Effect of silencing bmi-1 by RNA interference on function of K562 cell line.
- Author:
Xiao-Li CHEN
1
;
Qian REN
;
Zhen-Ping CHEN
;
Ze-Ping ZHOU
;
Qin-Jun ZHAO
;
Zhi-Yong QIU
;
Chun-Lan DONG
;
Zhong-Chao HAN
Author Information
1. Institute of Hematology and Blood Disease Hospital, Chinese Academy of Medical Sciences, Tianjin 300020, China.
- Publication Type:Journal Article
- MeSH:
Cell Proliferation;
Cell Survival;
Genetic Vectors;
Humans;
K562 Cells;
Leukemia, Myelogenous, Chronic, BCR-ABL Positive;
genetics;
Nuclear Proteins;
genetics;
Polycomb Repressive Complex 1;
Proto-Oncogene Proteins;
genetics;
RNA Interference;
RNA, Small Interfering;
genetics;
Repressor Proteins;
genetics;
Transfection
- From:
Journal of Experimental Hematology
2009;17(2):266-270
- CountryChina
- Language:Chinese
-
Abstract:
Bmi-1 is a transcriptional repressor, which belongs to the polycomb group family. It has been demon- started that over-expression of Bmi-1 occurs in a variety of cancers, including several types of leukemia. Bmi-1 gene plays a key role in regulation of self-renewal in normal and leukemic stem cells. Acute myeloid leukemic cells lacking Bmi-1 undergo proliferation arrest and show signs of differentiation and apoptosis, which leads to the proposal of Bmi-1 as a potential target for therapeutic intervention in leukemia. The purpose of this study was to investigate the effect of short hairpin RNA (shRNA) targeting Bmi-1 on functions of K562 cell line. The shRNA eukaryotic expression vector targeting Bmi-1 was constructed and transfected into K562 cells through lipofectamine 2000. The mRNA and protein levels of Bmi-1 were detected by PCR and Western blot respectively. The proliferation of K562 after Bmi-1 silencing was measured by using MTT assay and clone formation assay. The cell cycle was detected by flow cytometry. The results indicated that among the four shRNA designed, there was a shRNA which efficiently interfered with the expression of Bmi-1. The results of PCR and Western blot validated that the Bmi-1 gene of K562 cells transfected with such a Bmi-1 shRNA was suppressed successfully. Although levels of Bmi-1 mRNA and protein were significantly reduced, delivery of this siRNAs had no effect on cell viability or growth. Flow cytometry analysis suggested that Bmi-1 inhibition did not affect the cell cycle. It is concluded that the suppression of Bmi-1 expression is not able to reduce proliferation of K562 cells, suggesting existence of some other parallel signaling pathways, which are fundamental for leukemic transformation and are independent of Bmi-1 over-expression. Bmi-1 over-expression may play a secondary role in chronic myeloid leukemia transformation.
