Analysis of histones in Xenopus laevis I. a distinct index of enriched variants and modifications exists in each cell type and is remodeled during developmental transitions

David Shechter, Joshua J. Nicklay, Raghu K. Chitta, Jeffrey Shabanowitz, Donald F. Hunt, C. David Allis

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Histone proteins contain epigenetic information that is encoded both in the relative abundance of core histones and variants and particularly in the post-translational modification of these proteins. We determined the presence of such variants and covalent modifications in seven tissue types of the anuran Xenopus laevis, including oocyte, egg, sperm, early embryo equivalent (pronuclei incubated in egg extract), S3 neurula cells, A6 kidney cells, and erythrocytes. We first developed a new robust method for isolating the stored, predeposition histones from oocytes and eggs via chromatography on heparin-Sepha-rose, whereas we isolated chromatinized histones via conventional acid extraction. We identified two previously unknown H1 isoforms (H1fx and H1B.Sp) present on sperm chromatin. We immunoblotted this global collection of histones with many specific post-translational modification antibodies, including antibodies against methylated histone H3 on Lys4, Lys9, Lys 27, Lys79, Arg2, Arg17, and Arg 26; methylated histone H4 on Lys20; methylated H2A and H4 on Arg3; acetylated H4 on Lys5, Lys8, Lys 12, and Lys16 and H3 on Lys9 and Lys 14; and phosphorylated H3 on Ser10 and H2A/H4 on Ser 1. Furthermore, we subjected a subset of these histones to two-dimensional gel analysis and subsequent immunoblotting and mass spectrometry to determine the global remodeling of histone modifications that occurs as development proceeds. Overall, our observations suggest that each metazoan cell type may have a unique histone modification signature correlated with its differentiation status.

Original languageEnglish (US)
Pages (from-to)1064-1074
Number of pages11
JournalJournal of Biological Chemistry
Volume284
Issue number2
DOIs
StatePublished - Jan 9 2009
Externally publishedYes

Fingerprint

Xenopus laevis
Histones
Histone Code
Post Translational Protein Processing
Oocytes
Ovum
Spermatozoa
Antibodies
Immunoblotting
Epigenomics
Eggs
Chromatin
Heparin
Chromatography
Mass Spectrometry
Protein Isoforms
Embryonic Structures
Erythrocytes
Gels
Mass spectrometry

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Analysis of histones in Xenopus laevis I. a distinct index of enriched variants and modifications exists in each cell type and is remodeled during developmental transitions. / Shechter, David; Nicklay, Joshua J.; Chitta, Raghu K.; Shabanowitz, Jeffrey; Hunt, Donald F.; David Allis, C.

In: Journal of Biological Chemistry, Vol. 284, No. 2, 09.01.2009, p. 1064-1074.

Research output: Contribution to journalArticle

Shechter, David ; Nicklay, Joshua J. ; Chitta, Raghu K. ; Shabanowitz, Jeffrey ; Hunt, Donald F. ; David Allis, C. / Analysis of histones in Xenopus laevis I. a distinct index of enriched variants and modifications exists in each cell type and is remodeled during developmental transitions. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 2. pp. 1064-1074.
@article{4a1ffdf8052c4bbd9616d3c06d99bf73,
title = "Analysis of histones in Xenopus laevis I. a distinct index of enriched variants and modifications exists in each cell type and is remodeled during developmental transitions",
abstract = "Histone proteins contain epigenetic information that is encoded both in the relative abundance of core histones and variants and particularly in the post-translational modification of these proteins. We determined the presence of such variants and covalent modifications in seven tissue types of the anuran Xenopus laevis, including oocyte, egg, sperm, early embryo equivalent (pronuclei incubated in egg extract), S3 neurula cells, A6 kidney cells, and erythrocytes. We first developed a new robust method for isolating the stored, predeposition histones from oocytes and eggs via chromatography on heparin-Sepha-rose, whereas we isolated chromatinized histones via conventional acid extraction. We identified two previously unknown H1 isoforms (H1fx and H1B.Sp) present on sperm chromatin. We immunoblotted this global collection of histones with many specific post-translational modification antibodies, including antibodies against methylated histone H3 on Lys4, Lys9, Lys 27, Lys79, Arg2, Arg17, and Arg 26; methylated histone H4 on Lys20; methylated H2A and H4 on Arg3; acetylated H4 on Lys5, Lys8, Lys 12, and Lys16 and H3 on Lys9 and Lys 14; and phosphorylated H3 on Ser10 and H2A/H4 on Ser 1. Furthermore, we subjected a subset of these histones to two-dimensional gel analysis and subsequent immunoblotting and mass spectrometry to determine the global remodeling of histone modifications that occurs as development proceeds. Overall, our observations suggest that each metazoan cell type may have a unique histone modification signature correlated with its differentiation status.",
author = "David Shechter and Nicklay, {Joshua J.} and Chitta, {Raghu K.} and Jeffrey Shabanowitz and Hunt, {Donald F.} and {David Allis}, C.",
year = "2009",
month = "1",
day = "9",
doi = "10.1074/jbc.M807273200",
language = "English (US)",
volume = "284",
pages = "1064--1074",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "2",

