Metastatic melanoma is an aggressive, incurable cancer with historically few therapeutic options. The discovery that 60% of melanomas harbor the oncogenic BRAF_V600E mutation, which constitutively activates the MAPK pathway, has provided a promising new therapeutic axis. Although MAPK pathway inhibitor therapy has shown striking clinical results in BRAF_V600-mutant melanoma, this approach faces three limitations. First, 10-20% of BRAF_V600-mutant melanomas never achieve meaningful response to MAPK pathway inhibitor therapy (intrinsic resistance). Second, among BRAF_V600-mutant melanomas initially responding to MAPK pathway inhibitor therapy, relapse is universal (acquired resistance). Third, approximately 40% of melanomas lack BRAF_V600 mutations and so are not currently candidates for MAPK pathway inhibitor therapy. We sought to address each of these problems: by characterizing the phenomenon of intrinsic MAPK pathway inhibitor resistance, by finding ways to perturb mechanisms of acquired MAPK pathway inhibitor resistance, and by identifying novel dependencies in melanoma outside of the MAPK pathway. Intriguingly, the NF-kappa B pathway emerged as a common theme across these investigations. In particular, we establish that MAPK pathway inhibitor sensitive and resistant melanomas display distinct transcriptional signatures. Unlike most BRAF_V600-mutant melanomas, which highly express the melanocytic lineage transcription factor MITF, MAPK pathway inhibitor resistant lines display low MITF expression but high levels of NF-kappa B signaling. These divergent transcriptional states, which arise in melanocytes from aberrant MAPK pathway activation by BRAF_V600E, remain plastic and mutually antagonistic in established melanomas. Together, these results characterize a dichotomy between MITF and NF-kappa B cellular states as a determinant of intrinsic sensitivity versus resistance to MAPK pathway inhibitors in BRAF_V600-mutant melanoma. In separate investigations, we have shown that, NFKB1 p105, a member of the NF-kappa B family, intimately regulates levels of COT, a known effector of resistance to MAPK pathway inhibitors. Moreover, we have used shRNA screening to nominate particular nodes within the NF-kappa B pathway, including MYD88 and IRF3, as candidate melanoma lineage-specific dependencies. Cumulatively, although these studies use diverse approaches to investigate the limitations of MAPK pathway inhibitor therapy in melanoma, they converge in nominating the NF-kappa B pathway as a previously underappreciated feature of melanoma biology and suggest the relevance of this pathway for future investigation.

Les bactéries ont réussi à coloniser toutes les niches écologiques de la planète. Le passage d’un environnement à un autre s’accompagne de la fluctuation de nombreux paramètres environnementaux aboutissant à un stress cellulaire. Directement en contact avec le milieu environnant, l’enveloppe bactérienne est la première barrière contre ces stress extracellulaires. Toute rupture de son intégrité aura des conséquences délétères pour la cellule. Parmi les mécanismes permettant aux bactéries de détecter les changements de conditions environnementales, il existe des systèmes spécifiques appelés ESR (Envelope Stress Response). Ces systèmes maintiennent l’intégrité membranaire en réparant les dommages de l’enveloppe. Ce travail de thèse s’inscrit dans l’étude des mécanismes intrinsèques de résistance chez les bactéries, par la caractérisation d’un nouvel ESR d’E. coli : le système ZraPSR (Zinc Resistance Associated Protein Sensor Regulator). ZraPSR est un système à deux composants, composé d’un senseur ZraS, d’un régulateur transcriptionnel ZraR et d’une protéine périplasmique accessoire ZraP. La cascade ZraS-R est activée par des concentrations élevées en Zn et Pb. Ce travail a montré que ZraP établit un rétrocontrôle négatif sur la cascade de signalisation ZraSR par un mécanisme nécessitant sa métallation. Malgré une induction en présence de métaux, nous avons montré que le système ZraPSR ne possède aucun rôle dans l’homéostasie métallique. A contrario, en réponse à des signaux de stress, ZraSR va contribuer à la résistance intrinsèque à certains antimicrobiens. De plus, l’étude du régulon de ZraR a permis de commencer à entrevoir les mécanismes sous-jacents de réponse aux stress antimicrobiens médiée par ZraPSR. Cette réponse intègre des signaux de l’état physiologique de la cellule par l’intermédiaire de régulateurs globaux du métabolisme aboutissant à une réponse optimale. Le système ZraPSR semble donc être un nouveau mécanisme de résistance-croisée aux stress environnementaux. / Bacteria succeed in colonizing all the ecological niches on earth. Transition from one environment to another comes along with the fluctuation in numerous environmental parameters wich induce cellular stress. Directly in contact with the surrounding environment, the bacterial envelope is the first barrier against these extracellular stresses. Any break of its integrity will have deleterious consequences for the cell. Among mechanisms allowing bacteria to detect environmental changes, specific systems called ESR (Envelope Stress Response) have been studied. Such systems maintain membrane integrity by repairing envelope damages. This work takes part in the study of the intrinsic mechanisms of antimicrobial resistance in bacteria, by the characterization of a new ESR of E. coli: the ZraSR (Zinc Resistance Associated Protein Sensor Regulator) system. ZraPSR is a two-component system consisting of a ZraS sensor, a ZraR transcriptional regulator and a ZraP accessory periplasmic protein. The ZraS-R cascade is activated by high concentrations of Zn and Pb. In this study, we showed that ZraP establishes a negative feedback on the ZraSR pathway by a mechanism requiring its metallation. Despite the observed induction in the presence of metals, we showed that the ZraPSR system is not required for metal homeostasis. Whereas, in response to stress signals, ZraSR contribute to intrinsic resistance to certain antimicrobials. Futhermore, the study of the ZraR regulon allowed us to begin glimpsing the underlying mechanisms of antimicrobial stress response mediated by ZraPSR. This response incorporates signals from the physiological state of the cell through global regulators of the metabolism leading to an optimal response. The ZraPSR system seems to be a new cross-resistance mechanism to environmental stresses.