Phosphorylation and arginine methylation mark histone H2A prior to deposition during Xenopus laevis development

Wei Lin Wang, Lissa C. Anderson, Joshua J. Nicklay, Hongshan Chen, Matthew J. Gamble, Jeffrey Shabanowitz, Donald F. Hunt, David Shechter

Research output: Contribution to journalArticle

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Abstract

Background: Stored, soluble histones in eggs are essential for early development, in particular during the maternally controlled early cell cycles in the absence of transcription. Histone post-translational modifications (PTMs) direct and regulate chromatin-templated transactions, so understanding the nature and function of pre-deposition maternal histones is essential to deciphering mechanisms of regulation of development, chromatin assembly, and transcription. Little is known about histone H2A pre-deposition modifications nor known about the transitions that occur upon the onset of zygotic control of the cell cycle and transcription at the mid-blastula transition (MBT).

Results: We isolated histones from staged Xenopus laevis oocytes, eggs, embryos, and assembled pronuclei to identify changes in histone H2A modifications prior to deposition and in chromatin. Soluble and chromatin-bound histones from eggs and embryos demonstrated distinct patterns of maternal and zygotic H2A PTMs, with significant pre-deposition quantities of S1ph and R3me1, and R3me2s. We observed the first functional distinction between H2A and H4 S1 phosphorylation, as we showed that H2A and H2A.X-F (also known as H2A.X.3) serine 1 (S1) is phosphorylated concomitant with germinal vesicle breakdown (GVBD) while H4 serine 1 phosphorylation occurs post-MBT. In egg extract H2A/H4 S1 phosphorylation is independent of the cell cycle, chromatin assembly, and DNA replication. H2AS1ph is highly enriched on blastula chromatin during repression of zygotic gene expression while H4S1ph is correlated with the beginning of maternal gene expression and the lengthening of the cell cycle, consistent with distinct biological roles for H2A and H4 S1 phosphorylation. We isolated soluble H2A and H2A.X-F from the egg and chromatin-bound in pronuclei and analyzed them by mass spectrometry analysis to quantitatively determine abundances of S1ph and R3 methylation. We show that H2A and H4 S1ph, R3me1 and R3me2s are enriched on nucleosomes containing both active and repressive histone PTMs in human A549 cells and Xenopus embryos.

Conclusions: Significantly, we demonstrated that H2A phosphorylation and H4 arginine methylation form a new class of bona fide pre-deposition modifications in the vertebrate embryo. We show that S1ph and R3me containing chromatin domains are not correlated with H3 regulatory PTMs, suggesting a unique role for phosphorylation and arginine methylation.

Original languageEnglish (US)
Article number22
JournalEpigenetics and Chromatin
Volume7
Issue number1
DOIs
StatePublished - Sep 6 2014

Fingerprint

Histone Code
Xenopus laevis
Histones
Methylation
Arginine
Chromatin
Phosphorylation
Serine
Post Translational Protein Processing
Blastula
Embryonic Structures
Eggs
Cell Cycle
Chromatin Assembly and Disassembly
Mothers
Ovum
Gene Expression
Nucleosomes
Xenopus
Cell Cycle Checkpoints

Keywords

  • Early development
  • H2A
  • Histone arginine methylation
  • Histone deposition
  • Histone phosphorylation
  • Histones
  • Xenopus laevis

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology

Cite this

Phosphorylation and arginine methylation mark histone H2A prior to deposition during Xenopus laevis development. / Wang, Wei Lin; Anderson, Lissa C.; Nicklay, Joshua J.; Chen, Hongshan; Gamble, Matthew J.; Shabanowitz, Jeffrey; Hunt, Donald F.; Shechter, David.

In: Epigenetics and Chromatin, Vol. 7, No. 1, 22, 06.09.2014.

Research output: Contribution to journalArticle

Wang, Wei Lin ; Anderson, Lissa C. ; Nicklay, Joshua J. ; Chen, Hongshan ; Gamble, Matthew J. ; Shabanowitz, Jeffrey ; Hunt, Donald F. ; Shechter, David. / Phosphorylation and arginine methylation mark histone H2A prior to deposition during Xenopus laevis development. In: Epigenetics and Chromatin. 2014 ; Vol. 7, No. 1.
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AU - Wang, Wei Lin

AU - Anderson, Lissa C.

AU - Nicklay, Joshua J.

AU - Chen, Hongshan

AU - Gamble, Matthew J.

AU - Shabanowitz, Jeffrey

AU - Hunt, Donald F.

AU - Shechter, David

PY - 2014/9/6

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N2 - Background: Stored, soluble histones in eggs are essential for early development, in particular during the maternally controlled early cell cycles in the absence of transcription. Histone post-translational modifications (PTMs) direct and regulate chromatin-templated transactions, so understanding the nature and function of pre-deposition maternal histones is essential to deciphering mechanisms of regulation of development, chromatin assembly, and transcription. Little is known about histone H2A pre-deposition modifications nor known about the transitions that occur upon the onset of zygotic control of the cell cycle and transcription at the mid-blastula transition (MBT).Results: We isolated histones from staged Xenopus laevis oocytes, eggs, embryos, and assembled pronuclei to identify changes in histone H2A modifications prior to deposition and in chromatin. Soluble and chromatin-bound histones from eggs and embryos demonstrated distinct patterns of maternal and zygotic H2A PTMs, with significant pre-deposition quantities of S1ph and R3me1, and R3me2s. We observed the first functional distinction between H2A and H4 S1 phosphorylation, as we showed that H2A and H2A.X-F (also known as H2A.X.3) serine 1 (S1) is phosphorylated concomitant with germinal vesicle breakdown (GVBD) while H4 serine 1 phosphorylation occurs post-MBT. In egg extract H2A/H4 S1 phosphorylation is independent of the cell cycle, chromatin assembly, and DNA replication. H2AS1ph is highly enriched on blastula chromatin during repression of zygotic gene expression while H4S1ph is correlated with the beginning of maternal gene expression and the lengthening of the cell cycle, consistent with distinct biological roles for H2A and H4 S1 phosphorylation. We isolated soluble H2A and H2A.X-F from the egg and chromatin-bound in pronuclei and analyzed them by mass spectrometry analysis to quantitatively determine abundances of S1ph and R3 methylation. We show that H2A and H4 S1ph, R3me1 and R3me2s are enriched on nucleosomes containing both active and repressive histone PTMs in human A549 cells and Xenopus embryos.Conclusions: Significantly, we demonstrated that H2A phosphorylation and H4 arginine methylation form a new class of bona fide pre-deposition modifications in the vertebrate embryo. We show that S1ph and R3me containing chromatin domains are not correlated with H3 regulatory PTMs, suggesting a unique role for phosphorylation and arginine methylation.

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KW - Early development

KW - H2A

KW - Histone arginine methylation

KW - Histone deposition

KW - Histone phosphorylation

KW - Histones

KW - Xenopus laevis

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