CD19 is a major B cell receptor–independent activator of MYC-driven B-lymphomagenesis
Elaine Y. Chung, James N. Psathas, Duonan Yu, Yimei Li, Mitchell J. Weiss, Andrei Thomas-Tikhonenko
Elaine Y. Chung, James N. Psathas, Duonan Yu, Yimei Li, Mitchell J. Weiss, Andrei Thomas-Tikhonenko
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Research Article Oncology

CD19 is a major B cell receptor–independent activator of MYC-driven B-lymphomagenesis

Abstract

PAX5, a B cell–specific transcription factor, is overexpressed through chromosomal translocations in a subset of B cell lymphomas. Previously, we had shown that activation of immunoreceptor tyrosine-based activation motif (ITAM) proteins and B cell receptor (BCR) signaling by PAX5 contributes to B-lymphomagenesis. However, the effect of PAX5 on other oncogenic transcription factor-controlled pathways is unknown. Using a MYC-induced murine lymphoma model as well as MYC-transformed human B cell lines, we found that PAX5 controls c-MYC protein stability and steady-state levels. This promoter-independent, posttranslational mechanism of c-MYC regulation was independent of ITAM/BCR activity. Instead it was controlled by another PAX5 target, CD19, through the PI3K-AKT-GSK3β axis. Consequently, MYC levels in B cells from CD19-deficient mice were sharply reduced. Conversely, reexpression of CD19 in murine lymphomas with spontaneous silencing of PAX5 boosted MYC levels, expression of its key target genes, cell proliferation in vitro, and overall tumor growth in vivo. In human B-lymphomas, CD19 mRNA levels were found to correlate with those of MYC-activated genes. They also negatively correlated with the overall survival of patients with lymphoma in the same way that MYC levels do. Thus, CD19 is a major BCR-independent regulator of MYC-driven neoplastic growth in B cell neoplasms.

Authors

Elaine Y. Chung, James N. Psathas, Duonan Yu, Yimei Li, Mitchell J. Weiss, Andrei Thomas-Tikhonenko

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Figure 2

PAX5 regulates c-MYC protein levels through CD19.

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PAX5 regulates c-MYC protein levels through CD19. All panels except E an...
All panels except E and H represent immunoblotting analyses of proteins indicated. Actin was used as a loading control. (A) Whole cell lysates from MYC5 cells expressing HA-tagged wild-type or mutant ITAM (ITAMmut) were subjected to immunoprecipitation with α-HA antibody, followed by immunoblotting with antibody against total Lyn, Tyr396-phospho-Lyn, or another α-HA antibody (bracketed panels). Lysates from the same cells were immunoblotted with antibodies against c-MYC and β-actin (bottom panels). (B) CD19 and MYC levels in MYC5 cells transduced with empty vector (GFP) or CD19-encoding retrovirus (CD19). (C) PAX5, CD19, and MYC levels in P493-6 cells electroporated with increasing concentrations of control or α-CD19 siRNA. (D) CD19 and MYC levels in bone marrow–derived B cells from wild-type and CD19-null (KO) mice. (E) Flow cytometric detection of CD19 expression on the surface of MYC5 cells expressing PAX5 or low/high levels of CD19. Mean fluorescent intensities are shown in each plot. (F) CD19 and MYC protein expression in the same cultures. (G) MYC levels in PAX5- and CD19-reconstituted cell lines after treatment with CHX. CHX was added at the concentration of 1 μg/ml for 7.5 to 30 minutes. MYC-specific bands were quantitated by densitometry, normalized to actin, and plotted against time after treatment. Note that different exposures are shown for parental and PAX5/CD19-transduced cells to allow accurate band quantitation. Symbols represent individual timepoints. (H) Steady-state MYC levels in the same cultures, detected by radioimmunoprecipitation. Cells were pulse-labeled with 35S-methionine for 30 minutes, followed by a 0 to 30 minutes chase with “cold” amino acids.