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Establishment of chromatin states in mouse sperm and their relevance for paternal epigenetic inheritance

English title Establishment of chromatin states in mouse sperm and their relevance for paternal epigenetic inheritance
Applicant Peters Antoine Hendrik Felix Marie
Number 146293
Funding scheme Project funding (Div. I-III)
Research institution Friedrich Miescher Institute for Biomedical Research
Institution of higher education Institute Friedrich Miescher - FMI
Main discipline Genetics
Start/End 01.10.2013 - 30.09.2016
Approved amount 638'880.00
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All Disciplines (3)

Discipline
Genetics
Biochemistry
Embryology, Developmental Biology

Keywords (8)

histone variants; epigenetics; paternal intergenerational epigenetic inheritance; histone dynamics and turnover; spermatogenesis; chromatin remodeling; histone methylation; pre-implantation development

Lay Summary (German)

Lead
Wir untersuchen die molekularen Prozesse, die die Heranreifung der männlichen Keimzellen und die Übermittlung von epigenetischen Informationen vom Vater auf die Nachkommen kontrollieren.
Lay summary

Die Identität eukariotischer Zellen und Organismen wird durch die genetische Information, die in der DNA jeder Zelle gespeichert wird, definiert. Das Ablesen der genetischen Information in den Genen wird durch Proteine kontrolliert, die die DNA in zugängliche und unzugängliche Bereiche verpacken. Die Grundbausteine des Chromatins sind die Nukleosomen, welche aus der DNA und einem Oktamer aus Histonproteinen bestehen. Nukleosomen werden durch verschiedene Enzymklassen chemisch modifiziert und durch grosse Umbaumaschinen, die die Histonoktamere entlang der DNA verschieben, und durch andere Proteine, die an die Histone binden, in unterschiedliche Strukturen zusammengeführt. Es wird vermutet, dass einige Veränderungen an der DNA und an Histonen als epigenetisches Gedächtnis fungieren und Informationen zum Status der väterlichen Zellen an die Tochterzellen weitergeben.

Während der Ausreifung der männlichen Keimzellen der Maus und des Menschen werden die Nukleosomen durch andere Proteine ersetzt, die es ermöglichen die DNA in den sehr kleinen Spermien unterzubringen. Wir zeigten kürzlich, dass in Spermien ein paar wenige Nukleosomen an regulatorischen Sequenzen der Gene zurückbleiben und dass so die Möglichkeit besteht, dass sie väterliche epigenetische Informationen vererben.

Wir versuchen die Mechanismen zu ergründen, die den extensiven, genomweiten Umbau des Chromatins während der Heranreifung der männlichen Keimzellen regulieren. Ausserdem möchten wir die Mechanismen erforschen, die zur Zurückbehaltung der Nukleosomen an spezifischen, regulatorischen Gensequenzen im Spermium führen.

Wir hoffen eine wichtige Frage beantworten zu können, ob Nukleosomen in Mausspermien die Übertragung väterlicher, in Histonen kodierter, epigenetischer Information zwischen Generationen ermöglichen und so die Genexpression in der nächsten Generation kontrollieren.

 

 

Direct link to Lay Summary Last update: 28.11.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Alternative Computational Analysis Shows No Evidence for Nucleosome Enrichment at Repetitive Sequences in Mammalian Spermatozoa.
Royo Hélène, Stadler Michael Beda, Peters Antoine Hendrik Felix Marie (2016), Alternative Computational Analysis Shows No Evidence for Nucleosome Enrichment at Repetitive Sequences in Mammalian Spermatozoa., in Developmental cell, 37(1), 98-104.
The methyltransferase Setdb1 is essential for meiosis and mitosis in mouse oocytes and early embryos.
Eymery Angeline, Liu Zichuan, Ozonov Evgeniy A, Stadler Michael B, Peters Antoine H F M (2016), The methyltransferase Setdb1 is essential for meiosis and mitosis in mouse oocytes and early embryos., in Development (Cambridge, England), 143(15), 2767-79.
Disruption of histone methylation in developing sperm impairs offspring health transgenerationally.
Siklenka Keith, Erkek Serap, Godmann Maren, Lambrot Romain, McGraw Serge, Lafleur Christine, Cohen Tamara, Xia Jianguo, Suderman Matthew, Hallett Michael, Trasler Jacquetta, Peters Antoine H F M, Kimmins Sarah (2015), Disruption of histone methylation in developing sperm impairs offspring health transgenerationally., in Science (New York, N.Y.), 350(6261), 2006-2006.

