Lactogenic hormone activation of Stat5 and transcription of the beta-casein gene in mammary epithelial cells is independent of p42 ERK2 mitogen-activated protein kinase activity
Lactogenic hormone activation of Stat5 and transcription of the beta-casein gene in mammary epithelial cells is independent of p42 ERK2 mitogen-activated protein kinase activity
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DOI：
10.1074/jbc.271.50.31863
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发表时间：
1996-12-13
影响因子：
4.8
通讯作者：
Hynes, NE
中科院分区：
文献类型：
--
作者：
Wartmann, M;Cella, N;Hynes, NE
HC11乳腺上皮细胞已被用于表征乳蛋白基因表达调控的分子事件，乳原激素催乳素、胰岛素和糖皮质激素对HC11细胞的处理导致β -酪蛋白基因的转录，催乳素诱导一个信号事件，涉及乳腺因子Stat5的酪氨酸磷酸化，Stat5是信号转导和转录激活因子家族的成员（Stat）。我们发现HC11细胞表达两种Stat5蛋白，Stat5a和Stat5b。从磷酸盐标记的HC11细胞中免疫沉淀的Stat5a和Stat5b的磷酸肽和磷酸氨基酸分析显示，这两种蛋白都在丝氨酸上被组成性磷酸化，乳原激素处理导致Stat5蛋白中出现酪氨酸磷酸化肽，与此观察结果一致，对Stat5a和Stat5b的Western blot分析显示，乳原激素诱导了快速的，磷酸酪氨酸的短暂增加与Stat5在β -酪蛋白基因启动子中的同源识别序列的结合平行，乳原激素处理HC11细胞也导致有丝分裂原活化蛋白（MAP）激酶途径的快速激活。我们使用特异性MAP激酶抑制剂PD98059检测了该途径在β -酪蛋白转录中的作用。浓度的PD98059完全消除了乳原诱导的MAP激酶激活，但并未影响Stat5的磷酸化状态、其DNA结合活性或β -酪蛋白报告结构的转录激活，这表明MAP激酶途径与乳原激素诱导β -酪蛋白基因无关。
HC11 mammary epithelial cells have been used to characterize molecular events involved in the regulation of milk protein gene expression, Treatment of HC11 cells with the lactogenic hormones prolactin, insulin, and glucocorticoids results in transcription of the beta-casein gene, Prolactin induces a signaling event which involves tyrosine phosphorylation of the mammary gland factor, Stat5, a member of the family of signal transducers and activators of transcription (Stat). Here we show that HC11 cells express two Stat5 proteins, Stat5a and Stat5b. Phosphopeptide and phosphoamino acid analysis of Stat5a and Stat5b immunoprecipitated from phosphate-labeled HC11 cells revealed that both proteins were constitutively phosphorylated on serine, Lactogenic hormone treatment resulted in the appearance of a tyrosine-phosphorylated peptide in both Stat5 proteins, Consistent with this observation, a Western blot analysis of Stat5a and Stat5b showed that lactogenic hormones induced a rapid, transient increase in phosphotyrosine which paralleled the binding of Stat5 to its cognate recognition sequence in the beta-casein gene promoter, Lactogenic hormone treatment of the HC11 cells also led to a rapid activation of the mitogen activated protein (MAP) kinase pathway, We examined the role of this pathway in beta-casein transcription using a specific MAP kinase kinase inhibitor, PD98059, Concentrations of PD98059 which completely abrogated lactogen-induced MAP kinase activation did not affect the phosphorylation state of Stat5, its DNA binding activity, or transcriptional activation of a beta-casein reporter construct, This indicates that the MAP kinase pathway does not contribute to lactogenic hormone induction of the beta-casein gene.