BRCA1和BRCA2与DNA损伤修复及转录调控

乳腺癌和卵巢癌敏感基因BRCA1和BRCA2与同源重组,DNA损伤修复,胚胎生长,转录调控及遍在蛋白化有关,其中,BRCA1和BRCA2在DNA损伤修复和转录调控中功能的确定,将有助于探讨和阐明两者的肿瘤抑制功能及其机理,作者将综述近年来有关BRCA1和BRCA2在DNA损伤修复和转录调控中功能研究的最新进展。     

Intracellular Free Calcium Concentration and Cisplatin Resistance in Human Lung Adenocarcinoma A549 Cells

Human lung adenocarcinoma A₅₄₉ cells sensitive and A₅₄₉/DDP cells resistantto Cis-dichlorodiammine platinum[II] (cisplatin) exhibit differentintracellular free calcium and calcium fluorescence images labeled withFura-2/AM and Fluo-3/AM as judged by dual-excitation fluorescence assay,Miracal Imaging and Laser Scanning Confocal Microscopy (LSCM) of singlecells. The concentration of intracellular free calcium of the resistantA₅₄₉/DDP cells is one third that of the sensitive A₅₄₉ cells. The effluxof Rhodamine 123 in resistant A₅₄₉/DDP cells is faster than that insensitive A₅₄₉ cells. In addition, A₅₄₉/DDP cells have an increase ofPhosphatidylinositol 4-kinase (PtdIns 4-kinase) activity in the plasmamembrane. So it is tentatively suggested that the increase in PtIns4-kinase activity resulting from lower intracellular Ca²⁺concentration leads to an increase of its enzymaticproducts——PIP and PIP₂, which may stimulate the activity ofP-glycoprotein.     

Knockdown of HIF1A‐AS2 suppresses TRIM44 to protect cardiomyocytes against hypoxia‐induced injury

Myocardial infarction (MI) is a common cardiovascular disease characterized by an interruption of blood and oxygen supply to the heart, which results in gradual damage to the myocardial tissue and ultimately heart failure. The role of long non‐coding RNAs in the pathology of MI remains in its infancy, but has been implicated in MI and other heart conditions. For example, the expression of a non‐coding RNA hypoxia‐inducible factor 1α (HIF1A)‐antisense RNA 2 (HIF1A‐AS2) has previously been linked to coronary heart disease, however, whether HIF1A‐AS2 expression is also high in MI has not been addressed. Here, we report that HIF1A‐AS2 is upregulated in hypoxia‐treated human cardiomyocytes (HMCs) compared with normal cardiomyocytes, and that silenced HIF1A‐AS2 inhibited apoptosis and facilitated viability, migration, and invasion of HMCs. Our data suggested that in MI, HIF1A‐AS2 upregulation was associated with miR‐623, which promoted expression of tripartite motif containing 44 (TRIM44). Moreover, by upregulating TRIM44 we were able to remedy the HIF1A‐AS2 repression of apoptosis in HMCs. Thus, we conclude that cardiomyocytes can be protected against hypoxic‐treated injury by knockdown of HIF1A‐AS2, which suppresses TRIM44, and that HIF1A‐AS2 overexpression is a prognostic indicator of MI.     

晚期非小细胞肺癌组织中BRCA1、STMN1、RRM1的表达及其临床意义

目的：探讨晚期非小细胞肺癌（NSCLC）组织中DNA修复基因家族成员BRCA1、STMN1、RRM1的表达及其临床意义。方法：回顾性分析本院2009年1月至2012年1月86例经组织学或细胞学证实的IIIB／IV期非小细胞肺癌患者,以分支DNA-液相芯片法检测肿瘤标本的BRCA1、STMN1、RRM1基因mRNA表达,并对检测结果应用SPSS13．0进行统计分析。结果：BRCA1中高表达与患者的性别无明显相关性（x毫0．1003,P=0．7514）,STMN1中高表达与肿瘤的分化程度相关（卡方=18．3002,P=0．000）。分析基因mRNA的表达情况与患者的化疗有效率,提示BRCA1中高表达患者完全缓解0例、部分缓解23例、稳定17例、进展16例,低表达患者分别为0例、12例、14例、4例（P〉0．05）,而STMN1表达阳性患者与阴性患者的临床疗效分别为0例、14例、21例、16例和0例、21例、10例、4例（P〈0．05）;RRM1表达阳性患者与阴性患者的临床疗效分别为0例、17例、19例、20例和0例、18例、12例、0例（P〈0．05）。结论：通过对BRCA1、STMN1、RRM1基因mRNA表达的个体化治疗靶标检测,对患者的预后以及化疗疗效有一定的预测价值。     

