|Table of Contents|

HuR gene clone and construction of its eukaryotic expressional vector

Journal Of Modern Oncology[ISSN:1672-4992/CN:61-1415/R]

Issue:
2018 01
Page:
1-4
Research Field:
Publishing date:

Info

Title:
HuR gene clone and construction of its eukaryotic expressional vector
Author(s):
Zhang Kailin1Wang Shuhua1Shi Song1Rao Min2Sun Daquan1
1.Department of Biochemistry and Molecular Biology,Guizhou Medical University,Guizhou Guiyang 550025,China;2.Guizhou Provincial People's Hospital,Guizhou Guiyang 550000,China.
Keywords:
HuR genehepatocellular carcinoma celllipofectioneukaryotic expression
PACS:
R735.7
DOI:
10.3969/j.issn.1672-4992.2018.01.001
Abstract:
Objective:To construct the eukaryotic expression vector pEGFP-HuR and screen its stable transfected hepatocellular carcinoma cells.Methods:The coding region of HuR gene sequence was amplified from SMMC-7721 cells by reversing transcription polymerase chain reaction (RT-PCR).The fragment was inserted into green fluorescent protein (GFP) expression vector pEGFP-C1 digested by Xho I and EcoR I enzymes,the same as PCR product digestion.The recombinant plasmid was transfected into SMMC-7721 cells and the pEGFP-HuR stable transfected hepatocellular carcinoma cells were screened by G418 drug.The expression of HuR gene was detected by Western blot.Results:Both double digestion analysis by Xho I and EcoR I and DNA sequencing result verified that the pEGFP-HuR eukaryotic expression vector was successfully constructed,the HuR protein level in pEGFP-HuR stable transfection cells was obviously raised.Conclusion:The pEGFP-HuR eukaryotic expression vector was successfully constructed and HuR gene was significantly expressed in stable transfection cells,which provides the help for the study of HuR gene.

References:

[1]Kim HH,Abdelmohsen K,Lal A,et al.Nuclear HuR aecumulation through phosphorylation by Cdkl[J].Genes Dev,2008,22(13):1804-1815.
[2]Giaginis C,Alexandrou P,Delladetsima I,et al.Clinical significance of Hu-antigen receptor (HuR) and cyclooxygenase-2 (COX-2) expression in human malignant and benign thyroid lesions[J].Pathol Oncol Res,2016,22(1):189-196.
[3]Sun DQ,Wang Y,Liu DG.Cancer cell growth suppression by a 62nt AU-rich RNA from C/EBPβ 3'UTR through competitive binding with HuR[J].Biochem Biophys Res Commun,2012,426(1):122-128.
[4]Zhang J,Chen X.Posttranscriptional regulation of p53 and its targets by RNA-binding proteins[J].Curr Mol Med,2008,8(8):845-849.
[5]Quann EJ,Khwaja F,Djakiew D.The p38 MAPK pathway mediates aryl propionic acid induced messenger RNA stability of p75 NTR in prostate cancer cells[J].Cancer Res,2007,67(23):11402-11410.
[6]Kang MJ,Ryu BK,Lee MG,et al.NF-κB activates transcription of the RNA-binding factor HuR,via PI3K-AKT signaling,to promote gastric tumorigenesis[J].Gastroenterology,2008,135(6):2030-2042.
[7]Zhou Y,Chang R,Ji W,et al.Loss of scribble promotes Snail translation through translocation of HuR and enhances cancer drug resistance[J].J Biol Chem,2016,291(1):291-302.
[8]López E,López I,Sequí J,et al.Discovering and validating unknown phospho-sites from p38 and HuR protein kinases in vitro by phosphoproteomic and bioinformatic tools[J].J Clin Bioinforma,2011,1(1):16.
[9]Ko CY,Wang WL,Li CF,et al.IL-18-induced interaction between IMP3 and HuR contributes to COX-2 mRNA stabilization in acute myeloid leukemia[J].J Leukoc Biol,2016,99(1):131-141.
[10]Blanco FF,Preet R,Aguado A,et al.Impact of HuR inhibition by the small molecule MS-444 on colorectal cancer cell tumorigenesis[J].Oncotarget,2016,7(45):74043-74058.
[11]Kurosu T,Ohga N,Hida Y,et al.HuR keeps an angiogenic switch on by stabilising mRNA of VEGF and COX-2 in tumour endothelium[J].Br J Cancer,2011,104(5):819-829.
[12]Tiedje C,Ronkina N,Tehrani M,et al.The p38/MK2-driven exchange between tristetraprolin and HuR regulates AU-rich element-dependent translation[J].PLoS Genet,2012,8(9):e1002977.
[13]Kotta-Loizou I,Vasilopoulos SN,Coutts RH,et al.Current evidence and future perspectives on HuR and breast cancer development,prognosis,and treatment[J].Neoplasia,2016,18(11):674-688.
[14]Zhang Z,Huang A,Zhang A,et al.HuR promotes breast cancer cell proliferation and survival via binding to CDK3 mRNA[J].Biomed Pharmacother,2017,91:788-795.
[15]Badawi A,Hehlgans S,Pfeilschifter J,et al.Silencing of the mRNA-binding protein HuR increases the sensitivity of colorectal cancer cells to ionizing radiation through upregulation of caspase-2[J].Cancer Lett,2017,393:103-112.
[16]Muralidharan R,Babu A,Amreddy N,et al.Folate receptor-targeted nanoparticle delivery of HuR-RNAi suppresses lung cancer cell proliferation and migration[J].J Nanobiotechnology,2016,14(1):47.
[17]Chai Y,Liu J,Zhang Z,et al.HuR-regulated lncRNA NEAT1 stability in tumorigenesis and progression of ovarian cancer[J].Cancer Med,2016,5(7):1588-1598.
[18]Huang YH,Peng W,Furuuchi N,et al.Insights from HuR biology point to potential improvement for second-line ovarian cancertherapy[J].Oncotarget,2016,7(16):21812-21824.
[19]To KK,Leung WW,Ng SS.Exploiting a novel miR-519c-HuR-ABCG2 regulatory pathway to overcome chemoresistance in colorectal cancer[J].Exp Cell Res,2015,338(2):222-231.

Memo

Memo:
National Natural Science Foundation of China(No.81560390);国家自然科学基金(编号:81560390);贵州省科学技术基金(编号:黔科合J字[2014]2025号,黔科合J字[2017]1142号);贵阳医学院院基金(编号:017号,020号)
Last Update: 2017-11-30