Roles of EZH2 and KDM6B during EMT


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Camille Lachat1,Amandine Etcheverry2,Marc Aubry2,Jean Mosser2,Michael Herfs3,Jean-Paul Feugeas4,Michael Boyer-Guittaut4,Paul Peixoto5,Eric Hervouet5
1INSERM UMR1098,2CHU Rennes, Service de Génétique Moléculaire et Génomique, Rennes F-35033, France ; CNRS, UMR 6290, Institut de Génétique et Développement de Rennes (IGDR), Rennes F-35043, France ; Université Rennes1, UEB, UMS 3480 Biosit, Faculté de Médecine, Rennes F-35043, France ; Plate-forme Génomique Environnementale et Humaine Biosit, Université Rennes1, Rennes F-35043, France,3Laboratory of Experimental Pathology, GIGA-Cancer, University of Liege, Liege, Belgium,4Univ. Bourgogne Franche-Comté, INSERM, EFS BFC, UMR1098, Interactions Hôte-Greffon-Tumeur/Ingénierie Cellulaire et Génique, F-25000 Besançon, France,5Univ. Bourgogne Franche-Comté, INSERM, EFS BFC, UMR1098, Interactions Hôte-Greffon-Tumeur/Ingénierie Cellulaire et Génique, F-25000 Besançon, France ; EPIGENEXP Platform, University of Bourgogne Franche-Comté, F-25000 Besançon, France

Abstract

Background

Epigenetic enzymes EZH2 and KDM6B catalyze methylation and demethylation of the epigenetic mark H3K27me3 (trimethylation of lysine 27 of histone H3), respectively. Trimethylation of H3K27 lead to the inhibition of transcription of downstream genes. Despite their opposite roles, these enzymes have already been described to be involved in Epithelial to Mesenchymal Transition (EMT) and/or tumor aggressiveness and metastasis formation. Studying the roles of these enzymes during EMT could therefore lead to the improvement of cancer treatment efficiency since metastasis formation is one of the main factors linked to bad prognosis in cancer.

Method

We analyzed EZH2 and KDM6B expressions in vivo in lung tumor biopsies (n=100) using IHC assay and in vitro in the lung cancer cell line A549. EMT marker expressions, cellular migration and invasion capacities were analyzed after modulation of EZH2 or KDM6B activity and expression. Moreover, ChIP-sequencing analysis was performed against H3K27me3, EZH2 and KDM6B to identify epigenetically-regulated promoters of genes involved in EMT. Then, ChIP-qPCR were performed during EMT to confirm these data. Finally, we used siRNA targeting these genes to characterize their roles during EMT.  

Results

In our cohort, KDM6B and EZH2 expressions have been correlated with the mesenchymal marker VIMENTIN. In A549 cell line, overexpression of EZH2 and KDM6B both activated the mesenchymal phenotype. Moreover, chemical inhibition of EZH2 and KDM6B induced a mesenchymal phenotype, both at the molecular and the functional level. We therefore hypothesized that KDM6B and EZH2 may present similar or different target genes during EMT. Thanks to ChIP-seq data, we found several of these target genes.

Conclusion

Altogether our data will allow us to: 1) characterize the molecular roles of the lysine demethylase KDM6B and the histone methyltransferase EZH2 during EMT; 2) identify new EMT biomarkers regulated by epigenetic modifications and, 3) propose new epigenetic-based therapy strategies to regulate EMT/metastatic cancer cells.