LncRNA XIST promotes human lung adenocarcinoma cells to cisplatin resistance via let-7i/BAG-1 axis

Long noncoding RNAs (lncRNAs) have been identified as oncogenes or tumor suppressors that are involved in tumorigenesis and chemoresistance. LncRNA XIST expression is upregulated in several cancers, however, its biologic role in the development of the chemotherapy of human lung adenocarcinoma (LAD) has not been elucidated. This study aimed to observe the expression of LncRNA XIST in LAD and to evaluate its biologic role and clinical significance in the resistance of LAD cells to cisplatin. LncRNA XIST expression was markedly increased in cisplatin-resistant A549/DDP cells compared with parental A549 cells as shown by qRT-PCR. LncRNA XIST overexpression in A549 cells increased their chemosensitivity to cisplatin both in vitro and in vivo by protecting cells from apoptosis and promoting cell proliferation. By contrast, LncRNA XIST knockdown in A549/DDP cells decreased the chemoresistance. We revealed that XIST functioned as competing endogenous RNA to repress let-7i, which controlled its down-stream target BAG-1. We proposed that XIST was responsible for cisplatin resistance of LAD cells and XIST exerted its function through the let-7i/BAG-1 axis. Our findings suggested that lncRNA XIST may be a new marker of poor response to cisplatin and could be a potential therapeutic target for LAD chemotherapy.     

Retracted: Chemoresistance in gastric cancer is attributed to the overexpression of excision repair cross-complementing 1 (ERCC1) caused by microRNA-122 dysregulation

MicroRNAs are deemed as key regulators of gene expression. In particular, the elevated expression of excision repair cross-complementing 1 (ERCC1) significantly reduced the effectiveness of gastric cancer treatment by cisplatin (CDDP)-based therapies. In this paper, qRT-PCR and western blot were adopted to measure miR-122 and ERCC1 messenger RNA (mRNA) expression in all samples. Luciferase assay was carried out to verify the role of ERCC1 as a target of miR-122. The CCK-8 assay was carried out to study the effect of ERCC1 and miR-122 on cell survival and apoptosis. The results demonstrated that miR-122 expression was reduced in cisplatin-resistant gastric cancer. Using bioinformatic analysis, miR-122 was shown to target the 3′-UTR of human ERCC1. A dual-luciferase assay demonstrated that miR-122 downregulated ERCC1 expression, while the mutations in ERCC1 3′-UTR abolished its interaction with miR-122. Transfection of miR-122 mimics decreased the levels of ERCC1 mRNA and protein expression, while the transfection of miR-122 inhibitors increased the levels of both ERCC1 mRNA and protein expression. Furthermore, we found that overexpressed miR-122 promoted the proliferation of MKN74 cells and reduced their apoptotic by targeting ERCC1. In addition, the levels of miR-122 and ERCC1 were negatively correlated in gastric cancer samples. In summary, the reduced miR-122 expression may play an essential role in the induction of cisplatin-resistance by increasing ERCC1 expression.     

BRCA1 epigenetic inactivation predicts sensitivity to platinum-based chemotherapy in breast and ovarian cancer

Germline mutations in the BRCA1 or BRCA2 genes are associated with an increased risk of breast and ovarian cancer development. Both genes are involved in DNA repair, and tumors harboring genetic defects in them are thought to be more sensitive to DNA-damaging agents used in chemotherapy. However, as only a minority of breast and ovarian cancer patients carry BRCA1 or BRCA2 mutations, few patients are likely to benefit from these pharmacogenetic biomarkers. Herein, we show that, in cancer cell lines and xenografted tumors, BRCA1 CpG island promoter hypermethylation-associated silencing also predicts enhanced sensitivity to platinum-derived drugs to the same extent as BRCA1 mutations. Most importantly, BRCA1 hypermethylation proves to be a predictor of longer time to relapse and improved overall survival in ovarian cancer patients undergoing chemotherapy with cisplatin.     

BRCA1 epigenetic inactivation predicts sensitivity to platinum-based chemotherapy in breast and ovarian cancer

Germline mutations in the BRCA1 or BRCA2 genes are associated with an increased risk of breast and ovarian cancer development. Both genes are involved in DNA repair, and tumors harboring genetic defects in them are thought to be more sensitive to DNA-damaging agents used in chemotherapy. However, as only a minority of breast and ovarian cancer patients carry BRCA1 or BRCA2 mutations, few patients are likely to benefit from these pharmacogenetic biomarkers. Herein, we show that, in cancer cell lines and xenografted tumors, BRCA1 CpG island promoter hypermethylation-associated silencing also predicts enhanced sensitivity to platinum-derived drugs to the same extent as BRCA1 mutations. Most importantly, BRCA1 hypermethylation proves to be a predictor of longer time to relapse and improved overall survival in ovarian cancer patients undergoing chemotherapy with cisplatin.     

