E2f8 mediates tumor suppression in postnatal liver development
Lindsey N. Kent, Jessica B. Rakijas, Shusil K. Pandit, Bart Westendorp, Hui-Zi Chen, Justin T. Huntington, Xing Tang, Sooin Bae, Arunima Srivastava, Shantibhusan Senapati, Christopher Koivisto, Chelsea K. Martin, Maria C. Cuitino, Miguel Perez, Julian M. Clouse, Veda Chokshi, Neelam Shinde, Raleigh Kladney, Daokun Sun, Antonio Perez-Castro, Ramadhan B. Matondo, Sathidpak Nantasanti, Michal Mokry, Kun Huang, Raghu Machiraju, Soledad Fernandez, Thomas J. Rosol, Vincenzo Coppola, Kamal S. Pohar, James M. Pipas, Carl R. Schmidt, Alain de Bruin, Gustavo Leone
Lindsey N. Kent, Jessica B. Rakijas, Shusil K. Pandit, Bart Westendorp, Hui-Zi Chen, Justin T. Huntington, Xing Tang, Sooin Bae, Arunima Srivastava, Shantibhusan Senapati, Christopher Koivisto, Chelsea K. Martin, Maria C. Cuitino, Miguel Perez, Julian M. Clouse, Veda Chokshi, Neelam Shinde, Raleigh Kladney, Daokun Sun, Antonio Perez-Castro, Ramadhan B. Matondo, Sathidpak Nantasanti, Michal Mokry, Kun Huang, Raghu Machiraju, Soledad Fernandez, Thomas J. Rosol, Vincenzo Coppola, Kamal S. Pohar, James M. Pipas, Carl R. Schmidt, Alain de Bruin, Gustavo Leone
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Research Article Oncology

E2f8 mediates tumor suppression in postnatal liver development

Abstract

E2F-mediated transcriptional repression of cell cycle–dependent gene expression is critical for the control of cellular proliferation, survival, and development. E2F signaling also interacts with transcriptional programs that are downstream of genetic predictors for cancer development, including hepatocellular carcinoma (HCC). Here, we evaluated the function of the atypical repressor genes E2f7 and E2f8 in adult liver physiology. Using several loss-of-function alleles in mice, we determined that combined deletion of E2f7 and E2f8 in hepatocytes leads to HCC. Temporal-specific ablation strategies revealed that E2f8’s tumor suppressor role is critical during the first 2 weeks of life, which correspond to a highly proliferative stage of postnatal liver development. Disruption of E2F8’s DNA binding activity phenocopied the effects of an E2f8 null allele and led to HCC. Finally, a profile of chromatin occupancy and gene expression in young and tumor-bearing mice identified a set of shared targets for E2F7 and E2F8 whose increased expression during early postnatal liver development is associated with HCC progression in mice. Increased expression of E2F8-specific target genes was also observed in human liver biopsies from HCC patients compared to healthy patients. In summary, these studies suggest that E2F8-mediated transcriptional repression is a critical tumor suppressor mechanism during postnatal liver development.

Authors

Lindsey N. Kent, Jessica B. Rakijas, Shusil K. Pandit, Bart Westendorp, Hui-Zi Chen, Justin T. Huntington, Xing Tang, Sooin Bae, Arunima Srivastava, Shantibhusan Senapati, Christopher Koivisto, Chelsea K. Martin, Maria C. Cuitino, Miguel Perez, Julian M. Clouse, Veda Chokshi, Neelam Shinde, Raleigh Kladney, Daokun Sun, Antonio Perez-Castro, Ramadhan B. Matondo, Sathidpak Nantasanti, Michal Mokry, Kun Huang, Raghu Machiraju, Soledad Fernandez, Thomas J. Rosol, Vincenzo Coppola, Kamal S. Pohar, James M. Pipas, Carl R. Schmidt, Alain de Bruin, Gustavo Leone

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Figure 7

Identification of HCC-relevant E2F targets by ChIP-seq.

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Identification of HCC-relevant E2F targets by ChIP-seq. (A) Tag-intensit...
(A) Tag-intensity heat map showing the distribution of tags for all E2F7 and E2F8 peaks identified by ChIP-seq. Peaks were centered on E2F8-specific samples except for peaks that were specific to E2F7. (B) Percentage of E2F7- and E2F8-specific peaks in different gene regions. Gene regions were defined by distance from the transcriptional start site (TSS) as follows: 5′ distal (–50 Gb to –50 kb), 5′ proximal (–50 kb to –5 kb), promoter (–5 kb to +2 kb), gene body (+2 kb to end of transcript), 3′ distal (end of transcript to +30 Gb). Number of peaks for each gene region is indicated above bars. (C) Graph depicting the frequency of E2F7 and E2F8 tags relative to the TSS (0). The promoter region (–5 kb to +2 kb from the TSS) is highlighted and the consensus binding sequence at the promoter identified by HOMER is depicted. (D) Examples of E2F7 and E2F8 occupancy at selected promoters. (E) ChIP-qPCR validation using IgG, E2F7, or E2F8 antibodies in HepG2 cells. Selected target promoters are shown (CHEK1, FDPS, MCM2, PRIM1, RAD51, TIMELESS, and TOP2A). A nonpromoter region of TOP2A (TOP2A neg) was used as a negative control. % input values were normalized to IgG. Primers were designed to amplify ChIP-seq–identified peak regions. (F) Gene ontology using ingenuity pathway analysis (IPA) software depicts the estimated contribution of gene functions associated with E2F7- or E2F8-bound promoters. Functional categories related to cell cycle, cancer, and liver disease with the lowest P values are shown. Bars indicate the Benjamini-Hochberg–adjusted (B-H) P value; the threshold of P = 0.05 is shown.