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, Résumé en français
, le dogme central de la biologie moléculaire consistait en un flux d'information allant de l'ADN, à l'ARN-messager (ARNm) puis aux protéines, comme déterminants principaux de l'identité cellulaire. Cependant, les nouvelles techniques de séquençage à haut-débit ont révélées que parmi les 3 milliards de bases qui composent le génome humain, seules 2% codent pour des protéines. En revanche, 97% du génome humain sont transcrits en ARN, dont la grande majorité provient ainsi de régions dites « non-codantes » (nc) (Djebali et al, 2000.
, Ces lncARN sont transcrits par l'ARN Polymérase II et subissent une maturation similaire à celle des ARNm puisqu'ils sont généralement coiffés, épissés et poly-adénylés, Ils sont impliqués dans de nombreux processus biologiques, 2011.
, Grace à leur expression hautement spécifique, ces lncARN ont été proposés comme des biomarqueurs de diagnostic et classification (Li et al, 2013) ou même comme des acteurs de la cancérogénèse, 2016.
, Le travail présenté ici se concentre donc sur ces lncARN liés au cancer et plus spécifiquement sur leur association à la transition épithélio-mésenchymateuse (TEM)
, Récemment, le changement strict de l'identité épithéliale à mésenchymateuse a été remis en question et des états hybrides/intermédiaires ont été identifiés, avec notamment des phénotypes variables de caractère « souche », de plasticité cellulaire ou de capacité d'invasion et migration. Ces traits sont particulièrement importants dans le développement tumoral et sont associés à la résistance aux traitements, aux métastase et à la récurrence tumorale, Bien que les gènes codants impliqués dans la TEM aient été caractérisés dans le développement et plus récemment dans le cancer, le rôle des lncARN n'a que très peu été décrit. Il est cependant indéniable puisque dans les cinq dernières années, 2006.
des lncARN dans la régulation de la TEM en identifiant les lncARN différentiellement exprimés entre les cellules épithéliales et mésenchymateuses. J'ai d'abord caractérisé le rôle du lncARN connu HOTAIR dans la TEM (chapitre 4) puis j'ai identifié de nouveaux candidats en définissant ceux qui étaient fonctionnels au travers d'un criblage génétique par CRISPR (chapitre 5). Dans ce but, j'ai utilisé un système HEK-TEM ,
, qui reposent sur un modèle original provenant de cellules primaires humaines d'épithélium de rein (HEK) et dont ont été tirées une lignée cellulaire épithéliale « Epi » et une lignée mésenchymateuse « Mes
,
Comme première preuve de concept de l'étude de lncARN dans le système HEK-TEM, je me suis concentré sur le rôle de HOTAIR dans la régulation de la TEM, HOTAIR comme étant surexprimé dans les cellules Mes, 2019. ,
, Il a été montré que HOTAIR recrute des protéines de modifications de la chromatine, pour réprimer l'expression génique, grâce à des domaines structuraux situées à ses extrémités en 5' et 3'. Ces domaines interagissent avec les complexes PRC2 et Lsd1-CoREST-REST, respectivement. Nous avons d'abord défini l'importance de ces deux domaines dans la régulation de la TEM en surexprimant des variants tronqués de HOTAIR dans les cellules Epi