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La myoferline est une oncoprotéine émergente qui est surexprimée dans différents types de cancer dont le cancer du pancréas. Le Laboratoire de Recherches sur les Métastases a démontré que la myoferline contrôle la dynamique mitochondriale et la fonction respiratoire. Récemment, plusieurs publications ont suggéré que le cytosquelette d’actine, en plus de son rôle dans la mobilité cellulaire et la dynamique mitochondriale, a une fonction régulatrice sur la phosphorylation oxydative. Nous avons émis l’hypothèse que les effets observés sur la mitochondrie lors de la déplétion de la myoferline pourraient être indirects et impliquer le cytosquelette. Par conséquent, nous avons effectué une analyse in silico d’un ensemble de données obtenues à partir de patients atteints du PDAC (TCGA), ainsi que sur des données obtenues par RNA-Seq et générées à partir de cellules Panc-1 déplétées en myoferline. Nous avons découvert que un ensemble de gènes du cytosquelette d’actine (GO:0015629) était significativement surexprimé dans les deux jeux de données. Ensuite, nous avons analysé des images de microscopie électronique à transmission obtenues à partir de cellules Panc-1 déplétées en myoferline et avons observé une altération de la distribution du cytosquelette dans le cytoplasme. Afin de renforcer ces observations, nous avons évalué l’abondance de certains composants du cytosquelette par analyse en western blot. Nous avons démontré que l’abondance de la b-actine diminuait de 30% lorsque la myoferline était déplétée tandis que l’abondance d’autres constituants, la lamine B1, l’a-tubuline et l’a-actine des muscles lisses ne variait pas. De plus, l’abondance des transcrits de ACTB était significativement diminuée dans les mêmes conditions. Dans le but de caractériser davantage les microfilaments du cytosquelette lors de la déplétion en myoferline, nous avons observé leur organisation par microscopie confocale grâce à une coloration avec de la phalloïdine fluorescente. Alors que de nombreuses fibres épaisses sont apparues dans le cytoplasme des cellules Panc-1 témoins, la coloration était principalement ponctuée et disséminée dans tout le cytoplasme lors de la déplétion en myoferline. Connaissant la fonction du cytosquelette d’actine dans la migration cellulaire, et afin d’aborder un aspect fonctionnel dans le projet, nous avons évalué la migration cellulaire des Panc-1 en utilisant un test « wound healing ». De manière intéressante, en accord avec nos observations précédentes, la déplétion de la myoferline dans les cellules Panc-1 réduit de manière significative sa capacité de migration bidimensionnelle. Nous avons ensuite voulu aborder l’aspect mécanistique de nos observations. Nous avons donc analysé l'abondance et la phosphorylation de la machinerie centrale de la dynamique des microfilaments, à savoir la cofiline, la kinase LIMK1 et la phosphorylase SSH1. Malheureusement, nous n'avons pas été en mesure d'obtenir un ensemble de résultats fiables pour cette dernière partie de notre travail. En conclusion, la déplétion de la myoferline dans une lignée cellulaire de PDAC a un impact sur l'organisation du cytosquelette de b-actine, conduisant à une diminution de la mobilité cellulaire. Myoferlin is an emerging oncoprotein described as overexpressed in different types of cancer including pancreatic cancer. The Metastasis Research Laboratory has demonstrated that myoferlin controls mitochondrial dynamics and respiratory function. Recently, several publications have suggested that the actin cytoskeleton, in addition to its role in cell movement and mitochondrial dynamics, has a regulatory function of oxidative phosphorylation. We hypothesize that the mitochondrial effects of myoferlin depletion could be indirect and involve the actin cytoskeleton. Consequently, we performed in silico analysis of a TCGA data set obtained from PDAC patients, as well as of RNA-seq data generated from Panc-1 cells depleted for myoferlin. We discovered that the actin cytoskeleton gene set (GO:0015629) was significantly enriched in both data sets. We then analyzed electronic microscopy images obtained from myoferlin-depleted Panc-1 cells and reported an alteration of the cytoskeleton distribution inside the cytoplasm. In order to strengthen these observations, we evaluated the abundance of several constituents of cytoskeleton by western blot. We showed that b-actin abundance was decreased by 30% when myoferlin was depleted while other cytoskeleton components, lamin B1, a-tubulin, and a-smooth muscle actin, were not altered. In addition, the abundance of ACTB transcripts were significatively decreased in the same conditions. To further characterized the microfilaments of the cytoskeleton upon myoferlin depletion, we observed its organization by confocal microscopy thanks to a fluorescent-phalloidin staining. While numerous and thick fibers appeared in the cytoplasm of control Panc-1 cells, the staining was mainly punctuated and disseminated throughout all the cytosol upon myoferlin silencing. Owing the known function of b-actin cytoskeleton in cell migration, and in order to tackle a functional aspect of the project, we assessed Panc-1 cell migration using a wound healing assay. Interestingly, and in agreement with previous observations, the depletion of myoferlin in Panc-1 cells reduced significantly its two-dimensional migration ability. We then wanted to address the mechanistic aspect of our observations. Consequently, we analyzed the abundance and the phosphorylation status of the core machinery of microfilaments dynamics, namely cofilin, its kinase LIMK1 and its phosphorylase SSH1. Unfortunately, we were unable to obtain a confident set of results for this last part of my work. In conclusion, myoferlin depletion in a PDAC cell line impact the organization of b-actin cytoskeleton, leading to a decrease of the cell mobility.

