Multiple mineralocorticoid response elements localized in different introns regulate intermediate conductance K+ (Kcnn4) channel expression in the rat distal colon.
Multiple mineralocorticoid response elements localized in different introns regulate intermediate conductance K+ (Kcnn4) channel expression in the rat distal colon.
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DOI：
10.1371/journal.pone.0098695
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Rajendran VM
中科院分区：
文献类型：
--
作者：
O'Hara B;Alvarez de la Rosa D;Rajendran VM
血浆醛固酮升高和中间电导K+(IK/Kcnn4)通道的表达增加在结肠中是联系在一起的。这一观察结果表明，Kcnn4基因的表达是通过醛固酮对其同源受体(即盐皮质激素受体MR)的作用来调控的。为了确定这一点，我们使用正常和醛固酮处理的大鼠结肠上皮细胞的染色质，采用染色质免疫沉淀(CHIP)方法鉴定假定转录起始点(TSS)上游20kb和下游10kb区域的MR反应元件(MRE)。在醛固酮大鼠中，MRE在跨越第一和第二内含子的大约5kb区域内免疫沉淀。这些区域被分别克隆到缺乏增强子活性的荧光素酶表达载体中。用MR和醛固酮处理后，将这些克隆导入HEK293T和CaCo2细胞，检测其体外增强活性。研究发现，至少有四个区域对MR和醛固酮有反应。利用生物信息学工具确定了两个含有MRE的区域。这些克隆在假定的MRE发生突变后失去了增强子活性，从而确立了MRE的功能。第三个和第四个克隆不包含任何生物信息学上明显的MRE。此外，它们在额外的亚克隆时失去了活性，这表明在亚克隆时分离的区域之间存在协同作用。这些结果证明了Kcnn4中存在内含子MRE，并表明多个内含子反应元件之间存在高度协同的相互作用。
An elevated plasma aldosterone and an increased expression of the intermediate conductance K+ (IK/Kcnn4) channels are linked in colon. This observation suggests that the expression of Kcnn4 gene is controlled through the action of aldosterone on its cognate receptor (i.e., mineralocorticoid receptor; MR). In order to establish this, we performed chromatin immunoprecipitation (ChIP) assay to identify the MR response elements (MREs) in a region that spanned 20 kb upstream and 10 kb downstream of the presumed transcription start site (TSS) using chromatin from the colonic epithelial cells of normal and aldosterone-treated rats. MREs were immunoprecipitated in an approximately 5 kb region that spanned the first and second introns in the aldosterone rats. These regions were individually cloned in luciferase-expression vector lacking enhancer activity. These clones were tested for enhancer activity in vitro by transfecting in HEK293T and CaCo2 cells with MR and aldosterone treatment. At least four regions were found to be responsive to the MR and aldosterone. Two regions were identified to contain MREs using bioinformatics tools. These clones lost their enhancer activity after mutation of the presumptive MREs, and thus, established the functionality of the MREs. The third and fourth clones did not contain any bioinformatically obvious MREs. Further, they lost their activity upon additional sub-cloning, which suggest cooperativity between the regions that were separated upon sub-cloning. These results demonstrate the presence of intronic MREs in Kcnn4 and suggest a highly cooperative interaction between multiple intronic response elements.
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DOI：
10.1152/ajpgi.00101.2010
发表时间：
2010-09-01
影响因子：
4.5
作者：
Kumar, N. S. Nanda;Singh, Satish K.;Rajendran, Vazhaikkurichi M.
通讯作者：
Rajendran, Vazhaikkurichi M.
影响因子：
30.8
作者：
Carroll, Jason S.;Meyer, Clifford A.;Brown, Myles
通讯作者：
Brown, Myles
DOI：
10.1152/ajpgi.00337.2002
发表时间：
2003-07-01
影响因子：
4.5
作者：
Joiner, WJ;Basavappa, S;Rajendran, VM
通讯作者：
Rajendran, VM
影响因子：
3.8
作者：
Hubler, TR;Scammell, JG
通讯作者：
Scammell, JG
影响因子：
13.6
作者：
Gaeggeler, HP;Gonzalez-Rodriguez, E;Rossier, BC
通讯作者：
Rossier, BC