C/EBPβ represses p53 to promote cell survival downstream of DNA damage independent of oncogenic Ras and p19Arf

Abstract

CCAAT/enhancer-binding protein-β (C/EBPβ) is a mediator of cell survival and tumorigenesis. When C/EBPβ−/− mice are treated with carcinogens that produce oncogenic Ras mutations in keratinocytes, they respond with abnormally elevated keratinocyte apoptosis and a block in skin tumorigenesis. Although this aberrant carcinogen-induced apoptosis results from abnormal upregulation of p53, it is not known whether upregulated p53 results from oncogenic Ras and its ability to induce p19Arf and/or activate DNA-damage response pathways or from direct carcinogen-induced DNA damage. We report that p19Arf is dramatically elevated in C/EBPβ−/− epidermis and that C/EBPβ represses a p19Arf promoter reporter. To determine whether p19Arf is responsible for the proapoptotic phenotype in C/EBPβ−/− mice, C/EBPβ−/−;p19Arf−/− mice were generated. C/EBPβ−/−;p19Arf−/− mice responded to carcinogen treatment with increased p53 and apoptosis, indicating p19Arf is not essential. To ascertain whether oncogenic Ras activation induces aberrant p53 and apoptosis in C/EBPβ−/− epidermis, we generated K14-ER:Ras; C/EBPβ−/− mice. Oncogenic Ras activation induced by 4-hydroxytamoxifen did not produce increased p53 or apoptosis. Finally, when C/EBPβ−/− mice were treated with differing types of DNA-damaging agents, including alkylating chemotherapeutic agents, they displayed aberrant levels of p53 and apoptosis. These results indicate that C/EBPβ represses p53 to promote cell survival downstream of DNA damage and suggest that inhibition of C/EBPβ may be a target for cancer cotherapy to increase the efficacy of alkylating chemotherapeutic agents.

Arf Induction by Tgfβ Is Influenced by Sp1 and C/ebpβ in Opposing Directions

“…For example, C/ebpβ −/− mice are completely refractory to chemically induced skin cancer [40], which concept is consistent with higher p19 Arf expression as a tumor suppressor. However, Arf does not seem to play a role in tumor resistance in this model [26]. Nonetheless, our findings demonstrating increased Arf mRNA in the vitreous of C/ebpβ −/− embryos indicates that C/ebpβ can repress Arf in a normal developmental context.…”

“…The mice were generated by introducing a MCI-Neo poly(A)+ mutation at the 3′ terminus of C/ebpβ to abolish translation of the LAP and LIP isoforms [24]. As previously described [26], analysis of cultured MEFs derived from wild type and C/ebpβ −/− embryos demonstrated that basal Arf mRNA and p19 Arf protein were increased upon C/ebpβ loss (Figure 2C and D, lane 3 versus 1). Despite the increased baseline Arf expression, though, absence of C/ebpβ only minimally influenced the further induction of Arf mRNA by Tgfβ (Figure 2C, compare lane 4 versus 3 with 2 versus 1).…”

“…Among those, C/ebpβ was an attractive candidate because previous work had implicated it as an Arf repressor in primary epidermal keratinocytes [26], and putative consensus DNA binding elements are found within 500 bp 5′ to the Arf translation initiation codon (Figure 1A). Utilizing chromatin immunoprecipitation (ChIP), we demonstrated that C/ebpβ was bound to this region in cultured mouse embryo fibroblasts (MEFs) at passage 3 (YZ and SXS, unpublished data).…”

“…That C/ebpβ can repress Arf was previously suggested primarily by the elevated Arf mRNA and protein observed in C/ebpβ −/− keratinocytes in culture and in the adult mouse [26]. Sp1 is well known to bind to GC-rich promoter elements [37], [38], and the mouse and human Arf promoters contain numerous Sp1 binding sites within CpG islands [15], [33].…”

Non-receptor Tyrosine Kinases c-Abl and Arg Regulate the Activity of C/EBPβ

“…Many studies have shown that C/EBPβ has a pro-survival role by aberrant regulation of p53 level and function, or preventing TNF-induced apoptosis of fibroblasts by activating the expression of MnSOD. [52][53][54] Here, we show that tyrosine phosphorylation on Y79 of C/ EBPβ by c-Abl is vital for regulation of cell survival.…”