Retinoic acid and dexamethasone, respectively, increased activity of the RC3 promoter in neuroblastoma cells when a deletion construct containing the retinoic acid and the glucocorticoid responsive elements was cotransfected with retinoic acid receptor or glucocorticoid receptor expression vectors. When added together all-trans retinoic acid and dexamethasone had additive effects. Despite the fact that RC3 expression in vivo is thyroid hormone-dependent, no evidence for the presence of a thyroid hormone responsive element was found within the 2.4 kbp flanking region analyzed and thyroid hormone did not increase reporter activity after cotransfection of suitable constructs with thyroid hormone receptor expression vectors. Our results suggest that the expression of RC3 in vivo could be subject to complex physiological signals, including retinoids and steroid hormones in addition to thyroid hormones.
RC3编码一种甲状腺激素依赖性、钙调蛋白结合蛋白激酶C底物（神经颗粒蛋白，p17），存在于前脑离散神经元群体的树突棘中。其生理作用可能与突触可塑性、记忆及其他过程有关。在本研究中，我们分离并测序了转录起始点上游2.4kbp的基因组DNA，并确定了其核苷酸序列。RC3启动子的主要特征是缺乏TATA盒和CAAT盒，且在帽位点周围存在一个起始子序列。通过序列分析，我们鉴定了几个顺式作用调节元件，其中包括视黄酸和类固醇（糖皮质激素/孕酮）激素受体的反应元件。含有视黄酸反应元件的寡核苷酸在体外能特异性地与视黄酸受体结合，并在转染到COS-7细胞后赋予异源启动子视黄酸调节作用。当含有视黄酸和糖皮质激素反应元件的缺失构建体与视黄酸受体或糖皮质激素受体表达载体共转染时，视黄酸和地塞米松分别增加了神经母细胞瘤细胞中RC3启动子的活性。当全反式视黄酸和地塞米松一起添加时，具有相加作用。尽管RC3在体内的表达是甲状腺激素依赖性的，但在所分析的2.4kbp侧翼区域内未发现甲状腺激素反应元件的证据，并且在将合适的构建体与甲状腺激素受体表达载体共转染后，甲状腺激素并未增加报告基因的活性。我们的结果表明，RC3在体内的表达可能受到复杂生理信号的影响，除甲状腺激素外，还包括类视黄醇和类固醇激素。
