Our second main project aims at characterizing novel signalling pathways controlling stem cell potential. We chose the skin as a model tissue of a highly renewable epithelial tissue to study new mechanisms implicated in the control of adult stem cell functions. We are currently developping large-scale genetic and chemical screens based on epidermal stem cells in order to identify new signalling pathways and pharmacological modulators regulating stem cell potential. This part of our project may have broad applications in regenerative medecine and novel anti-tumor strategies.
Project 1: Characterization of E4F1 functions in the p53 pathway.
The exploration of the protein networks deregulated by viral oncoproteins led to the discovery in the 80ies of several essential regulators of cell division and cell survival. Among those, E4F1 was originally discovered as an ubiquitously expressed cellular transcription factor regulated by the viral oncoprotein E1A during adenoviral infection. While most other cellular targets of E1A discovered at the same time (E2F/pRB, CBP/p300, PCAF etc…) have been extensively studied and recognized as central regulators of cell proliferation and survival, E4F1 functions remained, inexplicably, poorly investigated until recently. However, our laboratory recently showed that E4F1 is a multifunctional protein that is essential during early embryogenesis and plays important roles in the proliferation/survival balance of different cell types including stem cells. Some, but not all, of E4F1 functions are linked to its implication in the p53 pathway. E4F1 physically interacts with several essential components of this pathway, including p53, p14ARF, and Bmi1. Interestingly, we found that E4F1 stimulated oligo-ubiquitylation of p53 does not lead to its proteasomal degradation but instead regulates its transcriptional activities. Altogether, our results indicate that E4F1 is both a transcription factor and a novel key post-translational regulator of the p53 pathway. We are more specifically focusing on one branch of this pathway implicating several E4F1-interacting proteins, the Bmi1-Ink4a/ARF-p53 pathway, which has been previously described to play important roles in cellular senescence, aging, and carcinogenesis.
Using a variety of approaches, including mouse mutants and biochemical analyses of E4F1-mediated post-translational modifications, we are investigating new levels of regulation of the p53 pathway implicating E4F1.
Project 2: Identification of novel signalling pathways regulating epidermal stem cell potential.
Throughout adult life, the epidermis and the hair follicle undergo continuous renewal due to stem cell activities. Stem cells in the epidermis and hair follicles not only ensure the maintenance of epidermal homeostasis and hair regeneration, but also contribute to repair of the epidermis after injury. It is also believed that accumulation of genetic lesions in those long lived adult stem cells actively contribute to tumorigenesis. One of our main objective is to identify and characterize new signalling pathways implicated in epidermal stem cell functions. To address these questions, we currently develop several large-scale genetic and chemical screens using primary keratinocytes isolated from several of our mouse mutants.
Our strategy should improve our knowledge of the molecular circuitries involved in epidermal stem cells activities and identify chemical compounds with potential clinical and industrial applications in stem cell-dependent skin physiology. We also investigate whether our findings can be extrapolated to other types of adult stem cells, such as the hematopoietic stem cells, and whether perturbations of those pathways contribute to tumor development.
Project 3: E4F1 functions in tumorigenesis
E4F1 directly interacts with / regulates / or is regulated by several bona fide tumor suppressors (p53, pRB, p14ARF and Rassf1A) and oncoproteins (E1A, HBX, Gam1, Bmi1, HMGA2), suggesting that it might itself play a role in carcinogenesis. We are currently investigating how E4F1-mediated activities influence tumor incidence and progression in different genetically engineered tumor prone mouse models in which we can conditionnally inactivate or overexpress E4F1. This project led us to identify p53-independent functions of E4F1 that are implicated in cell survival. So far, we mainly focused on leukemia models based on the genetic inactivation of the Ink4a/ARF locus and chemically-induced skin tumors. We are also evaluating E4F1 expression levels in human tumor samples. |
