High-Risk Human Papillomavirus E6 Protein Promotes Reprogramming of Fanconi Anemia Patient Cells through Repression of p53 but Does Not Allow for Sustained Growth of Induced Pluripotent Stem Cells

Abstract

DNA repair plays a crucial role in embryonic and somatic stem cell biology and cell reprogramming. The Fanconi anemia (FA) pathway, which promotes error-free repair of DNA double-strand breaks, is required for somatic cell reprogramming to induced pluripotent stem cells (iPSC). Thus, cells from Fanconi anemia patients, which lack this critical pathway, fail to be reprogrammed to iPSC under standard conditions unless the defective FA gene is complemented. In this study, we utilized the oncogenes of high-risk human papillomavirus 16 (HPV16) to overcome the resistance of FA patient cells to reprogramming. We found that E6, but not E7, recovers FA iPSC colony formation and, furthermore, that p53 inhibition is necessary and sufficient for this activity. The iPSC colonies resulting from each of these approaches stained positive for alkaline phosphatase, NANOG, and Tra-1-60, indicating that they were fully reprogrammed into pluripotent cells. However, FA iPSC were incapable of outgrowth into stable iPSC lines regardless of p53 suppression, whereas their FA-complemented counterparts grew efficiently. Thus, we conclude that the FA pathway is required for the growth of iPSC beyond reprogramming and that p53-independent mechanisms are involved. IMPORTANCEA novel approach is described whereby HPV oncogenes are used as tools to uncover DNA repair-related molecular mechanisms affecting somatic cell reprogramming. The findings indicate that p53-dependent mechanisms block FA cells from reprogramming but also uncover a previously unrecognized defect in FA iPSC proliferation independent of p53.

Natural history and management of Fanconi anemia patients with head and neck cancer: A 10‐year follow‐up

“…In addition, studies have demonstrated an interaction between the HPV16 E7 oncoprotein and the FA pathway. [38][39][40] It has also been hypothesized that the FA pathway contributes to the repair of DNA damage induced by HPV16 E7, providing one explanation for why FA patients are predisposed to HPV-associated HNSCCs. 41 In FA patients from North America, a high percentage (84%) of HNSCCs were found to be positive for highrisk HPV DNA.…”

Opportunities and challenges of radiotherapy for treating cancer

“…Under conditions of pathological stress, including radiation, 137,138 hypoxia, 139 and oncogene expression, 140 checkpoints that restrict developmental cellular outputs can be lifted, allowing cells to reprogramme for 'stemness' . 141 Such reprogramming could be an essential part of normal healing, but during fractionated courses of radiation it could generate a nidus for recurrence.…”

Fanconi anemia: a model disease for studies on human genetics and advanced therapeutics

“…Reprogramming induces the DDR [99] and activates the FA pathway [94], leading to P53-mediated apoptosis and low reprogramming efficiency [96]. This is partially circumvented by preventing reactive oxygen species (ROS)-mediated DNA damage [94] or by suppressing P53 during reprogramming using RNA interference [96] or human papillomavirus P53-repressing E6 protein [100]. Safe and controlled FANCA gene correction was achieved using integration-free genome editing by genetic recombination with helper-dependent adenoviral vectors [96] or by targeting FANCA insertion into the safe locus AAVS1 [97] using engineered nucleases ("safe harbor" strategy) [101].…”