Effects of WT1 gene isoform transfection on gene expression profiles in leukemia cell line NB4

Objective:To explore the potential effect of change in proportions of WT1 gene isoforms on gene expression profiles in acute promyelocytic leukemia cell line NB4. Methods:The eukaryotic expression recombinant vector (pCB6+/WTA) containing full-length of human WT1 isoform (WTA:-17AA/-KTS) cDNA and the pCB6+ vector alone were transfected into the leukemia cell line NB4 by electroperforation. The positive cell clones (NB4/WTA) were selected with G418. The WTA mRNA and protein in transfected cells were detected by reversed transcriptase polymerase chain reaction (RT-PCR) and western blotting, respectively. cDNA microarray was used to explore the effect of transfection of exogenous WT1 gene isoforms on gene expression profiles in leukemia NB4 cells. Expression of cyclinA1 and CDK7 genes were determined by semi-quantitative RT-PCR. Results:Exogenous WTA gene isoforms had stable expressions at mRNA and protein levels in NB4 cells after transfection. The gene expression profiles were changed by transfection of exogenous WTA gene isoforms into NB4 cells. Among 4,096 gene clones on the microarray, 89 (2.17%) genes underwent more than 2.0-fold changes and most of them (nearly 88.7%) were down-regulated. These genes are cell cycle-related genes,apoptosis-related genes,oncogenes,signal transduction-related genes, transport protein genes, etc. RT-PCR showed that cyclin A1 gene expression was up-regulated and CDK7 gene expression was down-regulated. Conclusion:The introduction and expression of exogenous WTA gene isoforms can alter the gene expression profiles in NB4 cells. Alteration in gene expression profiles had influence on biological behaviors such as growth, differentiation,and apoptosis of leukemia NB4/WTA cells.