Akt-mediated phosphorylation of Bmi1 modulates its oncogenic potential, E3 ligase activity, and DNA damage repair activity in mouse prostate cancer
Karim Nacerddine, Jean-Bernard Beaudry, Vasudeva Ginjala, Bart Westerman, Francesca Mattiroli, Ji-Ying Song, Henk van der Poel, Olga Balagué Ponz, Colin Pritchard, Paulien Cornelissen-Steijger, John Zevenhoven, Ellen Tanger, Titia K. Sixma, Shridar Ganesan, Maarten van Lohuizen
Karim Nacerddine, Jean-Bernard Beaudry, Vasudeva Ginjala, Bart Westerman, Francesca Mattiroli, Ji-Ying Song, Henk van der Poel, Olga Balagué Ponz, Colin Pritchard, Paulien Cornelissen-Steijger, John Zevenhoven, Ellen Tanger, Titia K. Sixma, Shridar Ganesan, Maarten van Lohuizen
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Research Article Oncology

Akt-mediated phosphorylation of Bmi1 modulates its oncogenic potential, E3 ligase activity, and DNA damage repair activity in mouse prostate cancer

Abstract

Prostate cancer (PCa) is a major lethal malignancy in men, but the molecular events and their interplay underlying prostate carcinogenesis remain poorly understood. Epigenetic events and the upregulation of polycomb group silencing proteins including Bmi1 have been described to occur during PCa progression. Here, we found that conditional overexpression of Bmi1 in mice induced prostatic intraepithelial neoplasia, and elicited invasive adenocarcinoma when combined with PTEN haploinsufficiency. In addition, Bmi1 and the PI3K/Akt pathway were coactivated in a substantial fraction of human high-grade tumors. We found that Akt mediated Bmi1 phosphorylation, enhancing its oncogenic potential in an Ink4a/Arf-independent manner. This process also modulated the DNA damage response and affected genomic stability. Together, our findings demonstrate the etiological role of Bmi1 in PCa, unravel an oncogenic collaboration between Bmi1 and the PI3K/Akt pathway, and provide mechanistic insights into the modulation of Bmi1 function by phosphorylation during prostate carcinogenesis.

Authors

Karim Nacerddine, Jean-Bernard Beaudry, Vasudeva Ginjala, Bart Westerman, Francesca Mattiroli, Ji-Ying Song, Henk van der Poel, Olga Balagué Ponz, Colin Pritchard, Paulien Cornelissen-Steijger, John Zevenhoven, Ellen Tanger, Titia K. Sixma, Shridar Ganesan, Maarten van Lohuizen

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Figure 8

Phosphorylation of Bmi1 enhances HR and genomic instability.

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Phosphorylation of Bmi1 enhances HR and genomic instability. (A) Phospho...
(A) Phosphorylation of Bmi1 was required for efficient DNA DSB repair by HR. The HR GFP reporter is stably integrated into the genome of U2OS cell line. SceGFP is a GFP gene that contains an I-SceI endonuclease site within the coding region. Cleavage of the I-SceI site in vivo and repair by HR directed by the downstream iGFP repeat results in GFP+ cells. HR GFP U2OS cells were transfected with scrambled (Sc) siRNA or Bmi1 siRNA reconstituted with constructs expressing mouse empty vector, Bmi1-WT, or Bmi1-3A. Where indicated, cells were also transfected with an I-SceI expression vector. After 24 hours, the percentage of GFP+ cells was measured by flow cytometry. Data are means ± SD of 3 independent experiments. *P = 0.006 versus Bmi1-WT, unpaired t test. (B) Frequency of spontaneous SCEs in doxycycline-treated LNCaP-tet-shBmi1 prostate cancer cells infected with retroviruses expressing Bmi1-WT, Bmi1-3A, or control empty vector. SCEs were increased 2.3-fold with Bmi1-WT, but not Bmi1-3A, overexpression (*P < 0.0001, unpaired t test). Akt inhibition (Akti) suppressed Bmi1-WT–induced SCEs. Data are means ± SEM. Examples of SCE events (arrowheads) are shown in the inset, which depicts subsets of chromosomes in a metaphase spread labeled for SCE analysis (original magnification, ×200). (C) Immunohistochemistry for γH2AX revealed greater DNA damage in tumor areas of PbCre4;Bmi1LSL;PtenloxP/+ mutant prostates (n = 27) and Bmi1+pAkt+ human samples (n = 77). Scale bars: 100 μm.