cAMP-mediated inhibition of the epithelial brush border Na+/H+ exchanger, NHE3, requires an associated regulatory protein
cAMP-mediated inhibition of the epithelial brush border Na+/H+ exchanger, NHE3, requires an associated regulatory protein
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DOI：
10.1073/pnas.94.7.3010
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发表时间：
1997-04-01
影响因子：
11.1
通讯作者：
Donowitz, M
中科院分区：
文献类型：
--
作者：
Yun, CHC;Oh, S;Donowitz, M
NHE 3是位于肠和肾刷状缘膜上的Na+/H+交换器，在那里它在跨上皮Na+吸收中起作用。刷状缘Na+吸收过程被cAMP依赖性蛋白激酶的激活所强烈抑制，但这种抑制作用的分子机制知之甚少。我们已经鉴定了两种调节蛋白，E3 KARP和NHERF，它们与NHE 3相互作用，使cAMP抑制NHE 3。NHE 3基因的两个调控蛋白在结构上相似，在氨基酸序列上约有50%的同源性。将NHE 3基因转染到PS120成纤维细胞或Caco-2细胞中，cAMP不能抑制NHE 3的活性。北方印迹分析表明，PS120和Caco-2细胞均不表达E3 KARP和NHERF。相反，其他cAMP抑制NHE 3的细胞系，包括OK、CHO和LLC-PK 1细胞，表达NHERF相关的调节蛋白。为了确定它们在NHE 3的cAMP依赖性抑制中的功能，将E3 KARP和NHERF转染到PS120/NHE 3成纤维细胞中。用NHERF或E3 KARP转染PS120/NHE 3成纤维细胞重建了cAMP诱导的NHE 3诱导，导致这些细胞中25-30%的抑制。
NHE3 is the Na+/H+ exchanger located on the intestinal and renal brush border membrane, where it functions in transepithelial Na+ absorption. The brush border Na+ absorptive process is acutely inhibited by activation of cAMP-dependent protein kinase, but the molecular mechanism of this inhibitory effect is poorly understood, We have identified two regulatory proteins, E3KARP and NHERF, that interact with NHE3 to enable cAMP to inhibit NHE3. The twee regulatory proteins are structurally related, sharing approximate to 50% identity In amino acid sequences, It has been previously shown that when NHE3 is transfected into PS120 fibroblasts or Caco-2 cells, cAMP failed to inhibit NHE3 activity, Northern blot analysis showed that both PS120 and Caco-2 cells lacked the expression of both E3KARP and NHERF. In contrast, other cell lines in which cAMP inhibits NHE3, including OK, CHO, and LLC-PK1 cells, expressed NHERF-related regulatory proteins, To determine their functions in cAMP-dependent inhibition of NHE3, E3KARP and NHERF were transfected into PS120/NHE3 fibroblasts. Transfection in PS120/NHE3 fibroblasts with either NHERF or E3KARP reconstituted cAMP-induced induction of NHE3, resulting in 25-30% inhibition in these cells.