磷酸丝氨酸氨基转移酶1介导肺腺癌细胞粘附及其机制探究

摘要 目的：探究丝氨酸生物合成途径（SSP）关键酶磷酸丝氨酸氨基转移酶1（PSAT1）与肺腺癌细胞粘附的关系，并初步探讨其作用机制。方法：使用siRNA抑制PSAT1蛋白表达，观察肺腺癌细胞形态以及粘附变化，同时过表达PSAT1，反向观察PSAT1对肺腺癌细胞粘附的影响。初步探究其作用机制，采用免疫共沉淀-蛋白质谱法寻找与PSAT1直接相互作用的蛋白，筛选差异蛋白，并在过表达细胞体系中验证。结合临床公共数据库分析互作蛋白与患者预后关系。结果：发现敲低PSAT1引起肺腺癌细胞形态改变；敲低PSAT1抑制肺腺癌PC9、HCC827细胞粘附；过表达PSAT1增强PC9及HCC827细胞粘附；免疫共沉淀-蛋白质谱检测到2560个可能与PSAT1结合的蛋白，进一步通过免疫共沉淀-免疫印迹法验证PSAT1过表达使细胞中与间皮素（MSLN）结合显著上升；通过临床样本数据观察PSAT1与MSLN共同高表达的肺腺癌患者，其预后更差。结论：本文首次报道PSAT1可能通过与MSLN等蛋白-蛋白相互作用影响肺腺癌细胞粘附的新机制，提示PSAT1有望成为潜在抗肿瘤靶点，靶向其相互作用蛋白能为小分子抑制剂设计及患者个体化治疗提供新思路。     

Long noncoding RNA CCAT2 functions as a competitive endogenous RNA to regulate FOXC1 expression by sponging miR-23b-5p in lung adenocarcinoma

Long noncoding RNA (lncRNA) may regulate the process of tumor formation. Although lncRNA CCAT2 has been identified as a key point in many diseases, its pathophysiological mechanism in lung adenocarcinoma remains unknown. We measured the expression level of CCAT2 in lung adenocarcinoma cells and normal lung epithelial cell line BEAS-2B by quantitative real-time polymerase chain reaction (qRT-PCR). As well, cell migration and proliferation were detected by transwell detection and CCK8 assay. At the same time, the new target point of CCAT2 was confirmed with bioinformatics analysis and dual-luciferase reporter assay. In addition, potential mechanisms were studied by Western blot analysis and RNA immunoprecipitation (RIP) analysis. The expression of CCAT2 was upregulated obviously in lung adenocarcinoma cells. Cell function analysis showed that upregulation of CCAT2 significantly promoted cell proliferation and migration, and reduction of CCAT2 inhibited cell migration and proliferation. In addition, CCAT2 positively regulated the expression of FOXC1 by competitive binding with miR-23b-5p. These findings indicated that CCAT2 may act as a competitive endogenous RNA (ceRNA) to regulate FOXC1 expression by competitively binding miR-23b-5p in lung adenocarcinoma.     

OsVIL2 functions with PRC2 to induce flowering by repressing OsLFL1 in rice

Flowering is exquisitely regulated by both promotive and inhibitory factors. Molecular genetic studies with Arabidopsis have verified several epigenetic repressors that regulate flowering time. However, the roles of chromatin remodeling factors in developmental processes have not been well explored in Oryza sativa (rice). We identified a chromatin remodeling factor OsVIL2 (O. sativa VIN3‐LIKE 2) that promotes flowering. OsVIL2 contains a plant homeodomain (PHD) finger, which is a conserved motif of histone binding proteins. Insertion mutations in OsVIL2 caused late flowering under both long and short days. In osvil2 mutants OsLFL1 expression was increased, but that of Ehd1, Hd3a and RFT1 was reduced. We demonstrated that OsVIL2 is bound to native histone H3 in vitro. Chromatin immunoprecipitation analyses showed that OsVIL2 was directly associated with OsLFL1 chromatin. We also observed that H3K27me3 was significantly enriched by OsLFL1 chromatin in the wild type, but that this enrichment was diminished in the osvil2 mutants. These results indicated that OsVIL2 epigenetically represses OsLFL1 expression. We showed that OsVIL2 physically interacts with OsEMF2b, a component of polycomb repression complex 2. As observed from osvil2, a null mutation of OsEMF2b caused late flowering by increasing OsLFL1 expression and decreasing Ehd1 expression. Thus, we conclude that OsVIL2 functions together with PRC2 to induce flowering by repressing OsLFL1.