}

TY - JOUR

T1 - Analysis of histones in Xenopus laevis I. a distinct index of enriched variants and modifications exists in each cell type and is remodeled during developmental transitions

AU - Shechter, David

AU - Nicklay, Joshua J.

AU - Chitta, Raghu K.

AU - Shabanowitz, Jeffrey

AU - Hunt, Donald F.

AU - David Allis, C.

PY - 2009/1/9

Y1 - 2009/1/9

N2 - Histone proteins contain epigenetic information that is encoded both in the relative abundance of core histones and variants and particularly in the post-translational modification of these proteins. We determined the presence of such variants and covalent modifications in seven tissue types of the anuran Xenopus laevis, including oocyte, egg, sperm, early embryo equivalent (pronuclei incubated in egg extract), S3 neurula cells, A6 kidney cells, and erythrocytes. We first developed a new robust method for isolating the stored, predeposition histones from oocytes and eggs via chromatography on heparin-Sepha-rose, whereas we isolated chromatinized histones via conventional acid extraction. We identified two previously unknown H1 isoforms (H1fx and H1B.Sp) present on sperm chromatin. We immunoblotted this global collection of histones with many specific post-translational modification antibodies, including antibodies against methylated histone H3 on Lys4, Lys9, Lys 27, Lys79, Arg2, Arg17, and Arg 26; methylated histone H4 on Lys20; methylated H2A and H4 on Arg3; acetylated H4 on Lys5, Lys8, Lys 12, and Lys16 and H3 on Lys9 and Lys 14; and phosphorylated H3 on Ser10 and H2A/H4 on Ser 1. Furthermore, we subjected a subset of these histones to two-dimensional gel analysis and subsequent immunoblotting and mass spectrometry to determine the global remodeling of histone modifications that occurs as development proceeds. Overall, our observations suggest that each metazoan cell type may have a unique histone modification signature correlated with its differentiation status.

AB - Histone proteins contain epigenetic information that is encoded both in the relative abundance of core histones and variants and particularly in the post-translational modification of these proteins. We determined the presence of such variants and covalent modifications in seven tissue types of the anuran Xenopus laevis, including oocyte, egg, sperm, early embryo equivalent (pronuclei incubated in egg extract), S3 neurula cells, A6 kidney cells, and erythrocytes. We first developed a new robust method for isolating the stored, predeposition histones from oocytes and eggs via chromatography on heparin-Sepha-rose, whereas we isolated chromatinized histones via conventional acid extraction. We identified two previously unknown H1 isoforms (H1fx and H1B.Sp) present on sperm chromatin. We immunoblotted this global collection of histones with many specific post-translational modification antibodies, including antibodies against methylated histone H3 on Lys4, Lys9, Lys 27, Lys79, Arg2, Arg17, and Arg 26; methylated histone H4 on Lys20; methylated H2A and H4 on Arg3; acetylated H4 on Lys5, Lys8, Lys 12, and Lys16 and H3 on Lys9 and Lys 14; and phosphorylated H3 on Ser10 and H2A/H4 on Ser 1. Furthermore, we subjected a subset of these histones to two-dimensional gel analysis and subsequent immunoblotting and mass spectrometry to determine the global remodeling of histone modifications that occurs as development proceeds. Overall, our observations suggest that each metazoan cell type may have a unique histone modification signature correlated with its differentiation status.

UR - http://www.scopus.com/inward/record.url?scp=59449108085&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=59449108085&partnerID=8YFLogxK

U2 - 10.1074/jbc.M807273200

DO - 10.1074/jbc.M807273200

M3 - Article

VL - 284

SP - 1064

EP - 1074

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 2

ER -