The E2F4/p130 repressor complex cooperates with oncogenic deltaNp73alpha to inhibit gene expression in human papillomavirus 38 E6/E7-transformed keratinocytes and in cancer cells

Tumor suppressor p53 and its related proteins, p63 and p73, can be synthesized as multiple isoforms lacking part of the N- or C-terminal regions. Specifically, high expression of the deltaNp73alpha (DNp73a) isoform is notoriously associated with various human malignancies characterized by poor prognosis. This isoform is also accumulated by oncogenic viruses such as Epstein–Barr virus (EBV), as well as genus beta human papillomaviruses (HPV) that appear to be involved in carcinogenesis. To gain additional insight into DNp73a mechanisms, we have performed proteomics analyses using human keratinocytes transformed by the E6 and E7 proteins of the beta-HPV type 38 virus as an experimental model (38HK). We find that DNp73a associates with the E2F4/p130 repressor complex through a direct interaction with E2F4. This interaction is favored by the N-terminal truncation of p73 characteristic of DNp73 isoforms. Moreover, it is independent of the C-terminal splicing status, suggesting that it could represent a general feature of DNp73 isoforms. We show that the DNp73a-E2F4/p130 complex inhibits the expression of specific genes, including genes encoding for negative regulators of proliferation, both in 38HK and in HPV-negative cancer-derived cell lines. Such genes are not inhibited by E2F4/p130 in primary keratinocytes lacking DNp73a, indicating that the interaction with DNp73a rewires the E2F4 transcriptional program. In conclusion, we have identified and characterized a novel transcriptional regulatory complex with potential implications in oncogenesis.

肿瘤抑制因子p53及其相关蛋白p63和p73可合成多种异构体，这些异构体缺失部分N-或C-端区域。特别是，deltaNp73alpha (DNp73a)异构体的高表达与多种预后不良的人类恶性肿瘤密切相关。此异构体亦被诸如爱泼斯坦-巴尔病毒（EBV）和β型人乳头瘤病毒（HPV）等致癌病毒积累，这些病毒似乎与致癌过程有关。为了深入了解DNp73a的机制，我们采用38型β-HPV病毒E6和E7蛋白转化的人角质细胞（38HK）作为实验模型，进行了蛋白质组学分析。我们发现DNp73a通过与E2F4的直接触发与E2F4/p130抑制复合物结合。这种结合受DNp73异构体特有的p73 N端截断所促进。此外，这种结合独立于C端的剪接状态，表明它可能代表DNp73异构体的一般特征。我们展示DNp73a-E2F4/p130复合物抑制特定基因的表达，包括编码增殖负调节因子的基因，在38HK和HPV阴性癌细胞系中均如此。这些基因在缺乏DNp73a的原发性角质细胞中不被E2F4/p130抑制，表明DNp73a的相互作用重排了E2F4的转录程序。总之，我们已鉴定并表征了一种新的转录调控复合物，该复合物在肿瘤发生中可能具有潜在的意义。