Transfection-mediated expression of a dominant cAMP-resistant phenotype in the opossum kidney (OK) cell line prevents parathyroid hormone-induced inhibition of Na-phosphate cotransport. A protein kinase-A-mediated event.
Transfection-mediated expression of a dominant cAMP-resistant phenotype in the opossum kidney (OK) cell line prevents parathyroid hormone-induced inhibition of Na-phosphate cotransport. A protein kinase-A-mediated event.
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负鼠肾 (OK) 细胞系中转染介导的显性 cAMP 抗性表型表达可防止甲状旁腺激素诱导的磷酸钠共转运抑制。
DOI：
10.1172/jci114860
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发表时间：
1990
期刊：
影响因子：
--
通讯作者：
Pollock,AS
中科院分区：
文献类型：
--
作者：
Segal,JH;Pollock,AS
PTH应答负鼠肾（OK）细胞系中的钠-磷酸盐共转运被PTH、cAMP和蛋白激酶C激活剂抑制。为了探测cAMP的作用，我们稳定转染OK细胞的表达载体的cAMP结合突变的鼠蛋白激酶A调节亚基。从转染细胞的cAMP结合蛋白的二维电泳表明突变的调节单位的20倍的过度表达。来自这些细胞的蛋白激酶A具有20倍增加的半最大活化所需的cAMP浓度，2.8 μ M对野生型细胞的0.15 μ M。在转染的细胞中，Na-磷酸盐共转运对高达1 mM 8-Br-cAMP和1 μ M PTH不敏感，而这些相同的激动剂在野生型细胞中引起显著的转运抑制。蛋白激酶C激活剂油酰-乙酰甘油和十四烷酰-佛波醇乙酸酯，这是在野生型细胞中标记的钠-磷酸盐共转运的影响，仍然存在，虽然减弱，在转染的突变体。随着传代时间的延长，cAMP不敏感表型恢复为野生型cAMP敏感性，尽管继续选择共转染的neo标志物。回复突变体细胞对蛋白激酶A的半数最大激活需要正常的cAMP，0.13 μ M，并且恢复了PTH和cAMP敏感的Na-磷酸盐共转运抑制。我们认为，一个完整的和正常的cAMP敏感的蛋白激酶A途径是PTH抑制OK细胞中磷酸钠共转运的绝对必要条件。图片
Sodium-phosphate cotransport in the PTH-responsive opossum kidney (OK) cell line is inhibited by PTH, cAMP, and activators of protein kinase C. In order to probe the role of cAMP, we stably transfected OK cells with an expression vector for a cAMP-binding mutation of the murine protein kinase A regulatory subunit. Two-dimensional electrophoresis of cAMP-binding proteins from transfected cells indicated a 20-fold overexpression of the mutant regulatory unit. Protein kinase A from these cells had a 20-fold increase in the concentration of cAMP required for half-maximal activation, 2.8 microM vs. 0.15 microM for wild type cells. In the transfected cells, Na-phosphate cotransport was insensitive to up to 1 mM 8-Br-cAMP and 1 microM PTH, while these same agonists caused a significant inhibition of transport in the wild type cells. The effects on Na-phosphate cotransport of the protein kinase C activators oleoyl-acetyl glycerol and tetradecanoyl-phorbol acetate, which were marked in the wild type cells, were still present, although attenuated, in the transfected mutants. With prolonged passage, the cAMP-insensitive phenotype reverted to wild type cAMP sensitivity despite continued selection for the cotransfected neo marker. The revertant cells had a normal cAMP requirement for half-maximal activation of protein kinase A, 0.13 microM, and the PTH and cAMP-sensitive inhibition of Na-phosphate cotransport was restored. We suggest that an intact and normally cAMP-sensitive protein kinase A pathway is an absolute requirement for PTH inhibition of Na-phosphate cotransport in the OK cell.Images
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DOI：
10.1073/pnas.86.13.4887
发表时间：
1989-07-01
影响因子：
11.1
作者：
MELLON, PL;CLEGG, CH;MCKNIGHT, GS
通讯作者：
MCKNIGHT, GS
环AMP依赖性蛋白激酶同工酶自磷酸化的研究。
影响因子：
4.8
作者：
U. Walter;I. Uno;A. Y. Liu;P. Greengard
通讯作者：
P. Greengard
DOI：
10.1016/s0021-9258(18)45175-7
发表时间：
1987-09
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
C. Clegg;L. A. Correll;G. Cadd;G. McKnight
通讯作者：
C. Clegg;L. A. Correll;G. Cadd;G. McKnight
DOI：
--
发表时间：
1985-03
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
G. Grynkiewicz;M. Poenie;Roger Y. TsienB
通讯作者：
G. Grynkiewicz;M. Poenie;Roger Y. TsienB
影响因子：
4.8
作者：
A. Teitelbaum;G. Strewler
通讯作者：
A. Teitelbaum;G. Strewler