Activating transcription factor 3 (ATF3) is a common stress sensor, and its rapid induction by cellular stresses (e.g. DNA damage) is crucial for cells to mount appropriate responses (e.g. activating the tumor suppressor p53) and maintain homeostasis. Although emerging evidence suggests that dysregulation of ATF3 contributes to occurrences of human diseases including cancer, the mechanism(s) by which ATF3 expression is regulated is largely unknown. Here, we demonstrate that mouse double minute 2 (MDM2) is a bona fide E3 ubiquitin ligase for ATF3 and regulates ATF3 expression by promoting its degradation. MDM2 via its C-terminal RING finger can bind to the Basic region of ATF3 and mediate the addition of ubiquitin moieties to the ATF3 leucine zipper domain. As a consequence, ATF3, but not a mutant deficient in MDM2 binding (⌬80 -100), is degraded by MDM2-mediated proteolysis. Consistent with these results, ablation of MDM2 in cells not only increases basal ATF3 levels, but results in stabilization of ATF3 in late stages of DNA damage responses. Because ATF3 was recently identified as a p53 activator, these results suggest that MDM2 could inactivate p53 through an additional feedback mechanism involving ATF3. Therefore, we provide the first evidence demonstrating that ATF3 is regulated by a posttranslational mechanism. ATF32 is a member of the ATF/CREB family of transcription factors, and its expression is rapidly induced by a large variety of cellular stresses including DNA damage, wounds, and cellular injury (1). ATF3 can bind to DNA (via the ATF/CREB consensus sequence, 5Ј-TGACGTCA-3Ј) (1) and other proteins (e.g. Smad3, p53, and E6) (2-4), resulting in alterations in gene expression and cellular functions. Although consequences of stress-induced ATF3 expression are not well understood, recent evidence links ATF3 to several important pathways, including TGF␤ signaling (2), the Toll-like receptor 4 pathway (5), the eIF2 kinase-mediated endoplasmic reticulum stress response (6), as well as the p53 activation pathway (3), suggesting that dysregulation of ATF3 could contribute to occurrences of many human diseases including cancer. Indeed, although we previously showed that ATF3 deficiency promotes oncogenic transformation (3, 7), recent unbiased cDNA array studies have revealed that ATF3 expression is down-regulated in common human cancers (for a review of these data, see Ref. (8).Although induction of ATF3 expression is a common characteristic of stress responses (1), the mechanisms by which ATF3 expression is regulated during these processes remain largely unknown. It has been shown that ATF3 expression can be regulated by transcription factors such as ATF2, Smad3, and NF-B (2, 9, 10) and controlled by signaling mediated by p38 or JNK/SAPK (9, 11). Moreover, an atypical p53-binding site was identified in the ATF3 promoter (12). However, whether p53 regulates ATF3 expression still remains to be firmly validated, even though a marginal effect of p53 on ATF3 expression was reported in specific cells and in response...

Genomic integration of human papillomavirus (HPV) DNA accounts for more than 90% of cervical cancers. High-risk genital HPVs encode E6 proteins that can interact with a cellular ubiquitin ligase E6-associated protein (E6AP) and target the tumor suppressor p53 for ubiquitin-mediated proteolysis. Currently, how this critical event is regulated is largely unknown. Here we report that activating transcription factor 3 (ATF3), a broad DNA damage sensor whose expression is frequently downregulated in cervical cancer, interacted with E6 and prevented p53 from ubiquitination and degradation mediated by the viral protein. Consistent with its role as a potent E6 antagonist, ATF3 expressed enforcedly in HPV-positive SiHa cells activated p53, leading to expression of p53-target genes (e.g. p21 and PUMA), cell cycle arrest and apoptotic cell death. The leucine zipper domain of ATF3 appears indispensable for these effects as an ATF3 mutant lacking this domain failed to interact with E6 and activate p53 in the cervical cancer cells. The prevention of p53 degradation was unlikely caused by binding of ATF3 to the tumor suppressor, but rather was a consequence of disruption of the E6-E6AP interaction by ATF3. These results indicate that ATF3 plays a key role in a mechanism defending against HPV-induced carcinogenesis, and could serve as a novel therapeutic target for HPV-positive cancers. Human papillomavirus (HPV)3 infection is a major risk factor for cervical cancer (1, 2). Genomic integration of HPV DNA, occurring in more than 90% of cervical cancers, results in expression of a viral protein E6, which in turn inactivates the tumor suppressor p53 by driving its proteolysis (3). The E6 protein can bind to p53 at both the C terminus and the central DNA-binding region (4) and recruit a cellular protein E6AP to p53 (5, 6). E6AP belongs to the HECT family of E3 ubiquitin ligases and can catalyze the addition of ubiquitin moieties to p53, leading to its degradation by the 26 S proteasomes (6). Because the p53 gene is rarely mutated in cervical cancer, the E6-mediated degradation serves as the major mechanism inactivating p53 and promoting cervical carcinogenesis (7).In addition to p53 and E6AP, the E6 protein interacts with many other cellular proteins including Bak (8), Bax (9), CBP/ p300 (10), and BRCA1 (11), presumably through the four CXXC motifs sparsely distributed in the viral protein (12). These interactions account for the oncogenic activities of E6, which include not only promoting transformation but also enhancing cell proliferation and survival (13). Therefore, it is important to dissect the E6 interaction network for a better understanding of the molecular basis for cervical cancer and identification of therapeutic targets for the disease.ATF3 is a member of the ATF/CREB family of transcription factors and can be rapidly induced by DNA damage and other oncogenic stimuli (14,15). Whereas consequences of ATF3 induction are unclear, it is often assumed that ATF3 functions as a transcription factor to regulate gene expressio...