Cyclic AMP differentially regulates cell proliferation of normal human keratinocytes through ERK activation depending on the expression pattern of B-Raf
Cyclic AMP differentially regulates cell proliferation of normal human keratinocytes through ERK activation depending on the expression pattern of B-Raf
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DOI：
10.1007/s00403-004-0478-z
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发表时间：
2004-04-01
影响因子：
3
通讯作者：
Iizuka, H
中科院分区：
文献类型：
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作者：
Takahashi, H;Honma, M;Iizuka, H
细胞外刺激增加的细胞内环磷酸腺苷（cAMP）可诱导多种生物效应，例如细胞增殖、分化和迁移。先前关于 cAMP 对角质形成细胞增殖的影响的报道是相互矛盾的，并且表明该影响显然取决于细胞密度。最近的研究表明，cAMP 信号通过调节丝裂原激活蛋白激酶 (MAPK) 活性来调节细胞增殖。然而，cAMP 影响角质形成细胞增殖和/或 cAMP 和 MAPK 信号通路之间串扰的精确机制仍有待确定。使用正常人角质形成细胞 (NHK)，我们研究了 cAMP 对角质形成细胞增殖的影响及其细胞密度方面的分子机制。在汇合的 NHK 中，环 AMP 以 Ras 独立和 Rap1 依赖性方式降低细胞外调节激酶 (ERK) 磷酸化和细胞增殖。 MEK-1 抑制剂 PD98059 可以阻断 cAMP 导致的细胞增殖减少。相反，在未汇合的 NHK 中，cAMP 增加了 ERK 磷酸化和细胞增殖。 Western blot分析显示NHK表达B-Raf和Rap-1。尽管 95 kDa 和 62 kDa B-Raf 亚型均在亚汇合 NHK 中表达，但在汇合 NHK 中仅检测到 62 kDa B-Raf。将 95 kDa B-Raf 转染至汇合的 NHK 中，导致 ERK 磷酸化和细胞增殖的 cAMP 依赖性增加。这些发现表明，B-Raf 亚型的差异表达对于依赖于 ERK 磷酸化的 NHK 增殖的 cAMP 依赖性调节至关重要。
Intracellular cyclic AMP (cAMP) increased by extracellular stimuli induces various biological effects, such as cell proliferation, differentiation, and migration. Previous reports regarding the effect of cAMP on keratinocyte proliferation are contradictory and indicate that the effect apparently depends on cellular density. Recent studies have revealed that cAMP signaling regulates cell proliferation by modulating mitogen-activated protein kinase (MAPK) activity. The precise mechanism by which cAMP affects keratinocyte proliferation and/or the crosstalk between the cAMP and MAPK signaling pathways, however, remain to be determined. Using normal human keratinocytes (NHK), we investigated the effect of cAMP on keratinocyte proliferation and its molecular mechanism in terms of cellular density. In confluent NHK, cyclic AMP decreased extracellular regulated kinase (ERK) phosphorylation and cell proliferation in a Ras-independent and Rap1-dependent manner. The decreased cell proliferation by cAMP was blocked by the MEK-1 inhibitor, PD98059. In contrast, in subconfluent NHK, cAMP increased ERK phosphorylation and cell proliferation. Western blot analysis revealed that NHK expressed B-Raf and Rap-1. Although both 95 kDa and 62 kDa B-Raf isoforms were expressed in subconfluent NHK, only 62 kDa B-Raf was detected in confluent NHK. Transfection of 95 kDa B-Raf into confluent NHK resulted in a cAMP-dependent increase in ERK phosphorylation and cell proliferation. These findings indicate that differential expression of B-Raf isoforms is critical for cAMP-dependent regulation of NHK proliferation that depends on phosphorylation of ERK.