![]() ![]() These reporters should be useful for studying gene silencing and X-chromosome inactivation with high spatial and temporal resolution in intact cells and may also aid in the search for conjectured histone demethylase activity. ![]() Reporters developed using the chromodomains from HP1 and Polycomb respond to enzymatic methylation at the lysine 9 and lysine 27 positions of histone H3, respectively, giving 60% and 28% YFP/CFP emission ratio increases in vitro or in single living cells. Enzymatic methylation by a methyltransferase induces complexation of the methylated substrate peptide to the chromodomain, changing the FRET level between the flanking CFP and YFP domains. The reporters are four-part chimeric proteins consisting of a substrate peptide from the N-terminus of histone H3 fused to a chromodomain (a natural methyllysine-specific recognition domain), sandwiched between a fluorescence resonance energy transfer (FRET)-capable pair of fluorophores, cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP). We report the design and characterization of two genetically encoded fluorescent reporters of histone protein methylation.
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