c-MYC-Induced AP4 Attenuates DREAM-Mediated Repression by p53

SIMPLE SUMMARY: Deregulated expression of the c-MYC oncogene activates the tumor suppressor p53, which has been suggested to represent a failsafe mechanism against the uncontrolled expansion of tumor cells. Here, we analyzed the role of the c-MYC-induced TFAP4/AP4 gene in this context using a genetic approach in MCF-7 breast cancer cells. Inactivation of AP4 resulted in elevated levels of both spontaneous and c-MYC-induced DNA damage, senescence, and diminished cell proliferation. Inactivation of p53 in AP4-deficient cells reverted senescence and proliferative defects. Furthermore, loss of AP4 resulted in p53-dependenct, enhanced repression of DREAM and E2F target genes after the induction of c-MYC, which could be abrogated by the concomitant depletion of p21 or the DREAM complex component LIN37. These p53-dependent effects were reflected on the levels of gene expressions and clinical associations in primary breast cancer tumors from patient cohorts. Our results established AP4 as a pivotal factor at the crossroads of c-MYC, E2F, and p53-mediated target gene regulation. ABSTRACT: Background: The deregulated expression of the c-MYC oncogene activates p53, which is presumably mediated by ARF/INK4, as well as replication-stress-induced DNA damage. Here, we aimed to determine whether the c-MYC-inducible AP4 transcription factor plays a role in this context using a genetic approach. Methods: We used a CRISPR/Cas9 approach to generate AP4- and/or p53-deficient derivatives of MCF-7 breast cancer cells harboring an ectopic, inducible c-MYC allele. Cell proliferation, senescence, DNA damage, and comprehensive RNA expression profiles were determined after activation of c-MYC. In addition, we analyzed the expression data from primary breast cancer samples. Results: Loss of AP4 resulted in elevated levels of both spontaneous and c-MYC-induced DNA damage, senescence, and diminished cell proliferation. Deletion of p53 in AP4-deficient cells reverted senescence and proliferation defects without affecting DNA damage levels. RNA-Seq analyses showed that loss of AP4 enhanced repression of DREAM and E2F target genes after p53 activation by c-MYC. Depletion of p21 or the DREAM complex component LIN37 abrogated this effect. These p53-dependent effects were conserved on the level of clinical and gene expression associations found in primary breast cancer tumors. Conclusions: Our results establish AP4 as a pivotal factor at the crossroads of c-MYC, E2F, and p53 target gene regulation.