Establishment and utilization of a tetracycline-controlled inducible RNA interfering system to repress gene expression in chronic myelogenous leukemia cells.
RNA interference (RNAi), a posttranscriptional gene silencing process mediated by small double-stranded RNA specifically complementary to the targeted transcript, has been used extensively in the development of novel therapeutic approaches against various human diseases including chronic myelogenous leukemia (CML). Here, we report the successful construction of a tetracycline-controlled siRNA in CML cell line K562. A K562 cell line stably expressing the reverse tetracycline-controlled transactivator (rtTA) was constructed. A tetracycline responsive element (TRE) was integrated into the RNA polymerase III promoter region of pBS/U6 that was used to drive specific siRNA to target the novel cytokine receptor-like factor 3 (CRLF3) gene. The results show that rtTA was able to recognize the TRE to prevent siRNA-mediated exogenous and endogenous CRLF3 gene repressions. Moreover, CRLF3-siRNA mediated gene repression could be induced in a dose-dependent manner in the presence of doxycycline. Thus, the inducible siRNAi system in K562 cells might be useful for the study of RNAi-mediated therapeutic approaches against CML.
Yang
Fan
F
Department of Microbiology and Etiology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100005, China.
Zhang
Yun
Y
Cao
Ying-Li
YL
Wang
Shu-Hui
SH
Liu
Li
L
D001483:Base Sequence; D005786:Gene Expression Regulation; D018390:Gene Targeting; D005822:Genetic Vectors; D006801:Humans; D020014:K562 Cells; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012342:RNA, Small Nuclear; D018121:Receptors, Cytokine / Q000235:genetics; D020218:Response Elements; D013754:Tetracyclines / Q000494:pharmacology; D015534:Trans-Activators; D014162:Transfection
C507912:CRLF3 protein, human; D034741:RNA, Small Interfering; D012342:RNA, Small Nuclear; D018121:Receptors, Cytokine; D013754:Tetracyclines; D015534:Trans-Activators; C405486:U6 small nuclear RNA