RESUMO
Upregulated expression of the anti-apoptotic BCL2 oncogene is a common feature of various types of B-cell malignancies, from lymphoma to leukemia or myeloma. It is currently unclear how the various patterns of deregulation observed in pathology eventually impact the phenotype of malignant B cells and their microenvironment. Follicular lymphoma (FL) is the most common non-Hodgkin lymphoma arising from malignant germinal center (GC) B-cells, and its major hallmark is the t(14:18) translocation occurring in B cell progenitors and placing the BCL2 gene under the control of the immunoglobulin heavy chain locus regulatory region (IgH 3'RR), thus exposing it to constitutive expression and hypermutation. Translocation of BCL2 onto Ig light chain genes, BCL2 gene amplification, and other mechanisms yielding BCL2 over-expression are, in contrast, rare in FL and rather promote other types of B-cell lymphoma, leukemia, or multiple myeloma. In order to assess the impact of distinct BCL2 deregulation patterns on B-cell fate, two mouse models were designed that associated BCL2 and its full P1-P2 promoter region to either the IgH 3'RR, within a "3'RR-BCL2" transgene mimicking the situation seen in FL, or an Ig light chain locus context, through knock-in insertion at the Igκ locus ("Igκ-BCL2" model). While linkage to the IgH 3' RR mostly yielded expression in GC B-cells, the Igκ-driven up-regulation culminated in plasmablasts and plasma cells, boosting the plasma cell in-flow and the accumulation of long-lived plasma cells. These data demonstrate that the timing and level of BCL2 deregulation are crucial for the behavior of B cells inside GC, an observation that could strongly impact the lymphomagenesis process triggered by secondary genetic hits.
RESUMO
The MYC transcription factor regulates a vast number of genes and is implicated in many human malignancies. In some hematological malignancies, MYC is frequently subject to missense mutations that enhance its transformation activity. Here, we use a novel murine cell system to (i) characterize the transcriptional effects of progressively increasing MYC levels as normal primary B-cells transform to lymphoma cells and (ii) determine how this gene regulation program is modified by lymphoma-associated MYC mutations (T58A and T58I) that enhance its transformation activity. Unlike many previous studies, the cell system exploits primary B-cells that are transduced to allow regulated MYC expression under circumstances where apoptosis and senescence pathways are abrogated by the over-expression of the Bcl-xL and BMI1 proteins. In such cells, transition from a normal to a lymphoma phenotype is directly dependent on the MYC expression level, without a requirement for secondary events that are normally required during MYC-driven oncogenic transformation. A generalized linear model approach allowed an integrated analysis of RNA sequencing data to identify regulated genes in relation to both progressively increasing MYC level and wild type or mutant status. Using this design, a total of 7569 regulated genes were identified, of which the majority (n = 7263) were regulated in response to progressively increased levels of wild type MYC, while a smaller number of genes (n = 917) were differentially regulated, compared to wild type MYC, in T58A MYC- and/or T58I MYC-expressing cells. Unlike most genes that are similarly regulated by both wild type and mutant MYC genes, the set of 917 genes did not significantly overlap with known lipopolysaccharide regulated genes, which represent genes regulated by MYC in normal B cells. The genes that were differently regulated in cells expressing mutant MYC proteins were significantly enriched in DNA replication and G2 phase to mitosis transition genes. Thus, mutants affecting MYC proteins may augment quantitative oncogenic effects on the expression of normal MYC-target genes with qualitative oncogenic effects, by which sets of cell cycle genes are abnormally targeted by MYC as B cells transition into lymphoma cells. The T58A and T58I mutations augment MYC-driven transformation by distinct mechanisms.