Mapping of lysine monomethylation of linker histones in human breast and its cancer.
Linker histones H1 are key modulators of chromatin structure. Tightness of their binding to DNA is regulated by posttranslational modifications. In this study we have analyzed posttranslational modifications of five major variants of H1 in human tissue - H1.0, H1.2, H1.3, H1.4, and H1.5. To improve sequence coverage, tryptic peptides of H1 were separated by HPLC and the individual fractions were analyzed using a peptide on-chip implementation of nanoelectrospray (TriVersa), coupled to a linear ion trap-orbitrap hybrid instrument. For quantitative analysis of lysine methylation, ionization efficiencies of methylated and nonmethylated peptides were determined using synthetic peptides. Our analysis revealed that monomethylation of lysine residues alongside with phosphorylation of serine and threonine residues is the major modification of H1 in tissue. We found that most prominent methylation sites are in the N-terminal tail and the globular domain of H1. In the C- terminal domains we identified only few and less abundant methylation sites. Quantitative analysis revealed that up to 25% of H1.4 is methylated at K-26 in human tissues. Another prominent methylation site was mapped to K-27 in H1.5, which resembles the K-26 site in H1.4. In H1.0 five less abundant (<1% of H1.0) sites were identified. Analysis of patient matched pairs of cancer and adjacent normal breast demonstrated high variation in H1 methylation between individuals.
Lu
Aiping
A
Department of Proteomics and Signal Transduction, Max-Planck Institute for Biochemistry, Munich, Germany.
Zougman
Alexandre
A
Pudełko
Marek
M
Bebenek
Marek
M
Ziółkowski
Piotr
P
Mann
Matthias
M
Wiśniewski
Jacek R
JR
D000368:Aged; D001940:Breast / Q000737:chemistry / Q000378:metabolism*; D001943:Breast Neoplasms / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D006657:Histones / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D016014:Linear Models; D008239:Lysine / Q000737:chemistry* / Q000378:metabolism; D008745:Methylation; D008875:Middle Aged; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping / Q000379:methods*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry
D006657:Histones; D010446:Peptide Fragments; D008239:Lysine