GATA1通过上调NANOG促进胰腺癌肿瘤干细胞形成
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摘要
目的:探究GATA1在胰腺癌肿瘤干细胞形成中的功能和作用机制。方法:通过流式细胞术检测GATA1对胰腺癌肿瘤干细胞形成的影响;通过实时荧光定量PCR和Western印迹筛选和验证GATA1下游的干性基因;通过双萤光素酶报告基因实验和染色质免疫共沉淀明确GATA1的调控机制。结果:GATA1过表达细胞株中肿瘤干细胞含量增加;GATA1上调NANOG的mRNA和蛋白表达水平;GATA1可以增强NANOG启动子的活性;GATA1结合在NANOG启动子-527～-524 bp处的GATA序列。结论:GATA1可以通过结合在NANOG启动子上激活其转录,促进胰腺癌肿瘤干细胞的形成。
Objective: To investigate the role of GATA1 in the regulation of pancreatic cancer stem cells and its mechanism. Methods: Flow cytometry assay was applied to determine the ratio of cancer stem cells in control and GATA1 overexpressed cells. Real-time PCR and Western blotting were used to screen and verify the stemness-related genes targeted by GATA1. Dual-luciferase reporter assay and chromatin immunoprecipitation were conducted to explore the specific mechanism in the regulation of NANOG by GATA1. Results: The ratio of cancer stem cells was upregulated in GATA1 overexpressed cells. The mRNA and protein level of NANOG were upregulated by GATA1. GATA1 can enhance the transcriptional activity of NANOG promotor by binding to its GATA sequence on-527～-524 bp site. Conclusion: GATA1 upregulates NANOG expression by binding to its promoter and promotes the stemness of pancreatic cancer cells.
Objective: To investigate the role of GATA1 in the regulation of pancreatic cancer stem cells and its mechanism. Methods: Flow cytometry assay was applied to determine the ratio of cancer stem cells in control and GATA1 overexpressed cells. Real-time PCR and Western blotting were used to screen and verify the stemness-related genes targeted by GATA1. Dual-luciferase reporter assay and chromatin immunoprecipitation were conducted to explore the specific mechanism in the regulation of NANOG by GATA1. Results: The ratio of cancer stem cells was upregulated in GATA1 overexpressed cells. The mRNA and protein level of NANOG were upregulated by GATA1. GATA1 can enhance the transcriptional activity of NANOG promotor by binding to its GATA sequence on-527～-524 bp site. Conclusion: GATA1 upregulates NANOG expression by binding to its promoter and promotes the stemness of pancreatic cancer cells.
引文
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[26]Zbinden M,Duquet A,Lorente-Trigos A,et al.NANOG regulates glioma stem cells and is essential in vivo acting in a cross-functional network with GLI1 and p53[J].EMBO J,2010,29(15):2659-2674.
[27]Wilkinson-White L,Lester K L,Ripin N,et al.GA-TA1 directly mediates interactions with closely spaced pseudopalindromic but not distantly spaced double GA-TA sites on DNA[J].Protein Sci,2015,24(10):1649-1659.
[28]Lu Y,Zhu H,Shan H,et al.Knockdown of Oct4and Nanog expression inhibits the stemness of pancreatic cancer cells[J].Cancer Lett,2013,340(1):113-123.
[2]孙可欣,郑荣寿,张思维,等.2015年中国分地区恶性肿瘤发病和死亡分析[J].中国肿瘤,2019,28(1):1-11.
[3]Zhou P,Li B,Liu F,et al.The epithelial to mesenchymal transition(EMT)and cancer stem cells:implication for treatment resistance in pancreatic cancer[J].Mol Cancer,2017,16(1):52-63.
[4]Batlle E,Clevers H.Cancer stem cells revisited[J].Nat Med,2017,23(10):1124-1134.
[5]Zhan H X,Xu J W,Wu D,et al.Pancreatic cancer stem cells:new insight into a stubborn disease[J].Cancer Lett,2015,357(2):429-437.
[6]van Schaijik B,Davis P F,Wickremesekera A C,et al.Subcellular localisation of the stem cell markers OCT4,SOX2,NANOG,KLF4 and c-MYC in cancer:a review[J].J Clin Pathol,2018,71(1):88-91.
