Characterization of hotspot exonuclease domain mutations in the DNA polymerase ϵ gene in endometrial cancer.
Characterization of hotspot exonuclease domain mutations in the DNA polymerase ϵ gene in endometrial cancer.
复制标题
DOI：
10.3389/fonc.2022.1018034
复制
发表时间：
2022
影响因子：
4.7
通讯作者：
Wang, Huaying
中科院分区：
文献类型：
--
作者：
Tian, Wenjuan;Ji, Zhaodong;Wang, Jingshu;Meng, Jiao;Bi, Rui;Ren, Yulan;Shan, Boer;Yang, Gong;Wang, Huaying
关键词：
本研究旨在分析子宫内膜癌（EC）中DNA聚合酶β 1（POLE）基因的热点核酸外切酶结构域突变（EDMs），并探讨EDMs对肿瘤细胞行为和POLE催化活性的影响。 在来自EC患者的肿瘤组织样品中进行POLE测序以鉴定热点EDM。生物信息学工具用于选择潜在的致病性EDM。评估了EDM与患者临床结局的相关性。用野生型POLE或POLE变体转染EC细胞，以检查EDM对EC细胞行为的影响，包括细胞周期、迁移和侵袭。采用免疫共沉淀法获得FLAG标记的野生型和突变型Pol γ催化亚基，然后评估聚合酶和核酸外切酶活性。除了先前报道的P286 R和V411 L之外，R375 Q和P452 L被鉴定为EC的新的和有害的POLE热点EDM。EDM组患者的临床结局明显优于其他队列患者。与野生型POLE相比，POLE突变体的过表达促进了EC细胞的顺铂耐药、G 0/G1细胞周期阻滞以及细胞的迁移和侵袭。POLE变体的过表达显著增加了HEK 293 T细胞中3 '-OH的丰度，并上调了DNA错配修复基因的表达。与野生型相比，在HEK 293细胞中存在错配核苷酸的情况下，Pol突变体表现出受损的聚合酶和核酸外切酶能力。我们鉴定了DNA聚合酶R411基因中的热点核酸外切酶结构域突变，并鉴定了P286 R、V411 L、R375 Q和P452 L为子宫内膜癌中的致病性POLE热点EDM。这些热点EDM与体外子宫内膜癌细胞的恶性行为和患者的良好预后相关，表明POLE影响DNA复制和校对以外的广泛细胞过程。
This study was aimed to profile hotspot exonuclease domain mutations (EDMs) of the DNA polymerase ϵ gene (POLE) in endometrial cancer (EC) and to investigate the effects of EDMs on tumor cell behavior and catalytic activities of Polϵ. POLE sequencing was performed in tumor tissue samples from patients with EC to identify hotspot EDMs. Bioinformatics tools were used to select the potential pathogenic EDMs. The association of EDMs with the clinical outcomes of patients was assessed. EC cells were transfected with wildtype POLE or POLE variants to examine the effects of the EDMs on EC cell behavior, including cell cycle, migration, and invasion. Co-immunoprecipitation was employed to obtain FLAG-tagged wildtype and mutant catalytic subunits of Polϵ, followed by the assessment of polymerase and exonuclease activities. In addition to previously reported P286R and V411L, R375Q and P452L were identified as novel, and deleterious POLE hotspot EDMs of EC. Patients in EDM group had significantly better clinical outcomes than the rest of the cohort. Compared with wildtype POLE, overexpression of POLE variants promoted cisplatin resistance, G0/G1 cell cycle arrest, and cell migration and invasion in EC cells. Overexpression of POLE variants significantly increased the abundance of 3’-OH and upregulated the expression of DNA mismatch repair genes in HEK293T cells. Compared with wildtype Polϵ, Pol ϵ mutants exhibited undermined polymerase and exonuclease abilities in the presence of mismatched nucleotides in HEK293 cells. We characterized the of hotspot exonuclease domain mutations in the DNA polymerase ϵ gene and identified P286R, V411L, R375Q, and P452L as pathogenic POLE hotspot EDMs in endometrial cancer. These hotspot EDMs are associated with the malignant behavior of endometrial cancer cells in vitro and favorable prognosis in patients, suggesting that POLE affects a wide range of cellular processes beyond DNA replication and proofreading.
登录
查看更多内容
影响因子：
7
作者：
Shinbrot E;Henninger EE;Weinhold N;Covington KR;Göksenin AY;Schultz N;Chao H;Doddapaneni H;Muzny DM;Gibbs RA;Sander C;Pursell ZF;Wheeler DA
通讯作者：
Wheeler DA
影响因子：
6.2
作者：
Billingsley, Caroline C.;Cohn, David E.;Mutch, David G.;Stephens, Julie A.;Suarez, Adrian A.;Goodfellow, Paul J.
通讯作者：
Goodfellow, Paul J.
影响因子：
11.5
作者：
McConechy, Melissa K.;Talhouk, Aline;McAlpine, Jessica N.
通讯作者：
McAlpine, Jessica N.
影响因子：
--
作者：
Cosgun, Erdal;Oh, Min
通讯作者：
Oh, Min
影响因子：
14.8
作者：
通讯作者：
--