Downregulation of human telomerase reverse transcriptase through anti-C-myc siRNA in human colon cancer Colo 320 cells.
The c-Myc and human telomerase reverse transcriptase gene (hTERT) gene are frequently deregulated and overexpressed in malignancy. hTERT activity is induced by c-Myc and strategies designed to inhibit c-Myc expression in cancer cells may have considerable therapeutic value. We designed and used a short hairpin RNA to inhibit c-Myc expression in Colo 320 cells and validated its effect on cell proliferation. In this study, four c-Myc-shRNA expression vectors were constructed and introduced into Colo 320 cells. The effects of c-Myc silencing on tumor cell growth was assessed by soft agar assay and DNA synthesis and MTT experiments. The expressions of c-Myc and hTERT were also assessed by real-time reverse transcription-polymerase chain reaction and Western blot analysis. Upon transient transfection with plasmid encoding shRNA, it was found that expression of c-Myc and hTERT decreased in shRNA-transfected cells. The downregulation of c-Myc and hTERT inhibited cell growth, shortened telomere lengths, and suppressed telomerase activity. In conclusion, our findings demonstrate that shRNA of c-Myc can inhibit the DNA replication in Colo 320 cells effectively and reduce telomere length and telomerase activity; therefore, it could be used as a new potential anticancer tool for therapy of human colon cancer.
Hu
Xiaoyun
X
Department of Adolescent Health and Maternal and Child Health Care, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, PR China.
Yu
Nancai
N
Fu
Lei
L
Su
Wen
W
Huang
Guofu
G
Wu
Yanxia
Y
Huang
Hanju
H
Huang
Hao
H
D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003110:Colonic Neoplasms / Q000628:therapy*; D015536:Down-Regulation; D016259:Genes, myc / Q000235:genetics*; D006801:Humans; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012320:RNA Polymerase III / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D019098:Telomerase / Q000235:genetics / Q000378:metabolism*; D016615:Telomere / Q000737:chemistry
D034741:RNA, Small Interfering; C509186:TERT protein, human; D019098:Telomerase; D012320:RNA Polymerase III