5 NF-κB and AKT signaling prevent DNA damage by suppressing RAG1/2 | 133 the course of the experiments described, whereas the primary human (BCR-ABL-negative) B-ALL samples displayed a much greater induction of cell death (ranging from 50% to 80% specific cell death) upon treatment (data not shown). Our findings indicate that RAG expression induced after AKT and NF-kB inhibition does not coincide with cell cycle exit in Abl cells, which could have oncogenic potential because RAG activity during S phase is linked to lymphomagenesis.10 Consistent with this, our study shows that AKT and NF-kB inhibition leads to increased RAG-dependent DNA damage, most prominently in large cycling cells, which may provoke genomic instability. In agreement, our LM-PCR analysis confirms that RAG is active in S phase under these conditions. Figure 7. Model indicating the potential mechanism by which IKKbi and AKTi stimulate RAG expression in pre-B cells. Arrows represent positive and/or stimulatory effects. Horizontal bars represent inhibitory and/or repressive effects. Erag, conserved transcriptional enhancer in the RAG locus. Inhibition of NF-kB and AKT also increased RAG protein expression in primary BCR-ABL-negative human B-ALL cells, similar to that in mouse Abl cells and human BCR-ABL-positive B-ALL cell lines, suggesting that these pathways are important for suppressing RAG expression in primary human leukemic cells. Moreover, NF-kB and AKT inhibition resulted in increased surface IgM expression on leukemic blasts, which suggests that these cells may have experienced RAG activity. In support, by reanalysis of gene expression data, we found that NF-kB negatively correlated with RAG expression in primary BCR-ABL-negative B-ALL patients, although it must be noted that the differences between
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