STAT3-Activated GM-CSFRa Translocates to the Nucleus and Protects CLL Cells from Apoptosis

Here, it was determined that chronic lymphocytic leukemia (CLL) cells express the a subunit, but not the b subunit, of the granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR/CSF2R). GM-CSFRa was detected on the surface, in the cytosol, and in the nucleus of CLL cells via confocal microscopy, cell fractionation, and GM-CSFRa antibody epitope mapping. Because STAT3 is frequently activated in CLL and the GM-CSFRa promoter harbors putative STAT3 consensus binding sites, MM1 cells were transfected with truncated forms of theGM-CSFRa promoter, then stimulatedwith IL6 to activate STAT3 and to identify STAT3binding sites. Chromatin immunoprecipitation (ChIP) and an electoromobility shift assay (EMSA) confirmed STAT3 occupancy to those promoter regions in both IL6-stimulated MM1 and CLL cells. Transfection of MM1 cells with STAT3-siRNA or CLL cells with STAT3-shRNA significantly downregulated GM-CSFRamRNA and protein levels. RNA transcripts, involved in regulating cell survival pathways, and the proteinsKAP1 (TRIM28) and ISG15 coimmunoprecipitated with GM-CSFRa. GM-CSFRa–bound KAP1 enhanced the transcriptional activity of STAT3, whereas GM-CSFRa-bound ISG15 inhibited the NF-kB pathway. Nevertheless, overexpression of GM-CSFRa protected MM1 cells from dexamethasone-induced apoptosis, and GM-CSFRa knockdown induced apoptosis in CLL cells, suggesting that GM-CSFRa provides a ligand-independent survival advantage. Implications: Constitutively, activation of STAT3 induces the expression of GM-CSFRa that protects CLL cells from apoptosis, suggesting that inhibition of STAT3 or GM-CSFRamay benefit patients with CLL.Mol Cancer Res; 1–16. 2014 AACR.