Myoferline --- Cytosquelette --- Pancréas --- Pancreas --- Cancer --- Myoferlin --- Cytoskeleton --- Cancer --- Sciences du vivant > Biochimie, biophysique & biologie moléculaire

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Despite the efficiency of current cancer treatments, cancer is still a deadly disease for too many. In 2008, 7.6 million people died of cancer; with the current development, it is estimated that the annual cancer death number will grow to 13 million by 2030. There is clearly a need for not only more research but also more innovative and out of the mainstream scientific ideas to discover and develop even better cancer treatments. This book presents the collective works published in the recent Special Issue entitled “Killing Cancer: Discovery and Selection of New Target Molecules”. These articles comprise a selection of studies, ideas, and opinions that aim to facilitate knowledge, thoughts, and discussion about which biological and molecular mechanisms in cancer we should target and how we should target them.

Research & information: general --- Biology, life sciences --- ferlin --- myoferlin --- dysferlin --- otoferlin --- C2 domain --- plasma membrane --- sulconazole --- NF-κB --- IL-8 --- mammosphere --- breast cancer stem cells --- AF1Q --- MLLT11 --- WNT --- STAT --- esophageal cancer --- prognosis --- mTORC1 --- mTORC2 --- metabolism --- rapalogs --- mTOR inhibitors --- cancer metabolism --- mTOR in immunotherapy --- nutrient metabolism --- kinase inhibitors --- mTOR signaling --- MAPK kinase --- ERK1 --- ERK2 --- CD domain --- Rolled --- SCH772984 --- VRT-11E --- sevenmaker --- cancer therapy --- EMT --- lysosome --- lysosome-mediated invasion --- MZF1 --- phosphorylation --- PAK4 --- SUMOylation --- transcription factor --- zinc finger --- glucocorticoids --- 3D growth --- nuclear factor kappa-light-chain-enhancer of activated B-cells (NF-κB) --- epithelial-mesenchymal transition --- anoikis --- proliferation --- targeted cancer therapy --- disulfiram --- NPL4 --- replication stress --- DNA damage --- BRCA1 --- BRCA2 --- ATR pathway --- PDAC --- TCIRG1 --- ATP6V0a3 --- invasion --- migration --- matrix degradation --- pH-regulation --- autophagy --- multidrug resistance in cancer --- drug efflux pumps --- ATP-binding cassette transporter --- breast cancer resistance protein (BCRP) --- ABCG2 --- pyrazolo-pyrimidine derivative --- SCO-201 --- colorectal cancer --- immunotherapy --- inflammation --- microsatellite instability --- oncofetal chondroitin sulfate --- chondroitin sulfate --- cancer --- solid tumors --- target --- pediatric cancer --- VAR2 --- dexamethasone --- thyroid cancer --- microgravity --- space environment --- ferlin --- myoferlin --- dysferlin --- otoferlin --- C2 domain --- plasma membrane --- sulconazole --- NF-κB --- IL-8 --- mammosphere --- breast cancer stem cells --- AF1Q --- MLLT11 --- WNT --- STAT --- esophageal cancer --- prognosis --- mTORC1 --- mTORC2 --- metabolism --- rapalogs --- mTOR inhibitors --- cancer metabolism --- mTOR in immunotherapy --- nutrient metabolism --- kinase inhibitors --- mTOR signaling --- MAPK kinase --- ERK1 --- ERK2 --- CD domain --- Rolled --- SCH772984 --- VRT-11E --- sevenmaker --- cancer therapy --- EMT --- lysosome --- lysosome-mediated invasion --- MZF1 --- phosphorylation --- PAK4 --- SUMOylation --- transcription factor --- zinc finger --- glucocorticoids --- 3D growth --- nuclear factor kappa-light-chain-enhancer of activated B-cells (NF-κB) --- epithelial-mesenchymal transition --- anoikis --- proliferation --- targeted cancer therapy --- disulfiram --- NPL4 --- replication stress --- DNA damage --- BRCA1 --- BRCA2 --- ATR pathway --- PDAC --- TCIRG1 --- ATP6V0a3 --- invasion --- migration --- matrix degradation --- pH-regulation --- autophagy --- multidrug resistance in cancer --- drug efflux pumps --- ATP-binding cassette transporter --- breast cancer resistance protein (BCRP) --- ABCG2 --- pyrazolo-pyrimidine derivative --- SCO-201 --- colorectal cancer --- immunotherapy --- inflammation --- microsatellite instability --- oncofetal chondroitin sulfate --- chondroitin sulfate --- cancer --- solid tumors --- target --- pediatric cancer --- VAR2 --- dexamethasone --- thyroid cancer --- microgravity --- space environment