The role of MIER1 and G9a in histone 3 lysine 27 (H3K27) methylation

Derwish, Leena (2014) The role of MIER1 and G9a in histone 3 lysine 27 (H3K27) methylation. Masters thesis, Memorial University of Newfoundland.

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Abstract

G9a is a lysine methyltransferase, which is responsible for mono- and di-methylation of lysine 9 on histone 3 in euchromatic loci containing transcriptionally active genes. G9a has been shown to be implicated in the methylation of lysine 27 on histone 3 as well. This study focused on characterizing the interaction between G9a and mesoderm induction early response 1 (MIER1), a fibroblast growth factor (FGF)-activated transcriptional regulator. Glutathione S-transferase pull-down assay showed that MIER1 interacts with G9a via the SANT domain, a domain common in MIER1 isoforms. Co-immunoprecipitations using lysates from human embryonic kidney cells (HEK-293) confirmed the validity of the interaction between these two proteins in vivo. Co-immunoprecipitations also revealed MIER1 to interact with chromodomain protein, Y-like (CDYL). Moreover, CDYL bridges enhancer of zeste homolog 2 (EZH2), the methyltransferase subunit of PRC2 complex, to MIER1. Methyltransferase assays using histone H3 N-terminal [H3(21-44)] peptide or recombinant histone H3 monomer as substrates showed recombinant G9a to have no detectable methylation activity towards H3K27. GST-MIER1 does not have intrinsic histone methyltransferase activity and does not methylate H3K27. However, the MIER1 complex immunoprecipiated from HEK-293 cells does methylate H3K27. A methyltransferase associated with MIER1 in HEK-293 cells, such as G9a or EZH2, could be responsible for this methylation.

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