A critical BCL6-related feedback loop explains the unique phenotype of germinal center B-cells and their malignant transformation.

RANUNCOLO, SM; POLO, JM; DIEROV, J; CARROLL, M; MELNICK, AM

Orlando, Florida.

Congreso; 48th Annual Meeting of the American Society of Hematology; 2006

American Society of Hematology

The BCL6 (B-Cell-Lymphoma-6) transcriptional repressor is a critical oncogene in B-cell lymphomas and is required for establishment of germinal centers by normal B-cells. However, the mechanisms by which BCL6 licenses germinal center formation and lymphomagenesis remain unknown. To characterize this mechanism we identified BCL6 target genes by expression arrays and high throughput chromatin immunoprecipitation. Remarkably, a number of these target genes were critical mediators of DNA damage sensing checkpoints including ATR and p53. Therefore, we hypothesized that BCL6 could attenuate DNA damage response by silencing these genes, which is likely a critical attribute for survival and proliferation of germinal center B-cells undergoing somatic hypermutation (SHM) and class switch recombination (CSR). Accordingly, we found that expression of BCL6 in primary naïve B cells isolated from human tonsils could block cellular sensing of DNA damage as demonstrated by loss of histone 2AX (H2AX) phosphorylation and delayed repair of double strand breaks. Repression of ATR (but not p53 or other targets) was required for this phenotype. Reciprocally, shRNA knockdown of BCL6 in B-cell lymphomas rescued repression of ATR, enhanced H2AX phosphorylation and accelerated repair of double strand breaks, independent of the status of p53. shRNA knockdown of BCL6 caused a marked increase of apoptosis in lymphoma cells in response to DNA damage, due to restored DNA damage checkpoint functions. Cell cycle profile analysis showed that BCL6 suppresses an S-phase checkpoint possible due to ATR repression. Importantly, BCL6 knockdown had an identical effect on ATR levels, H2AX phosphorylation, DNA damage, and survival in purified primary human tonsilar germinal center centroblasts. These results suggest that a major role of BCL6 in germinal center formation is to attenuate cellular response to DNA damage occurring as a byproduct of CSR and SHM. The same mechanism also seems to be required for lymphomagenesis, since we observed that sustained BCL6 expression in human primary mature B-cells leads to aberrant survival properties and genomic instability. Moreover, BCL6 blockade using a specific inhibitor molecule designed by our lab induces apoptosis in lymphoma cells and synergizes with DNA damaging agents. Therefore, we have identified a critical mechanism of action of the BCL6 oncoprotein in normal and pathogenic states and show that specific targeting of BCL6 could synergize with chemotherapy drugs for the therapy of B-cell lymphomas.