肝癌SMMC-7721细胞系HSF4调控的靶基因图谱分析
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摘要
热休克转录因子4(heat shock transcription factor 4,HSF4)是热休克转录因子HSF家族成员,近年发现HSF4除发挥调控热休克蛋白(heat shock protein,HSP)表达的功能外,亦直接或间接调节许多非热休克基因的表达,参与细胞生长发育多种生理病理过程。现通过染色质免疫共沉淀联合测序(chromatin immunoprecipitation sequencing,ChIP-Seq)方法对HSF4潜在的靶基因进行检测,并用GO(gene ontology)分析方法对这些靶基因进行分析,以探求HSF4发挥的生物学功能。ChIP-Seq结果显示:在肝癌SMMC-7721细胞系基因组DNA中共有1 726个HSF4结合区域,其中102个在启动子区。GO分析结果显示:这些潜在的靶基因分别参与细胞发育、增殖和对外部刺激应答的生物过程,具有与核酸和蛋白质结合及蛋白质激活的分子功能,参与药物和有害异物代谢以及化学致癌的信号传导通路。此外,MEME4.12.0软件推测出在SMMC-7721细胞系中HSF4与DNA启动子区结合的6种模式。分析HSF4在肝癌SMMC-7721细胞系中可能调控的靶基因图谱为进一步研究HSF4在肝癌发生机制中的作用提供了有效的实验数据和理论基础。
Heat shock transcription factor 4(HSF4) is a member of HSF family. Recently, it was reported that HSF4 not only regulates the expression of heat shock protein(HSP), but also directly or indirectly controls many non-heat shock gene expressions, involved in cell growth and some physiological and pathological processes. The aim of this study was to clarify the target genes regulated by HSF4 in the hepatoma cell line SMMC-7721. The potential target genes of HSF4 were detected by chromatin immunoprecipitation sequencing(ChIP-Seq) method. Then they were analyzed by gene ontology(GO) to explore the biological function of HSF4.In ChIP-Seq analysis, 1 726 binding peaks were identified, and among them, 102 were located in the pro-moter region. The results of GO analysis indicated that these target genes were associated with cell develop-ment, proliferation and response to stimuli in biological processes. Some of these genes could bind proteins or nucleic acids and activate proteins. They were also involved in drug metabolism, xenobiotics metabolism and chemical carcinogenesis pathways. Furthermore, MEME4.12.0 software was used to identify the HSF4 binding DNA motifs, and three motifs were discovered for coding genes and the other three motifs for lnc R-NAs. These results clarify the target genes of HSF4, and provide the experimental basis for further exploring the roles of HSF4 in tumorigenesis of liver cancer.
Heat shock transcription factor 4(HSF4) is a member of HSF family. Recently, it was reported that HSF4 not only regulates the expression of heat shock protein(HSP), but also directly or indirectly controls many non-heat shock gene expressions, involved in cell growth and some physiological and pathological processes. The aim of this study was to clarify the target genes regulated by HSF4 in the hepatoma cell line SMMC-7721. The potential target genes of HSF4 were detected by chromatin immunoprecipitation sequencing(ChIP-Seq) method. Then they were analyzed by gene ontology(GO) to explore the biological function of HSF4.In ChIP-Seq analysis, 1 726 binding peaks were identified, and among them, 102 were located in the pro-moter region. The results of GO analysis indicated that these target genes were associated with cell develop-ment, proliferation and response to stimuli in biological processes. Some of these genes could bind proteins or nucleic acids and activate proteins. They were also involved in drug metabolism, xenobiotics metabolism and chemical carcinogenesis pathways. Furthermore, MEME4.12.0 software was used to identify the HSF4 binding DNA motifs, and three motifs were discovered for coding genes and the other three motifs for lnc R-NAs. These results clarify the target genes of HSF4, and provide the experimental basis for further exploring the roles of HSF4 in tumorigenesis of liver cancer.
引文
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[2]Pirkkala L,Nykanen P,Sistonen L.Roles of the heat shock transcription factors in regulation of the heat shock response and beyond[J].FASEB Journal,2001,15(7):1118-1131.
[3]Gomez P R,Burchfiel E T,Thiele D J.Regulation of heat shock transcription factors and their roles in physiology and disease[J].Nature Reviews.Molecular Cell Biology,2018,19(1):4-19.
[4]Nakai A,Tanabe M,Kawazoe Y,et al.HSF4,a new member of the human heat shock factor family which lacks properties of a transcriptional activator[J].Molecular and Cellular Biology,1997,17(1):469-481.
[5]Min J N,Zhang Y,Moskophidis D,et al.Unique contribution of heat shock transcription factor 4 in ocular lens development and fiber cell differentiation[J].Genesis,2004,40(4):205-217.
[6]Fujimoto M,Izu H,Seki K,et al.HSF4 is required for normal cell growth and differentiation during mouse lens development[J].The EMBO Journal,2004,23(21):4297-4306.
[7]Bu L,Jin Y,Shi Y,et al.Mutant DNA-binding domain of HSF4 is associated with autosomal dominant lamellar and Marner cataract[J].Nature Genetics,2002,31(3):276-278.
