Identification of direct transcriptional targets of V600EBRAF/MEK signalling in melanoma
Identification of direct transcriptional targets of V600EBRAF/MEK signalling in melanoma
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DOI：
10.1111/j.1755-148x.2009.00618.x
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发表时间：
2009-12-01
影响因子：
4.3
通讯作者：
Marais, Richard
中科院分区：
文献类型：
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作者：
Packer, Leisl M.;East, Philip;Marais, Richard
P>BRAF 的致癌突变在黑色素瘤中很常见，并驱动 MEK/ERK 通路的组成性激活。为了阐明 V600EBRAF/MEK 信号下游的转录事件，我们对用 V600EBRAF 和 MEK 的有效选择性抑制剂（分别为 PLX4720 和 PD184352）处理的 A375 黑色素瘤细胞进行了基因表达谱分析。使用严格的贝叶斯方法，我们鉴定了 69 个转录物，它们似乎是该途径的直接转录靶标，并且其表达在途径抑制 6 小时后发生变化。我们还鉴定了几个额外的基因，其表达在途径抑制 24 小时后发生变化，并且可能是该途径的间接转录靶标。通过使用 RNA 干扰证明当 BRAF 耗尽时它们的表达受到类似的调节，以及通过在其他 BRAF 突变的黑色素瘤系中使用 qRT-PCR，证实了其中的一些。其中许多基因是转录因子和 ERK 通路的反馈抑制剂，并且在 NRAS 突变细胞中也受到 MEK 信号传导的调节。这项研究为了解黑色素瘤细胞中 V600EBRAF/MEK 信号传导调节的分子过程提供了基础。
P>Oncogenic mutations in BRAF are common in melanoma and drive constitutive activation of the MEK/ERK pathway. To elucidate the transcriptional events downstream of V600EBRAF/MEK signalling we performed gene expression profiling of A375 melanoma cells treated with potent and selective inhibitors of V600EBRAF and MEK (PLX4720 and PD184352 respectively). Using a stringent Bayesian approach, we identified 69 transcripts that appear to be direct transcriptional targets of this pathway and whose expression changed after 6 h of pathway inhibition. We also identified several additional genes whose expression changed after 24 h of pathway inhibition and which are likely to be indirect transcriptional targets of the pathway. Several of these were confirmed by demonstrating their expression to be similarly regulated when BRAF was depleted using RNA interference, and by using qRT-PCR in other BRAF mutated melanoma lines. Many of these genes are transcription factors and feedback inhibitors of the ERK pathway and are also regulated by MEK signalling in NRAS mutant cells. This study provides a basis for understanding the molecular processes that are regulated by V600EBRAF/MEK signalling in melanoma cells.