Restoration of C/EBPalpha expression in a BCR-ABL+ cell line induces terminal granulocytic differentiation.
The transcription factor C/EBPalpha plays a critical role in the process of granulocytic differentiation. Recently, mutations that abrogated transcriptional activation of C/EBPalpha were detected in acute myeloid leukemia patient samples. Moreover, the progression of chronic myelogenous leukemia (CML) to blast crisis in patients was correlated with down-modulation of C/EBPalpha. The KCL22 cell line, derived from BCR-ABL+ CML in blast crisis, expressed wild-type C/EBPepsilon protein but not a functional C/EBPalpha, -beta, and -gamma. Restoration of C/EBPalpha expression in KCL22 cells triggered a profound proliferative arrest, a block in the G2/M phase of the cell cycle and a gradual increase in apoptosis. Within 3 days of inducing expression of C/EBPalpha, a remarkable neutrophilic differentiation of the KCL22 blast cells occurred as shown by morphologic changes, induction of expression of CD11b, primary, secondary, and tertiary granule proteins, and granulocyte colony-stimulating factor receptor. Using high density oligonucleotide microarrays, the gene expression profile of KCL22 cells stably transfected with C/EBPalpha was compared with that of empty vector, and we identified genes not previously known to be regulated by C/EBPalpha. These included the up-regulation of those genes important for regulation of hematopoietic stem cell homing, granulocytic differentiation, and cell cycle, whereas down-regulation occurred for genes coding for signaling molecules and transcription factors that are implicated in regulation of proliferation and differentiation of hematopoietic cells. Our study showed that restoration of C/EBPalpha expression in BCR-ABL+ leukemic cells in blast crisis is sufficient for rapid neutrophil differentiation suggesting a potential therapeutic role for ectopic transfer of C/EBPalpha in acute phase of CML.
D017209:Apoptosis; D015152:Blotting, Northern; D015153:Blotting, Western; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000096:biosynthesis* / Q000737:chemistry; D039481:CD11b Antigen / Q000096:biosynthesis; D002454:Cell Differentiation; D002460:Cell Line; D045744:Cell Line, Tumor; D002469:Cell Separation; D015536:Down-Regulation; D005434:Flow Cytometry; D016044:Fusion Proteins, bcr-abl / Q000378:metabolism*; D016195:G2 Phase; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006098:Granulocytes / Q000166:cytology; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008938:Mitosis; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D012313:RNA / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection; D015854:Up-Regulation
D022763:CCAAT-Enhancer-Binding Protein-alpha; D039481:CD11b Antigen; D009841:Oligonucleotides; D012313:RNA; D016044:Fusion Proteins, bcr-abl