Limits...
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.


Serine/threonine O-acetylation in sugarcane histone H3.(A) MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the histone H3 peptide prKSacTGGKprAPR (residues 9–17) where S10 is acetylated. (B) MS/MS spectra of the doubly-charged precursor ion at m/z 598.8534 corresponding to H3T22 acetylation in the H3 peptide prKQLATacKprAAR (residues 18–26). Sequence of the modified peptide and the measured mass of the precursor ion are shown in the figure inset. N-terminal and lysine propionylation, products of the chemical derivatization, are indicated by pr.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4520453&req=5

pone.0134586.g003: Serine/threonine O-acetylation in sugarcane histone H3.(A) MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the histone H3 peptide prKSacTGGKprAPR (residues 9–17) where S10 is acetylated. (B) MS/MS spectra of the doubly-charged precursor ion at m/z 598.8534 corresponding to H3T22 acetylation in the H3 peptide prKQLATacKprAAR (residues 18–26). Sequence of the modified peptide and the measured mass of the precursor ion are shown in the figure inset. N-terminal and lysine propionylation, products of the chemical derivatization, are indicated by pr.

Mentions: Recently, low level O-acetylation at serine, threonine and tyrosine (S/T/Y) was shown to occur in the histone H3 of several model organisms [22]. H3S10ac in particular appears to be conserved from humans to yeast [22]. In our nanoLC-MS/MS analysis of sugarcane histones, we identified several peptides acetylated at H3S10, H3T22, H3S28, H3Y41 and H3Y54 (S3 Table). Acetylation of these residues, with exception of H3Y41, has been shown to occur in other organisms [22]. H3S10ac was identified from the MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the peptide prKSacTGGKAPR (residues 9–17) where H3S10 is acetylated (Fig 3A). However, the presence of multiple unassigned peaks of high intensity in the middle of the MS/MS spectrum in which H3S10ac was identified indicate there is a mix of peptides. Some of these peaks can be match to y5, y6 and y7 ions corresponding to prKme1SacTGGKacAPR (S5 Fig). Therefore, this spectrum corresponds to a mixture of H3S10ac and H3K9me1S10acK14ac peptides.


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)

Serine/threonine O-acetylation in sugarcane histone H3.(A) MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the histone H3 peptide prKSacTGGKprAPR (residues 9–17) where S10 is acetylated. (B) MS/MS spectra of the doubly-charged precursor ion at m/z 598.8534 corresponding to H3T22 acetylation in the H3 peptide prKQLATacKprAAR (residues 18–26). Sequence of the modified peptide and the measured mass of the precursor ion are shown in the figure inset. N-terminal and lysine propionylation, products of the chemical derivatization, are indicated by pr.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0134586.g003: Serine/threonine O-acetylation in sugarcane histone H3.(A) MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the histone H3 peptide prKSacTGGKprAPR (residues 9–17) where S10 is acetylated. (B) MS/MS spectra of the doubly-charged precursor ion at m/z 598.8534 corresponding to H3T22 acetylation in the H3 peptide prKQLATacKprAAR (residues 18–26). Sequence of the modified peptide and the measured mass of the precursor ion are shown in the figure inset. N-terminal and lysine propionylation, products of the chemical derivatization, are indicated by pr.
Mentions: Recently, low level O-acetylation at serine, threonine and tyrosine (S/T/Y) was shown to occur in the histone H3 of several model organisms [22]. H3S10ac in particular appears to be conserved from humans to yeast [22]. In our nanoLC-MS/MS analysis of sugarcane histones, we identified several peptides acetylated at H3S10, H3T22, H3S28, H3Y41 and H3Y54 (S3 Table). Acetylation of these residues, with exception of H3Y41, has been shown to occur in other organisms [22]. H3S10ac was identified from the MS/MS spectrum of the [M+2H]2+ ion (m/z 556.3089) that matched the peptide prKSacTGGKAPR (residues 9–17) where H3S10 is acetylated (Fig 3A). However, the presence of multiple unassigned peaks of high intensity in the middle of the MS/MS spectrum in which H3S10ac was identified indicate there is a mix of peptides. Some of these peaks can be match to y5, y6 and y7 ions corresponding to prKme1SacTGGKacAPR (S5 Fig). Therefore, this spectrum corresponds to a mixture of H3S10ac and H3K9me1S10acK14ac peptides.

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.