Distribution of histone H4 modifications as revealed by a panel of specific monoclonal antibodies

Yoko Hayashi-Takanaka, Kazumitsu Maehara, Akihito Harada, Takashi Umehara, Shigeyuki Yokoyama, Chikashi Obuse, Yasuyuki Ohkawa, Naohito Nozaki, Hiroshi Kimura

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

Post-translational histone modifications play a critical role in genome functions such as epigenetic gene regulation and genome maintenance. The tail of the histone H4 N-terminus contains several amino acids that can be acetylated and methylated. Some of these modifications are known to undergo drastic changes during the cell cycle. In this study, we generated a panel of mouse monoclonal antibodies against histone H4 modifications, including acetylation at K5, K8, K12, and K16, and different levels of methylation at K20. Their specificity was evaluated by ELISA and immunoblotting using synthetic peptide and recombinant proteins that harbor specific modifications or amino acid substitutions. Immunofluorescence confirmed the characteristic distributions of target modifications. An H4K5 acetylation (H4K5ac)-specific antibody CMA405 reacted with K5ac only when the neighboring K8 was unacetylated. This unique feature allowed us to detect newly assembled H4, which is diacetylated at K5 and K12, and distinguish it from hyperacetylated H4, where K5 and K8 are both acetylated. Chromatin immunoprecipiation combined with deep sequencing (ChIP-seq) revealed that acetylation of both H4K8 and H4K16 were enriched around transcription start sites. These extensively characterized and highly specific antibodies will be useful for future epigenetics and epigenome studies.

Original languageEnglish
Pages (from-to)753-766
Number of pages14
JournalChromosome Research
Volume23
Issue number4
DOIs
Publication statusPublished - Dec 1 2015

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Histone Code
Acetylation
Histones
Monoclonal Antibodies
Epigenomics
Genome
High-Throughput Nucleotide Sequencing
Antibodies
Transcription Initiation Site
Amino Acid Substitution
Post Translational Protein Processing
Recombinant Proteins
Immunoblotting
Methylation
Chromatin
Fluorescent Antibody Technique
Tail
Cell Cycle
Enzyme-Linked Immunosorbent Assay
Maintenance

All Science Journal Classification (ASJC) codes

  • Genetics

Cite this

Distribution of histone H4 modifications as revealed by a panel of specific monoclonal antibodies. / Hayashi-Takanaka, Yoko; Maehara, Kazumitsu; Harada, Akihito; Umehara, Takashi; Yokoyama, Shigeyuki; Obuse, Chikashi; Ohkawa, Yasuyuki; Nozaki, Naohito; Kimura, Hiroshi.

In: Chromosome Research, Vol. 23, No. 4, 01.12.2015, p. 753-766.

Research output: Contribution to journalArticle

Hayashi-Takanaka, Yoko ; Maehara, Kazumitsu ; Harada, Akihito ; Umehara, Takashi ; Yokoyama, Shigeyuki ; Obuse, Chikashi ; Ohkawa, Yasuyuki ; Nozaki, Naohito ; Kimura, Hiroshi. / Distribution of histone H4 modifications as revealed by a panel of specific monoclonal antibodies. In: Chromosome Research. 2015 ; Vol. 23, No. 4. pp. 753-766.
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