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Chemically ubiquitylated histone H2B stimulates hDot1L-mediated intranucleosomal methylation

Chemical ubiquitylation of histones Previous studies have suggested that histone H2B ubiquitylation and H3 K79 methylation are correlated; however, the underlying mechanism of this crosstalk is not understood, in a large part due to the difficulty of isolating homogenously ubiquitylated H2B (uH2B)....

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Published in:Nature 2008-06, Vol.453 (7196), p.812-816
Main Authors: McGinty, Robert K., Kim, Jaehoon, Chatterjee, Champak, Roeder, Robert G., Muir, Tom W.
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Language:English
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Summary:Chemical ubiquitylation of histones Previous studies have suggested that histone H2B ubiquitylation and H3 K79 methylation are correlated; however, the underlying mechanism of this crosstalk is not understood, in a large part due to the difficulty of isolating homogenously ubiquitylated H2B (uH2B). McGinty et al . overcame this obstacle by developing a synthetic route to obtain uH2B, employing two traceless chemical ligation strategies. Incorporation of this protein into chemically defined nucleosomes has allowed the demonstration that ubiquitylation of H2B directly stimulates methylation of H3 K79 by the methyltransferase hDot1L in an intranucleosomal fashion. An approach is described to synthesize chemically a protein that is ubiquitylated on a specific residue. Insight is provided into how ubiquitylated histone H2B regulates methylation of histone H3 in a trans histone crosstalk pathway. Numerous post-translational modifications of histones have been described in organisms ranging from yeast to humans 1 . Growing evidence for dynamic regulation of these modifications, position- and modification-specific protein interactions, and biochemical crosstalk between modifications has strengthened the ‘histone code’ hypothesis, in which histone modifications are integral to choreographing the expression of the genome 1 , 2 . One such modification, ubiquitylation of histone H2B (uH2B) on lysine 120 (K120) in humans 3 , and lysine 123 in yeast 4 , has been correlated with enhanced methylation of lysine 79 (K79) of histone H3 (refs 5–8 ), by K79-specific methyltransferase Dot1 (KMT4) 9 , 10 , 11 . However, the specific function of uH2B in this crosstalk pathway is not understood. Here we demonstrate, using chemically ubiquitylated H2B, a direct stimulation of hDot1L-mediated intranucleosomal methylation of H3 K79. Two traceless orthogonal expressed protein ligation (EPL) reactions were used to ubiquitylate H2B site-specifically. This strategy, using a photolytic ligation auxiliary and a desulphurization reaction, should be generally applicable to the chemical ubiquitylation of other proteins. Reconstitution of our uH2B into chemically defined nucleosomes, followed by biochemical analysis, revealed that uH2B directly activates methylation of H3 K79 by hDot1L. This effect is mediated through the catalytic domain of hDot1L, most likely through allosteric mechanisms. Furthermore, asymmetric incorporation of uH2B into dinucleosomes showed that the enhancement of methyla
ISSN:0028-0836
1476-4687
1476-4679
DOI:10.1038/nature06906