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Smyd3 regulates cancer cell phenotypes and catalyzes histone H4 lysine 5 methylation.

Van Aller GS, Reynoird N, Barbash O, Huddleston M, Liu S, Zmoos AF, McDevitt P, Sinnamon R, Le B, Mas G, Annan R, Sage J, Garcia BA, Tummino PJ, Gozani O, Kruger RG - Epigenetics (2012)

Bottom Line: Smyd3 is a lysine methyltransferase implicated in chromatin and cancer regulation.Here we show that Smyd3 catalyzes histone H4 methylation at lysine 5 (H4K5me).This novel histone methylation mark is detected in diverse cell types and its formation is attenuated by depletion of Smyd3 protein.

View Article: PubMed Central - PubMed

Affiliation: GlaxoSmithKline, Collegeville, PA, USA.

ABSTRACT
Smyd3 is a lysine methyltransferase implicated in chromatin and cancer regulation. Here we show that Smyd3 catalyzes histone H4 methylation at lysine 5 (H4K5me). This novel histone methylation mark is detected in diverse cell types and its formation is attenuated by depletion of Smyd3 protein. Further, Smyd3-driven cancer cell phenotypes require its enzymatic activity. Thus, Smyd3, via H4K5 methylation, provides a potential new link between chromatin dynamics and neoplastic disease.

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Related in: MedlinePlus

SMYD3 selectively methylates H4 in vitro. (A) Heat map summarizing results from a library of 327 histone peptides tested as Smyd3 methylation substrates. Peptides were derived from histones H2a, H2b, H3 and H4 and possessed various combinations of modifications (see Table S1). (B) Methylation assays using the indicated recombinant Smyd3 proteins on recombinant histone H4. Autoradiograph and Coomassie stained gels (loading control) are shown. (C) Quantitative evaluation of Smyd3 methylation using recombinant histones H3 and H4. (D) Methylation assays as in (b) using reconstituted nucleosomes. SET8 is known to methylate nucleosomal H4.9
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Figure 1: SMYD3 selectively methylates H4 in vitro. (A) Heat map summarizing results from a library of 327 histone peptides tested as Smyd3 methylation substrates. Peptides were derived from histones H2a, H2b, H3 and H4 and possessed various combinations of modifications (see Table S1). (B) Methylation assays using the indicated recombinant Smyd3 proteins on recombinant histone H4. Autoradiograph and Coomassie stained gels (loading control) are shown. (C) Quantitative evaluation of Smyd3 methylation using recombinant histones H3 and H4. (D) Methylation assays as in (b) using reconstituted nucleosomes. SET8 is known to methylate nucleosomal H4.9

Mentions: The methylation activity of recombinant full-length Smyd3 was screened against a library of 327 modified and unmodified histone peptides (see Table S1 for peptide content). The most highly methylated peptides were derived from histone H4 residues 1–21 (Fig. 1A, Table S1). None of the H3 peptides in the panel showed any activity as substrates including peptides that are unmethylated, mono- or di-methylated at H3K4 (Fig. 1A). Moreover, Smyd3 methylated recombinant histone, but three different Smyd3 catalytic mutants (Smyd3F183A, Smyd3N205A, and Smyd3Y239A) failed to do so (Fig. 1B and Fig. S1). Additionally, Smyd3 demonstrated 10-fold higher activity with recombinant histone H4 as compared to histone H3 (Fig. 1C). When reconstituted nucleosomes were used as substrates, Smyd3 methylated H4, like the positive control SET8/PR-Set7,9,10 but not the other core histones (Fig. 1D).


Smyd3 regulates cancer cell phenotypes and catalyzes histone H4 lysine 5 methylation.

Van Aller GS, Reynoird N, Barbash O, Huddleston M, Liu S, Zmoos AF, McDevitt P, Sinnamon R, Le B, Mas G, Annan R, Sage J, Garcia BA, Tummino PJ, Gozani O, Kruger RG - Epigenetics (2012)

SMYD3 selectively methylates H4 in vitro. (A) Heat map summarizing results from a library of 327 histone peptides tested as Smyd3 methylation substrates. Peptides were derived from histones H2a, H2b, H3 and H4 and possessed various combinations of modifications (see Table S1). (B) Methylation assays using the indicated recombinant Smyd3 proteins on recombinant histone H4. Autoradiograph and Coomassie stained gels (loading control) are shown. (C) Quantitative evaluation of Smyd3 methylation using recombinant histones H3 and H4. (D) Methylation assays as in (b) using reconstituted nucleosomes. SET8 is known to methylate nucleosomal H4.9
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3368817&req=5

Figure 1: SMYD3 selectively methylates H4 in vitro. (A) Heat map summarizing results from a library of 327 histone peptides tested as Smyd3 methylation substrates. Peptides were derived from histones H2a, H2b, H3 and H4 and possessed various combinations of modifications (see Table S1). (B) Methylation assays using the indicated recombinant Smyd3 proteins on recombinant histone H4. Autoradiograph and Coomassie stained gels (loading control) are shown. (C) Quantitative evaluation of Smyd3 methylation using recombinant histones H3 and H4. (D) Methylation assays as in (b) using reconstituted nucleosomes. SET8 is known to methylate nucleosomal H4.9
Mentions: The methylation activity of recombinant full-length Smyd3 was screened against a library of 327 modified and unmodified histone peptides (see Table S1 for peptide content). The most highly methylated peptides were derived from histone H4 residues 1–21 (Fig. 1A, Table S1). None of the H3 peptides in the panel showed any activity as substrates including peptides that are unmethylated, mono- or di-methylated at H3K4 (Fig. 1A). Moreover, Smyd3 methylated recombinant histone, but three different Smyd3 catalytic mutants (Smyd3F183A, Smyd3N205A, and Smyd3Y239A) failed to do so (Fig. 1B and Fig. S1). Additionally, Smyd3 demonstrated 10-fold higher activity with recombinant histone H4 as compared to histone H3 (Fig. 1C). When reconstituted nucleosomes were used as substrates, Smyd3 methylated H4, like the positive control SET8/PR-Set7,9,10 but not the other core histones (Fig. 1D).

Bottom Line: Smyd3 is a lysine methyltransferase implicated in chromatin and cancer regulation.Here we show that Smyd3 catalyzes histone H4 methylation at lysine 5 (H4K5me).This novel histone methylation mark is detected in diverse cell types and its formation is attenuated by depletion of Smyd3 protein.

View Article: PubMed Central - PubMed

Affiliation: GlaxoSmithKline, Collegeville, PA, USA.

ABSTRACT
Smyd3 is a lysine methyltransferase implicated in chromatin and cancer regulation. Here we show that Smyd3 catalyzes histone H4 methylation at lysine 5 (H4K5me). This novel histone methylation mark is detected in diverse cell types and its formation is attenuated by depletion of Smyd3 protein. Further, Smyd3-driven cancer cell phenotypes require its enzymatic activity. Thus, Smyd3, via H4K5 methylation, provides a potential new link between chromatin dynamics and neoplastic disease.

Show MeSH
Related in: MedlinePlus