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Invited seminar at Instituto Gulbenkian de Ciência, Oeiras, Portugal Individual talk CHROMATIN DYNAMICS AND EPIGENETIC INHERITANCE AT THE ONSET OF LIFE 18.11.2016 Oeiras, Portugal Peters Antoine Hendrik Felix Marie;
69th Annual Meeting of the Finnish Endocrine Society, Helsinki, Finland Talk given at a conference CHROMATIN DYNAMICS AND EPIGENETIC INHERITANCE AT THE ONSET OF LIFE 03.11.2016 Helsinki, Finland Peters Antoine Hendrik Felix Marie;
EMBO member Meeting Talk given at a conference Chromatin dynamics and epigenetic inheritance in mammalian reproduction 27.10.2016 Heidelberg, Germany Peters Antoine Hendrik Felix Marie;
2016 IMB Conference: Epigenetics in Development Talk given at a conference Chromatin dynamics in mammalian reproduction 22.10.2016 Mainz, Germany Peters Antoine Hendrik Felix Marie;
Invited seminar at the RIKEN Center for Integrative Medical Sciences, Yokohama, Japan Individual talk Histone variants and proteolysis during spermatogenesis 22.02.2016 Yokohama, Japan Peters Antoine Hendrik Felix Marie;
international Symposium on “Epigenome dynamics and regulation in germ cells” Talk given at a conference Histone variants and proteolysis during spermatogenesis 17.02.2016 Kyoto, Japan Peters Antoine Hendrik Felix Marie;
Hubert Bloch Lecture at the University of Basel Individual talk Epigenetic control of mammalian germline and early embryonic development 13.01.2016 Basel, Switzerland Peters Antoine Hendrik Felix Marie;
Invited seminar at the Technical University Dresden Individual talk Chromatin inheritance and dynamics in mouse germ cells and early embryos 09.11.2015 Dresden, Germany Peters Antoine Hendrik Felix Marie;
Invited seminar a the Erasmus University, Rotterdam, the Netherlands Individual talk Chromatin inheritance and dynamics at the onset of life 01.10.2015 Rotterdam, Netherlands Peters Antoine Hendrik Felix Marie;
48th Annual Meeting of the Society for the Study of Reproduction Talk given at a conference Chromatin inheritance and dynamics at the onset of life 20.06.2015 San Juan, Puerto Rico Peters Antoine Hendrik Felix Marie;
KOLLOQUIUM DES HESSISCHEN ZENTRUMS FÜR REPRODUKTIONSMEDIZIN (HZRM) SOWIE DER KFO 181 UND DES IRTG GIESSEN-MONASH Talk given at a conference Epigenetic control of mouse germ cell and early embryonic development 10.06.2015 Giessen, Germany Peters Antoine Hendrik Felix Marie;
Invited seminar at the Molecular Health Sciences Platform, ETH Zürich Individual talk Chromatin inheritance and dynamics at the onset of life: mechanisms and functional implications 27.03.2015 Zurich, Switzerland Peters Antoine Hendrik Felix Marie;
Invited seminar at the University of Kyoto Individual talk Epigenetic control of mammalian germ line and early embryonic development 27.11.2014 Kyoto, Japan Peters Antoine Hendrik Felix Marie;
The 37th Annual Meeting of the Molecular Biology Society of Japan Talk given at a conference Epigenetic control of mammalian germ line and early embryonic development 26.11.2014 Yokohama, Japan Peters Antoine Hendrik Felix Marie;
Invited seminar at the RIKEN Center for Integrative Medical Sciences Individual talk Epigenetic control of mammalian germ line and early embryonic development 25.11.2014 Yokohama, Japan Peters Antoine Hendrik Felix Marie;
Keystone Symposia meeting on Epigenetic Programming and Inheritance Individual talk Epigenetic control of mouse germ cell and early embryonic development 07.04.2014 Boston, United States of America Peters Antoine Hendrik Felix Marie;
International Meeting of the German Society of Cell Biology Talk given at a conference Epigenetic control of mouse germ cell and early embryonic development 20.03.2014 Regensburg, Germany Peters Antoine Hendrik Felix Marie;
Invited seminar at the Institute of Molecular Biology Individual talk Epigenetic control of mouse germ cell and early embryonic development 20.02.2014 Mainz, Germany Peters Antoine Hendrik Felix Marie;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media FMI report: Antoine Peters: Epigenetic control of mammalian germline and early embryonic development FMI website International 2016
Media relations: print media, online media Affecting health across generations FMI website International 2015
Media relations: print media, online media Detailing heterochromatin formation at the onset of life FMI website International 2015

