Effects of RNA interference on NPM-ALK fusion gene expression in anaplastic large-cell lymphoma cells
10.3969/j.issn.1008-8830.2005.03.003
- VernacularTitle:RNAi阻断NPM-ALK基因表达及对大细胞间变性淋巴瘤细胞的影响
- Author:
ZHAO YAN-XIA
1
;
GU LONG-JUN
;
YE QI-DONG
;
ZHAO JIN-CAI
Author Information
1. 上海第二医科大学附属新华医院
- Keywords:
RNA interference;
RNA,small interfering;
Gene fusion;
NPM-ALK;
Anaplastic large-cell lymphoma
- From:
Chinese Journal of Contemporary Pediatrics
2005;7(3):202-206
- CountryChina
- Language:Chinese
-
Abstract:
Objective To evaluate two small interfering RNAs (siRNAs) on the NPM-ALK fusion gene expression in anaplastic large-cell lymphoma cell line Karpas299, and to study the effect of RNA interference on Karpas299 cells proliferation. Methods Two siRNAs sequences (siRNA- I and siRNA-II) were designed to target the NPM-ALK fusion site in anaplastic large-cell lymphoma cell line Karpas299. An siRNA U6 expression system including U6 RNA-based polymerase III promoter was set up. The two siRNAs designed for down-regulation of the NPM-ALK fusion mRNA were transfected into Karpas299 cells by liposomal transfection reagents. The effect of RNAi on NPM-ALK mRNA expression was detected by real-time RT-PCR and Western blot. The anti-proliferative effects of the siRNA U6 system were assessed using MTT. Apoptosis was observed by fluorescence microscopy. Results The mRNA level of NPM-ALK in Karpas299 cells transfected with siRNA-I and siRNA-II decreased by approximately 75% and 35% respectively. The NPM-ALK protein expression was inhibited in Karpas299 cells at 72 hrs of siRNA-I transfection. The siRNA-II treatment had no effect on NPM-ALK protein expression. siRNA-I had inhibitory effects on Karpas299 cells proliferation and induced the cells apoptosis, while siRNA-II did not. Conclusions Sequence specific siRNAs targeting NPM-ALK was capable of suppressing NPM-ALK expression and inhibiting cellular proliferation. RNA interference may be a suitable technique for studying the function of NPM-ALK gene and may be used to develop siRNA-based targeted gene therapeutic approaches against NPM-ALK-positive lymphomas.
