Sequence-specific DNA binding activity of RNA helicase A to the p16INK4a promoter
Sequence-specific DNA binding activity of RNA helicase A to the p16INK4a promoter
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DOI：
10.1074/jbc.m004481200
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发表时间：
2001-01-12
影响因子：
4.8
通讯作者：
Baylin, SB
中科院分区：
文献类型：
--
作者：
Myöhänen, S;Baylin, SB
p16（INK 4a）在人类癌症中经常改变，通常通过表观遗传学介导的转录沉默。然而，很少有人知道这个基因的转录调控。为了更多地了解这种控制，我们开始研究与癌细胞中的启动子结合的蛋白质，这些蛋白质表达和不表达该基因。我们鉴定RNA解旋酶A（RHA）为在表达细胞中更好地结合p16（INK 4a）启动子的蛋白质。RHA以前没有被表征为表现出序列特异性DNA相互作用，但对p16（INK 4a）启动子中的序列5' CGG ACC GCG TGC GC 3'表现出序列特异性DNA相互作用。果蝇同源RHA，无雄性（Mle），在果蝇中的功能是激活雄性X染色体基因。在我们的实验环境中，RHA诱导p16（INK 4a）启动子的类似适度上调，这取决于其序列特异性相互作用。Mle与超乙酰化H4 Ac 16共定位在X染色体和一些常染色体基因座上。在我们的细胞中，RHA与p16（INK 4a）的结合减少，其中该基因是转录失活的，与乙酰化赖氨酸抗体免疫沉淀的RHA量减少有关。最后，我们显示RHA是caspase-3的细胞底物，其通过N末端的切割降低其与p16（INK 4a）的序列特异性结合。因此，我们已经确定了一个新的蛋白质相互作用的p16（INK 4a）启动子，涉及一个重要的蛋白质转录调节。这种相互作用在癌细胞中减少，其中该基因在转录上异常沉默。
p16(INK4a) is frequently altered in human cancer, often through epigenetically mediated transcriptional silencing. However, little is known about the transcriptional regulation of this gene. To learn more about such control, we initiated studies of proteins that bind to the promoter in cancer cells that do, and do not, express the gene. We identify RNA helicase A (RHA) as a protein that binds much better to the p16(INK4a) promoter in the expressing cells. RHA has not previously been characterized to manifest sequence-specific DNA interaction but does so to the sequence 5' CGG ACC GCG TGC GC 3' in the p16(INK4a) promoter. The Drosophila homologue to RHA, maleless (Mle), functions in the fly for a-fold activation of male X-chromosome genes. In our experimental setting, RHA induces a similar modest up-regulation of the p16(INK4a) promoter that is dependent upon its sequence-specific interaction. Mle colocalizes with hyperacetylated H4Ac16 on the X-chromosome and some autosomal loci. The decreased binding of RHA to p16(INK4a) in our cells, where the gene is transcriptionally inactive, is associated with decreased amounts of RHA that immunoprecipitate with acetylated lysine antibodies. Finally, we show RHA to be a cellular substrate for caspase-3, which decreases its sequence-specific binding to p16(INK4a) by cleavage of the N terminus. Thus, we have identified a new protein interaction with the p16(INK4a) promoter that involves an important protein for transcriptional modulation. This interaction is decreased in cancer cells, where this gene is aberrantly transcriptionally silent.