Janus kinase 2 (JAK2) regulates prolactin-mediated chloride transport in mouse mammary epithelial cells through tyrosine phosphorylation of Na+-K+-2Cl- cotransporter.
Janus kinase 2 (JAK2) regulates prolactin-mediated chloride transport in mouse mammary epithelial cells through tyrosine phosphorylation of Na+-K+-2Cl- cotransporter.
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DOI：
10.1210/mend.14.12.0568
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发表时间：
2000-12
影响因子：
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通讯作者：
N. Selvaraj;E. C. Omi;G. Gibori;M. Rao
中科院分区：
文献类型：
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作者：
N. Selvaraj;E. C. Omi;G. Gibori;M. Rao
上皮氯离子（Cl-）转运是通过同向转运体（如Na+- k +- 2cl -共转运体（NKCC1））和氯离子通道（如囊性纤维化跨膜电导调节剂（CFTR））的协调作用实现的。作为一种分泌组织，乳腺上皮细胞是这种机制的明显候选者，但在乳腺上皮细胞中Cl-转运及其激素调节的描述很少。我们确定乳腺上皮细胞系HC11是否运输氯化物，以及这是否受PRL（一种已知刺激离子运输的激素）的调节。HC11细胞同时表达CFTR和NKCC1。暴露于PRL或PGE1可增加HC11细胞中的Cl-转运。NKCC1受体阻滞剂速尿和Cl-通道抑制剂2-羧酸二苯胺可以抑制这种反应。PRL作用的剂量和时间过程表明，PRL在1 mg /ml和刺激10 min时对Cl-转运的影响最大。对信号通路的检查表明，PRL对Cl-转运的影响并不涉及[Ca2+]i或MAP激酶活性的增加。RT-PCR分析表明，HC11细胞表达Janus kinase 1 （JAK1）、JAK2和信号转换器和转录激活因子5 （STAT5）的mRNA，但不表达JAK3。PRL处理HC11细胞增加STAT5的磷酸化。JAK2抑制剂AG490阻断STAT5和prl诱导的磷酸化，但不阻断pge1诱导的Cl-转运。NKCC1，而非CFTR，在HC11细胞中被酪氨酸磷酸化。PRL增强了NKCC1的酪氨酸磷酸化，JAK2抑制剂AG490减弱了这种作用。这些结果首次证明了酪氨酸磷酸化的NKCC1和PRL-JAK2级联在调节Cl-转运中的作用。
Epithelial chloride (Cl-) transport is achieved by the coordinated action of symporters such as the Na+-K+-2Cl- cotransporter (NKCC1) and chloride channels such as the cystic fibrosis transmembrane conductance regulator (CFTR). As a secretory tissue, mammary epithelial cells are obvious candidates for such mechanisms, but Cl- transport and its hormonal regulation have been poorly delineated in mammary epithelial cells. We determined whether the mammary epithelial cell line, HC11, transports chloride and whether this was regulated by PRL, a hormone known to stimulate ion transport. HC11 cells express both CFTR and NKCC1. Exposure to PRL or PGE1 increased Cl- transport in HC11 cells. This was inhibited by the NKCC1 blocker, furosemide, and by the Cl- channel inhibitor, diphenylamine 2-carboxylate. Dose and time course of PRL action indicate that PRL had maximal effect on Cl- transport at 1 microg/ml and at 10 min of stimulation. Examination of the signaling pathways suggests that the PRL effect on Cl- transport does not involve an increase in [Ca2+]i or MAP kinase activity. RT-PCR analyses indicate that HC11 cells express mRNA for Janus kinase 1 (JAK1), JAK2, and signal transducer and activator of transcription 5 (STAT5) but not for JAK3. PRL treatment of HC11 cells increased phosphorylation of STAT5. The JAK2 inhibitor AG490 blocked phosphorylation of STAT5 and PRL-induced, but not PGE1-induced, Cl- transport. NKCC1, but not CFTR, is tyrosine phosphorylated in HC11 cells. PRL enhanced tyrosine phosphorylation of NKCC1, and this effect was attenuated by the JAK2 inhibitor AG490. These results are the first demonstrations of a role for tyrosine phosphorylation of NKCC1 and of the PRL-JAK2 cascade in the regulation of Cl- transport.