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Aberrant MYC expression is a common oncogenic event in human cancer. Paradoxically, MYC can either drive cell cycle progression or induce apoptosis. The latent ability of MYC to induce apoptosis has been termed "intrinsic tumor suppressor activity," and reactivating this apoptotic function in tumors is widely considered a valuable therapeutic goal. As a transcription factor, MYC controls the expression of many downstream targets, and for the majority of these, it remains unclear whether or not they play direct roles in MYC function. To identify the subset of genes specifically required for biological activity, we conducted a screen for functionally important MYC targets and identified BAG1, which encodes a prosurvival chaperone protein. Expression of BAG1 is regulated by MYC in both a mouse model of breast cancer and transformed human cells. Remarkably, BAG1 induction is essential for protecting cells from MYC-induced apoptosis. Ultimately, the synthetic lethality we have identified between MYC overexpression and BAG1 inhibition establishes a new pathway that might be exploited to reactivate the latent apoptotic potential of MYC as a cancer therapy.

Immunoglobulin heavy chain rearrangement (V(H)-to-DJ(H)) occurs only in B cells, suggesting it is inhibited in other lineages. Here we found that in the mouse V(H) locus, methylation of lysine 9 on histone H3 (H3-K9), a mark of inactive chromatin, was present in non-B lineage cells but was absent in B cells. As others have shown that H3-K9 methylation can inhibit V(D)J recombination on engineered substrates, our data support the idea that H3-K9 methylation inhibits endogenous V(H)-to-DJ(H) recombination. We also show that Pax5, a transcription factor required for B cell commitment, is necessary and sufficient for the removal of H3-K9 methylation in the V(H) locus and provide evidence that one function of Pax5 is to remove this inhibitory modification by a mechanism of histone exchange, thus allowing B cell-specific V(H)-to-DJ(H) recombination.

B lymphomagenesis is an uncontrolled expansion of immature precursors that fail to complete their differentiation program. This failure could be at least partly explained by inappropriate expression of several oncogenic transcription factors, such as Pax5 and Myc. Both Pax5 and c-Myc are implicated in the pathogenesis of non-Hodgkin lymphomas. To address their role in lymphomagenesis, we analyzed B-cell lymphomas derived from p53-null bone marrow progenitors infected in vivo by a Myc-encoding retrovirus. All Myc-induced lymphomas invariably maintained expression of Pax5, which is thought to be incompatible with terminal differentiation. However, upon culturing in vitro, several cell lines spontaneously down-regulated Pax5 and its target genes CD19, N-Myc, and MB1. Unexpectedly, other B-cell markers (eg, CD45R) were also down-regulated, and markers of myeloid lineage (CD11b and F4/80 antigen) were acquired instead. Moreover, cells assumed the morphology reminiscent of myeloid cells. A pool of F4/80-positive cells as well as several single-cell clones were obtained and reinjected into syngeneic mice. Remarkably, pooled cells rapidly re-expressed Pax5 and formed tumors of relatively mature lymphoid phenotype, with surface immunoglobulins being abundantly expressed. Approximately half of tumorigenic single-cell clones also abandoned myeloid differentiation and gave rise to B lymphomas. However, when secondary lymphoma cells were returned to in vitro conditions, they once again switched to myeloid differentiation. This process could be curbed via enforced expression of retrovirally encoded Pax5. Our data demonstrate that some Myc target cells are bipotent B-lymphoid/myeloid progenitors with the astonishing capacity to undergo successive rounds of lineage switching.

Myc is a pivotal protooncogenic transcription factor that contributes to the development of almost all Burkitt's lymphomas and about one-third of diffuse large B-cell lymphomas. How B-cells sustain their uncontrolled proliferation due to high Myc is not yet well defined. Here, we found that Myc trans-represses the expression of murine LAPTM5, a gene coding a lysosome-associated protein, by binding to two E-boxes in the LAPTM5 promoter. While the product of intact mRNA (CDS+3′UTR) of LAPTM5 failed to suppress the growth of B-lymphomas, either the protein coded by coding sequence (CDS) itself or the non-coding 3′-untranslated region (3′UTR) mRNA was able to inhibit the growth of B-lymphomas. Moreover, Myc trans-activated miR-17-3p, which promoted tumor growth. Strikingly, LAPTM5 3′UTR contains 11 miR-17-3p-binding sites through which the LAPTM5 protein synthesis was inhibited. The functional interplay between low LAPTM5 mRNA and high miR-17-3p due to high Myc in B-lymphomas leads to further dampening of tumor-suppressive LAPTM5 protein, which promotes tumor progression. Our results indicate that Myc inhibits LAPTM5 expression in B-lymphoma cells by transcriptional and post-transcriptional modifications. [ABSTRACT FROM AUTHOR]

miR-144/451 and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) regulate two antioxidative systems that have been identified to maintain redox homeostasis in erythroid cells by removing excess reactive oxygen species (ROS). Whether these two genes coordinate to affect ROS scavenging and the anemic phenotype, or which gene is more important for recovery from acute anemia, has not been explored. To address these questions, we crossed miR-144/451 knockout (KO) and Nrf2 KO mice and examined the phenotype change in the animals as well as the ROS levels in erythroid cells either at baseline or under stress condition. Several discoveries were made in this study. First, Nrf2/miR-144/451 double-KO mice unexpectedly exhibit similar anemic phenotypes as miR-144/451 single-KO mice during stable erythropoiesis, although compound mutations of miR-144/451 and Nrf2 lead to higher ROS levels in erythrocytes than single gene mutations. Second, Nrf2/miR-144/451 double-mutant mice exhibit more dramatic reticulocytosis than miR-144/451 or Nrf2 single-KO mice during days 3 to 7 after inducing acute hemolytic anemia using phenylhydrazine (PHZ), indicating a synergistic effect of miR-144/451 and Nrf2 on PHZ-induced stress erythropoiesis. However, the coordination does not persist during the whole recovery stage of PHZ-induced anemia; instead, Nrf2/miR-144/451 double-KO mice follow a recovery pattern similar to miR-144/451 single-KO mice during the remaining period of erythropoiesis. Third, the complete recovery from PHZ-induced acute anemia in miR-144/451 KO mice takes longer than in Nrf2 KO mice. Our findings demonstrate that complicated crosstalk between miR-144/451 and Nrf2 does exist and the crosstalk of these two antioxidant systems is development-stage-dependent. Our findings also demonstrate that miRNA deficiency could result in a more profound defect of erythropoiesis than dysfunctional transcription factors. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Summary: The transcription factor GATA-1 is required for terminal erythroid maturation and functions as an activator or repressor depending on gene context. Yet its in vivo site selectivity and ability to distinguish between activated versus repressed genes remain incompletely understood. In this study, we performed GATA-1 ChIP-seq in erythroid cells and compared it to GATA-1-induced gene expression changes. Bound and differentially expressed genes contain a greater number of GATA-binding motifs, a higher frequency of palindromic GATA sites, and closer occupancy to the transcriptional start site versus nondifferentially expressed genes. Moreover, we show that the transcription factor Zbtb7a occupies GATA-1-bound regions of some direct GATA-1 target genes, that the presence of SCL/TAL1 helps distinguish transcriptional activation versus repression, and that polycomb repressive complex 2 (PRC2) is involved in epigenetic silencing of a subset of GATA-1-repressed genes. These data provide insights into GATA-1-mediated gene regulation in vivo. [Copyright &y& Elsevier]