[8]Enoki Y,Mukoda Y,Furutanic C,et al.DNA-binding and transcriptional activities of human HSF4 containing mutations that associate with congenital and age-related cataracts[J].Biochimica et Biophysica Acta,2010,1802(9):749-753.
[9]Jin X,Eroglu B,Cho W,et al.Inactivation of heat shock factor Hsf4 induces cellular senescence and suppresses tumorigenesis in vivo[J].Molecular Cancer Research,2012,10(4):523-534.
[10]Huang M,Li D,Huang Y,et al.HSF4 promotes G1/S arrest in human lens epithelial cells by stabilizing p53[J].Biochimica et Biophysica Acta,2015,1853(8):1808-1817.
[11]Cui X,Zhang J,Du R,et al.HSF4 is involved in DNA damage repair through regulation of Rad51[J].Biochimica et Biophysica Acta-Molecular Basis of Disease,2012,1822(8):1308-1315.
[12]Miles R R,Crockett D K,Lim M S,et al.Analysis of BCL6-interacting proteins by tandem mass spectrometry[J].Molecular&Cellular Proteomics,2005,4(12):1898-1909.
[13]Hatzi K,Melnick A.Breaking bad in the germinal center:how deregulation of BCL6 contributes to lymphomagenesis[J].Trends in Molecular Medicine,2014,20(6):343-352
[14]Wei Z,Gao W,Wu Y,et al.Mutual interaction between BCL6and mi RNAs contributing to the pathogenesis of various cancers[J].Clinical and Translational Oncology,2015,17(11):841-846.
[15]Milarski K L,Morimoto R I.Expression of human HSP70 during the synthetic phase of the cell cycle[J].Proceedings of the National Academy of Sciences USA,1986,83(24):9517-9521.
[16]Gao M,Huang Y,Wang L,et al.HSF4 regulates lens fiber cell differentiation by activating p53 and its downstream regulators[J].Cell Death and Disease,2017,8(10):e3082.
[17]Jin X,Eroglu B,Moskophidis D,et al.Targeted deletion of Hsf1,2,and 4 genes in mice[J].Methods in Molecular Biology,2011,787:1-22.
[18]Liao S,Du R,Wang L,et al.BCAS2 interacts with HSF4 and negatively regulates its protein stability via ubiquitination[J].The International Journal of Biochemistry&Cell Biology,2015,68:78-86.
[19]Calderaoro J,Nault J C,Bioulac-Sage P,et al.ALDH3A1 is overexpressed in a subset of hepatocellular carcinoma characterised by activation of the Wnt/β-catenin pathway[J].Virchows Archiv,2014,464(1):53-60.
[20]Oraldi M,Saracino S,Maggoora M,et al.Importance of inverse correlation between ALDH3A1 and PPARγin tumor cells and tissue regeneration[J].Chemico-Biological Interactions,2011,191(1-3):171-176.
[21]Yan J,Demelo J,Cutz J C,et al.Aldehyde dehydrogenase 3A1associates with prostate tumorigenesis[J].British Journal of Cancer,2014,110(10):2593-2603.
[22]Benowitz N L,Sthelen G,Dempsey D A,et al.Disposition kinetics and metabolism of nicotine and cotinine in African American smokers:impact of CYP2A6 genetic variation and enzymatic activity[J].Pharmacogenetics and Genomics,2016,26(7):340-350.
[23]Egawa T.Runx and Th POK:a balancing act to regulate thymocyte lineage commitment[J].Journal of Cellular Biochemistry,2009,107(6):1037-1045.
[24]Rui J,Liu H,Zhu X,et al.Epigenetic silencing of Cd8 genes by Th POK-mediated deacetylation during CD4 T cell differentiation[J].The Journal of Immunology,2012,189(3):1380-1390.
[25]Mariani F,Sena P,Pedroni M,et al.Th inducing POZ-Kruppel Factor(Th POK)is a key regulator of the immune response since the early steps of colorectal carcinogenesis[J].PLo S One,2013,8(1):e54488.
[26]Xiao H,Lis J T.Germline transformation used to define key features of heat-shock response elements[J].Science,1988,239(4844):1139-1142.
[27]Lee T I,Rinaldi N J,Robert F,et al.Transcriptional regulatory networks in Saccharomyces cerevisiae[J].Science,2002,298(5594):799-804.
[28]Gwenael B,Esther T C,Harm V B,et al.A new library of yeast transcription factor motifs reveals a widespread function for Rsc3 in targeting nucleosome exclusion at promoters[J].Molecular Cell,2008,32(6):878-887.
[29]Reddy T E,De Lisi C,Shakhnovich B E.Binding site graphs:a new graph theoretical framework for prediction of transcription factor binding sites[J].PLo S Computational Biology,2007,3(5):e90.
[30]Harbison C T,Gordon D B,Lee T,et al.Transcriptional regulatory code of a eukaryotic genome[J].Nature,2004,431(7004):99-104.
[31]Bailey T L,Boden M,Buske F A,et al.MEME SUITE:tools for motif discovery and searching[J].Nucleic Acids Research,2009,37:W202-W208.