The HNF-1β-USP28-Claspin pathway upregulates DNA damage-induced Chk1 activation in ovarian clear cell carcinoma.
The HNF-1β-USP28-Claspin pathway upregulates DNA damage-induced Chk1 activation in ovarian clear cell carcinoma.
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DOI：
10.18632/oncotarget.24776
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发表时间：
2018-04-03
期刊：
影响因子：
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通讯作者：
Kobayashi H
中科院分区：
文献类型：
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作者：
Ito F;Yoshimoto C;Yamada Y;Sudo T;Kobayashi H
转录因子肝细胞核因子1-β（HNF-1β）增强检查点激酶1（Chk 1）激活，并促进卵巢透明细胞癌（CCC）暴露于包括博来霉素在内的多种遗传毒性药物后的G2/M细胞周期进展。然而，导致HNF-1β检查点激活的潜在机制仍然在很大程度上未知。为了阐明HNF-1β对细胞周期检查点的影响，用靶向HNF-1β、Claspin、USP 28或对照载体的siRNA转染人CCC细胞系。通过免疫沉淀评估HNF-1β对Claspin蛋白的泛素化和稳定化。使用RNAi介导的基因沉默的功能丧失研究表明，在用遗传毒性剂博来霉素处理后，HNF-1β促进Claspin表达，导致磷酸化Chk 1（p-Chk 1）的积累并促进CCC细胞系的存活。本研究首次证实了对Claspin表达至关重要的去泛素化酶USP 28是HNF-1β的靶基因之一。内源性USP 28的敲低抑制Claspin表达和p-Chk 1活化以及细胞活力。我们的研究结果确定了HNF-1β― USP 28 ―Claspin― Chk 1轴在DNA损伤后检查点信号放大中的新途径。靶向这一途径可能代表了卵巢CCC的一种推定的、新的抗癌策略。
Transcription factor hepatocyte nuclear factor 1-beta (HNF-1β) enhances checkpoint kinase 1 (Chk1) activation and promotes G2/M cell cycle progression in ovarian clear cell carcinoma (CCC) following exposure to diverse genotoxic agents including bleomycin. However, the underlying mechanism leading to checkpoint activation of HNF-1β still remains largely unknown. To clarify the effects of HNF-1β on cell cycle checkpoints, human CCC cell lines were transfected with siRNAs targeting HNF-1β, Claspin, USP28, or a control vector. Ubiquitination and stabilization of Claspin protein by HNF-1β was assessed by immunoprecipitation. Loss-of-function studies using RNAi-mediated gene silencing indicated that HNF-1β facilitated the Claspin expression after treatment with a genotoxic agent bleomycin, resulting in accumulation of phosphorylated Chk1 (p-Chk1) and promotion of survival in CCC cell lines. This study showed for the first time that USP28, a de-ubiquitinase crucial for Claspin expression, is one target gene of HNF-1β. Knockdown of endogenous USP28 suppressed the Claspin expression and p-Chk1 activation and cell viability. Our findings identify a novel pathway of the HNF-1β―USP28―Claspin―Chk1 axis in checkpoint signal amplification in response to DNA damage. Targeting this pathway may represent a putative, novel, anticancer strategy in ovarian CCC.