[7]Chambers I,Colby D,Robertson M,et al.Functional expression cloning of Nanog,a pluripotency sustaining factor in embryonic stem cells[J].Cell,2003,113(5):643-655.
[8]Zhang J,Espinoza L A,Kinders R J,et al.NANOGmodulates stemness in human colorectal cancer[J].Oncogene,2013,32(37):4397-4405.
[9]Noh K H,Kim B W,Song K H,et al.Nanog signaling in cancer promotes stem-like phenotype and immune evasion[J].J Clin Invest,2012,122(11):4077-4093.
[10]Meng A,Tang H,Yuan B,et al.Positive and negative cis-acting elements are required for hematopoietic expression of zebrafish GATA-1[J].Blood,1999,93(2):500-508.
[11]Li Y,Ke Q,Shao Y,et al.GATA1 induces epithelialmesenchymal transition in breast cancer cells through PAK5 oncogenic signaling[J].Oncotarget,2015,6(6):4345-4356.
[12]Chang Z,Zhang Y,Liu J,et al.GATA1 promotes gemcitabine resistance in pancreatic cancer through antiapoptotic pathway[J].J Oncol,2019,2019:9474273.
[13]Bonnet D,Dick J E.Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell[J].Nat Med,1997,3(7):730-737.
[14]Lapidot T,Sirard C,Vormoor J,et al.A cell initiating human acute myeloid leukaemia after transplantation into SCID mice[J].Nature,1994,367(6464):645-648.
[15]Al-Hajj M,Wicha M S,Benito-Hernandez A,et al.Prospective identification of tumorigenic breast cancer cells[J].Proc Natl Acad Sci USA,2003,100(7):3983-3988.
[16]Hemmati H D,Nakano I,Lazareff J A,et al.Cancerous stem cells can arise from pediatric brain tumors[J].Proc Natl Acad Sci USA,2003,100(25):15178-15183.
[17]Patrawala L,Calhoun T,Schneider-Broussard R,et al.Highly purified CD44+prostate cancer cells from xenograft human tumors are enriched in tumorigenic and metastatic progenitor cells[J].Oncogene,2006,25(12):1696-1708.
[18]Szotek P P,Pieretti-Vanmarcke R,Masiakos P T,et al.Ovarian cancer side population defines cells with stem cell-like characteristics and Mullerian Inhibiting Substance responsiveness[J].Proc Natl Acad Sci USA,2006,103(30):11154-11159.
[19]Li C,Heidt D G,Dalerba P,et al.Identification of pancreatic cancer stem cells[J].Cancer Res,2007,67(3):1030-1037.
[20]Hermann P C,Huber S L,Herrler T,et al.Distinct populations of cancer stem cells determine tumor growth and metastatic activity in human pancreatic cancer[J].Cell Stem Cell,2007,1(3):313-323.
[21]Sancho P,Alcala S,Usachov V,et al.The everchanging landscape of pancreatic cancer stem cells[J].Pancreatology,2016,16(4):489-496.
[22]Ercan G,Karlitepe A,Ozpolat B.Pancreatic cancer stem cells and therapeutic approaches[J].Anticancer Res,2017,37(6):2761-2775.
[23]Gong S,Li Q,Jeter C R,et al.Regulation of NANOG in cancer cells[J].Mol Carcinog,2015,54(9):679-687.
[24]Iv Santaliz-Ruiz L E,Xie X,Old M,et al.Emerging role of nanog in tumorigenesis and cancer stem cells[J].Int J Cancer,2014,135(12):2741-2748.
[25]Zhou J J,Chen R F,Deng X G,et al.Hepatitis C virus core protein regulates NANOG expression via the stat3 pathway[J].FEBS Lett,2014,588(4):566-573.
[26]Zbinden M,Duquet A,Lorente-Trigos A,et al.NANOG regulates glioma stem cells and is essential in vivo acting in a cross-functional network with GLI1 and p53[J].EMBO J,2010,29(15):2659-2674.
[27]Wilkinson-White L,Lester K L,Ripin N,et al.GA-TA1 directly mediates interactions with closely spaced pseudopalindromic but not distantly spaced double GA-TA sites on DNA[J].Protein Sci,2015,24(10):1649-1659.
[28]Lu Y,Zhu H,Shan H,et al.Knockdown of Oct4and Nanog expression inhibits the stemness of pancreatic cancer cells[J].Cancer Lett,2013,340(1):113-123.
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