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Analysis of Histones H3 and H4 Reveals Novel and Conserved Post-Translational Modifications in Sugarcane.

Moraes I, Yuan ZF, Liu S, Souza GM, Garcia BA, Casas-Mollano JA - PLoS ONE (2015)

Bottom Line: As a result, modifications, alone or in combination, are important determinants of chromatin states.Several modifications conserved in other plants, and also novel modified residues, were identified.Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, Brazil.

ABSTRACT
Histones are the main structural components of the nucleosome, hence targets of many regulatory proteins that mediate processes involving changes in chromatin. The functional outcome of many pathways is "written" in the histones in the form of post-translational modifications that determine the final gene expression readout. As a result, modifications, alone or in combination, are important determinants of chromatin states. Histone modifications are accomplished by the addition of different chemical groups such as methyl, acetyl and phosphate. Thus, identifying and characterizing these modifications and the proteins related to them is the initial step to understanding the mechanisms of gene regulation and in the future may even provide tools for breeding programs. Several studies over the past years have contributed to increase our knowledge of epigenetic gene regulation in model organisms like Arabidopsis, yet this field remains relatively unexplored in crops. In this study we identified and initially characterized histones H3 and H4 in the monocot crop sugarcane. We discovered a number of histone genes by searching the sugarcane ESTs database. The proteins encoded correspond to canonical histones, and their variants. We also purified bulk histones and used them to map post-translational modifications in the histones H3 and H4 using mass spectrometry. Several modifications conserved in other plants, and also novel modified residues, were identified. In particular, we report O-acetylation of serine, threonine and tyrosine, a recently identified modification conserved in several eukaryotes. Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species. To our knowledge, this is the first report of histones H3 and H4 as well as their post-translational modifications in sugarcane, and will provide a starting point for the study of chromatin regulation in this crop.

No MeSH data available.


Post-translational modifications identified in the histone H4 from sugarcane.Amino acid residues identified by the nanoLC-MS/MS analysis are indicated in red. The modification sites identified are shown on top of the sequence and the amino acid residue highlighted in blue (lysine) and purple (threonine). The first amino acid methionine was omitted from the sequence.
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pone.0134586.g007: Post-translational modifications identified in the histone H4 from sugarcane.Amino acid residues identified by the nanoLC-MS/MS analysis are indicated in red. The modification sites identified are shown on top of the sequence and the amino acid residue highlighted in blue (lysine) and purple (threonine). The first amino acid methionine was omitted from the sequence.

Mentions: In sugarcane we identified one protein with similarity to the canonical histone H4, Ss_H4.1, and a single variant, Ss_H4.2, with a single amino acid substitution Y72C. NanoLC-MS/MS analysis of bulk histone sequenced several peptides corresponding to histone H4. The protein coverage was 93.2% for Ss_H4.1 and 83.5% for the Ss_H4.2 variant (Fig 7). However, unique peptides generated from trypsin digestion of Ss_H4.2 were not found indicating that the variant may be present at very low levels, below the detection limit of the assay, or may be a tissue specific variant not expressed in leaf rolls. Therefore we consider all the modifications identified belonging only to Ss_H4.1.


Analysis of Histones H3 and H4 Reveals Novel and Conserved Post-Translational Modifications in Sugarcane.

Moraes I, Yuan ZF, Liu S, Souza GM, Garcia BA, Casas-Mollano JA - PLoS ONE (2015)

Post-translational modifications identified in the histone H4 from sugarcane.Amino acid residues identified by the nanoLC-MS/MS analysis are indicated in red. The modification sites identified are shown on top of the sequence and the amino acid residue highlighted in blue (lysine) and purple (threonine). The first amino acid methionine was omitted from the sequence.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4520453&req=5

pone.0134586.g007: Post-translational modifications identified in the histone H4 from sugarcane.Amino acid residues identified by the nanoLC-MS/MS analysis are indicated in red. The modification sites identified are shown on top of the sequence and the amino acid residue highlighted in blue (lysine) and purple (threonine). The first amino acid methionine was omitted from the sequence.
Mentions: In sugarcane we identified one protein with similarity to the canonical histone H4, Ss_H4.1, and a single variant, Ss_H4.2, with a single amino acid substitution Y72C. NanoLC-MS/MS analysis of bulk histone sequenced several peptides corresponding to histone H4. The protein coverage was 93.2% for Ss_H4.1 and 83.5% for the Ss_H4.2 variant (Fig 7). However, unique peptides generated from trypsin digestion of Ss_H4.2 were not found indicating that the variant may be present at very low levels, below the detection limit of the assay, or may be a tissue specific variant not expressed in leaf rolls. Therefore we consider all the modifications identified belonging only to Ss_H4.1.

Bottom Line: As a result, modifications, alone or in combination, are important determinants of chromatin states.Several modifications conserved in other plants, and also novel modified residues, were identified.Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, Brazil.

ABSTRACT
Histones are the main structural components of the nucleosome, hence targets of many regulatory proteins that mediate processes involving changes in chromatin. The functional outcome of many pathways is "written" in the histones in the form of post-translational modifications that determine the final gene expression readout. As a result, modifications, alone or in combination, are important determinants of chromatin states. Histone modifications are accomplished by the addition of different chemical groups such as methyl, acetyl and phosphate. Thus, identifying and characterizing these modifications and the proteins related to them is the initial step to understanding the mechanisms of gene regulation and in the future may even provide tools for breeding programs. Several studies over the past years have contributed to increase our knowledge of epigenetic gene regulation in model organisms like Arabidopsis, yet this field remains relatively unexplored in crops. In this study we identified and initially characterized histones H3 and H4 in the monocot crop sugarcane. We discovered a number of histone genes by searching the sugarcane ESTs database. The proteins encoded correspond to canonical histones, and their variants. We also purified bulk histones and used them to map post-translational modifications in the histones H3 and H4 using mass spectrometry. Several modifications conserved in other plants, and also novel modified residues, were identified. In particular, we report O-acetylation of serine, threonine and tyrosine, a recently identified modification conserved in several eukaryotes. Additionally, the sub-nuclear localization of some well-studied modifications (i.e., H3K4me3, H3K9me2, H3K27me3, H3K9ac, H3T3ph) is described and compared to other plant species. To our knowledge, this is the first report of histones H3 and H4 as well as their post-translational modifications in sugarcane, and will provide a starting point for the study of chromatin regulation in this crop.

No MeSH data available.