THE MECHANISM OF GENE REGULATION BY EXTRA CELLULAR CALCIUM THROUGH CALCIUM-SENSING RECEPTOR

钙敏感受体对细胞外钙的基因调控机制

基本信息

批准号：
07671116
负责人：
OKAZAKI Tomoki
金额：
$ 1.6万
依托单位：
UNIVERSITY OF TOKYO
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

Through the specific binding of a negative calcium responsive element (nCaRE) to its binding protein (nCaREB) in response to extracellular Ca (Ca^2+_e), nCaRE-bearing genes, such as the human parathyroid hormone (PTH) gene, are negatively regulated by Ca^2+_e. The Ku antigen mediated negative gene regulation by Ca^2+_e by interacting with a redox factor protein, refl. Though sequence-nonspecific DNA binding activity of the ku antigen has been well characterized, the mechanism of its sequence-specific DNA binding remained obscure, Here, we report that the specific binding of the Ku antigen to another protein, refl, leads to DNA-protein complex formation with a novel sequence-specificity and thereby regulates gene expression. We next examined whether the recently-identified membrane calcium-sensing receptor was involved in this type of gene regulation. Though cultured cells transfected with the calcium-sensing receptor expression vector potentiated the binding between nCaRE and nCaREB as exhibited repressed nCaRE-bearing reporter activity, these findings were observed irrespective of the extracellular calcium concentrations.On the other hand, we have found that one of the nCaREs, oligo B,is very well conserved among such vasoactive genes as the vasopressin and atrial natriuretic polypeptide genes. Further, the oligo B in the former gene is conserved throughout evolution. We demonstrate that the binding between oligo B and its binding nuclear proteins including a redox factor 1 (refl) was reduced by hyperosmolarity generated by sodium chloride (NaCl) but not by urea. Such attenuated binding was reversed by dephosphorylating some of the nuclear proteins by a potato acid phosphatase, suggesting that NaCl treatment elicited phosphorylation of these nuclear proteins to weaken their binding activity to oligo B.Furthermore, these nuclear events let to hyperosmolarity-mediated transcriptional stimulation of the genes bearing this DNA element in the cultured cells.

通过负钙反应元件（NCARE）与其结合蛋白（NCAREB）的特异性结合，响应于细胞外Ca（Ca^2+_e），NCARE含有基因，例如人类甲状旁腺激素（PTH）基因，由Ca^2+_e负调控。 KU抗原通过与氧化还原因子蛋白相互作用REFL通过CA^2+_e介导的阴性基因调节。尽管KU抗原的序列非特异性DNA结合活性已经很好地表征，但其序列特异性DNA结合的机制仍然晦涩难懂，在这里，我们报告说，KU抗原与另一种蛋白质REFL的特定结合导致DNA蛋白质复合物与新型序列特异性序列形成，并以一种新颖的序列形成，并在其上进行了调节。接下来，我们检查了最近识别的膜钙感应受体是否参与了这种基因调节。 Though cultured cells transfected with the calcium-sensing receptor expression vector potentiated the binding between nCaRE and nCaREB as exhibited repressed nCaRE-bearing reporter activity, these findings were observed irrespective of the extracellular calcium concentrations.On the other hand, we have found that one of the nCaREs, oligo B,is very well conserved among such vasoactive genes as the vasopressin and atrial亚钠多肽基因。此外，在整个进化过程中，前基因中的寡核B均得到保守。我们证明了寡核B与其结合核蛋白之间的结合（包括氧化还原因子1（REFL））通过氯化钠（NaCl）产生的高渗透性（REFL）降低，但不是由尿素。 Such attenuated binding was reversed by dephosphorylating some of the nuclear proteins by a potato acid phosphatase, suggesting that NaCl treatment elicited phosphorylation of these nuclear proteins to weaken their binding activity to oligo B.Furthermore, these nuclear events let to hyperosmolarity-mediated transcriptional stimulation of the genes bearing this DNA element in the cultured cells.