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Figure 3. MIXL1 up regulates c-REL expression to enhance anti apoptotic gene transcription(A) MIXL1-expressing clones show enhanced transcript levels for c-REL, BCL2A1, and BCL2L1. Quantitative RT-PCR results show the differences in c-REL, BCL2A1, and BCL2L1 expression levels between the U937 control, 1MIXL, and 2MIXL cells. Expression was normalized to 18S rRNA transcript levels. Error bars represent standard deviation between triplicates. *p < 0.05. (B) ChIP localizes endogenous MIXL1 to c-REL promoter in KG1 cells. Quantitative genomic PCR analysis shows specific enrichment of endogenous MIXL1 immunoprecipitated with either N-terminal or C-terminal MIXL1 antibodies on the c-REL promoter whereas an internal locus within the c-REL gene showed no MIXL1 occupancy. Error bars represent standard deviation between triplicates. (C) Knockdown of MIXL1 decreased while enforced expression of c-REL increased c-REL, BCL2A1, and BCL2L1 transcript levels. MIXL1 shRNA lentivirus and c-REL retrovirus were transduced into KG1 cells. RT-qPCR was performed in triplicate on RNAs isolated 48 hours after transduction. Expression was normalized to 18S rRNA levels, and error bars represent standard deviation between triplicates. (D) c-REL over-expression rescues MIXL1 knockdown–mediated growth arrest in KG1 cells. Growth was measured by MTS assay every 24 hours over a 4-day period in KG1 cells transduced with MIXL1 shRNA lentivirus and c-REL retrovirus. Absorbance was normalized to that of a non-transfected control sample. *p < 0.05. Image published in: Raymond A et al. (2014) Copyright: © 2014 Raymond et al. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View

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