p53-dependent p21 mRNA elongation is impaired when DNA replication is stalled
p53-dependent p21 mRNA elongation is impaired when DNA replication is stalled
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DOI：
10.1128/mcb.01520-06
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发表时间：
2007-02-01
影响因子：
5.3
通讯作者：
Prives, Carol
中科院分区：
文献类型：
--
作者：
Mattia, Melissa;Gottifredi, Vanesa;Prives, Carol
我们之前曾报道过，当某些人类细胞系中的 DNA 复制被阻断时，p53 诱导其关键靶基因子集（包括 p21(WAF1/CIP1)）的能力就会受损。在这里，我们通过比较两种药物的作用来研究这种损害的原因：羟基脲 (HU)，它使细胞停滞在早期 S 期并损害 p21 的诱导；柔红霉素，它引起 G(2) 阻滞，并导致 p53 对 p21 的强烈激活。 HU 处理被证明可以抑制 p21 mRNA 转录而不是改变其 mRNA 稳定性。然而，染色质免疫沉淀分析表明，HU 既不影响 p53 结合，也不影响 p21 启动子内历史 H3 和 H4 的乙酰化。此外，在 HU 和柔红霉素处理后，TFIID/TATA 结合蛋白复合物和 RNA 聚合酶 II (RNA Pol II) 大亚基的募集是相同的。然而，相对于柔红霉素处理，用 HU 处理的细胞中 p21 基因的转录延伸显着受损，起始位点下游区域的 RNA Pol II 占用率减少就证明了这一点。同样，在施用 HU 后的 p21 下游区域中，RNA Pol H（Pol H-C-末端结构域丝氨酸 2P）的特异性磷酸化形式较少，这种磷酸化仅在聚合酶延伸 RNA 时发生。我们认为，虽然 DNA 复制检查点不太可能调节 p21 启动子起始复合物的组装，但它会向参与转录延伸过程的一个或多个因子发出信号。
We have previously reported that when DNA replication is blocked in some human cell lines, p53 is impaired in its ability to induce a subset of its key target genes, including p21(WAF1/CIP1). Here, we investigated the reason for this impairment by comparing the effects of two agents, hydroxyurea (HU), which arrests cells in early S phase and impairs induction of p21, and daunorubicin, which causes a G(2) block and leads to robust activation of p21 by p53. HU treatment was shown to inhibit p21 mRNA transcription rather than alter its mRNA stability. Nevertheless, chromatin immunoprecipitation assays revealed that HU impacts neither p53 binding nor acetylation of histories H3 and H4 within the p21 promoter. Furthermore, recruitment of the TFIID/TATA-binding protein complex and the large subunit of RNA polymerase II (RNA Pol II) are equivalent after HU and daunorubicin treatments. Relative to daunorubicin treatment, however, transcription elongation of the p21 gene is significantly impaired in cells treated with HU, as evidenced by reduced occupancy of RNA Pol II at regions downstream of the start site. Likewise, in the p21 downstream region after administration of HU, there is less of a specifically phosphorylated form of RNA Pol H (Pol H-C-terminal domain serine 2P) which occurs only when the polymerase is elongating RNA. We propose that while the DNA replication checkpoint is unlikely to regulate the assembly of a p21 promoter initiation complex, it signals to one or more factors involved in the process of transcriptional elongation.