Awards

Title Year
EMBO member see: http://www.fmi.ch/news/releases/articles/embo.140507.html 2014

Associated projects

Number Title Start Funding scheme
125386 EPIGENETIC CONTROL OF EARLY MOUSE DEVELOPMENT 01.04.2009 Project funding (Div. I-III)
172873 Understanding molecular mechanisms underlying chromatin reprogramming during male germ cell development 01.04.2017 Project funding (Div. I-III)
172873 Understanding molecular mechanisms underlying chromatin reprogramming during male germ cell development 01.04.2017 Project funding (Div. I-III)

Abstract

Background: Histones and their post-translational modification have recently been suggested to mediate epigenetic inheritance of environmentally induced adap¬tive traits via the male germ line (Carone et al., 2010; Zeybel et al., 2012). In mammals, however, genomes of male germ cells are packaged in a highly condensed configuration. To enable the over 10-fold increase in compaction of the genome in spermatozoa, chromatin undergoes a dramatic remodeling along almost the entire genome at the end of spermatogenesis. Key events in this process are the removal of histones and their replacement first with transition proteins and subsequently with protamines (Govin et al., 2004). Nonetheless, this histone removal is not complete. In humans around 10% of histone proteins are retained on chromatin (Brykczynska et al., 2010; Hammoud et al., 2009), while in mice this number stands at 1%. These later findings raise the important question whether these retained histones would be involved in intergenerational transmission of epigenetic information, thereby contributing to gene regulation in offspring.Unpublished data from my laboratory shows that such retained histones are specifically positioned at regulatory regions of important developmental genes, even more explicitely in mouse than in human spermatozoa (Erkek et al., in preparation; Brykczynska et al., 2010). Furthermore, classes of genes serving distinct functions harbor different sets of post-translationally modified histones associated with either transcriptional activation or repression, in a manner that is predictive for expression states measured in early embryos. Importantly, we find that sequence composition is highly predictive for nucleosome enrichment genome-wide and that nucleosome occupancy inversely correlates with DNA methylation. Results from chromatin immunoprecipitation experiments with histone variant specific antibodies suggest specific functions for the canonical histones H3.1 and H3.2 versus the histone variant H3.3 in chromatin remodeling during spermiogenesis and possibly intergenerational epigenetic inheritance (Erkek et al., in preparation). Further preliminary data indicate the existence of an additional, evolutionary conserved, testis expressed histone H3 variant H3t with putative functions in histone eviction during spermiogenesis. Finally, we have observed, specifically in elongating spermatids, the presence of chromatin-bound histone H3 proteins that are cleaved at their N-termini. We speculate that histone clipping may function in the eviction of these proteins during spermiogenesis. On the basis of these data, we define a comprehensive research program that aims at elucidating the mechanisms underlying histone turnover and retention during spermatogenesis and paternal transmission of histone encoded epigenetic information directing gene expression in the next generation. Hypotheses:1.We hypothesize that nucleosomes containing different members of the histone H3 family are assembled onto and disassembled from chromatin during spermatogenesis by distinct pathways.2.We propose that these variant nucleosomes exhibit different properties on chromatin and are selectively evicted versus retained during spermiogenesis.3.We suggest that variant nucleosomes with specific posttranslational modifications that are retained in sperm play distinct roles in the regulation of gene expression in early embryos.Aims: To test these hypotheses we propose three aims that we will experimentally approach using a combination of biochemical, molecular genetic (transgenic and conditional deficiency mouse models), microscopy, epigenomic (ChIP and RNA sequencing) and bio-computational methods.1.To study the mechanisms underlying histone variant specific nucleosome remodeling during spermatogenesis and retention of H3.3 nucleosomes in spermatozoa. To investigate the role of H3.3 and H3t variants and of canonical H3.1 in spermatogenesis we will generate targeted inducible transgenic mice expressing epitope tagged versions of each of these H3 proteins. We will perform ChIP-Sequencing analyses in purified germ cells to determine the localization of specific H3 variants in the genome during spermatogenesis. We will use these transgenic mice to perform genetic pulse-chase experiments to determine when during spermatogenesis nucleosomes containing H3 variants become incorporated into chromatin. To identify chaperone and chromatin remodeler proteins interacting with the different H3 histones, we will perform immunoprecipitation experiments coupled to protein identification by mass spectrometry (IP/MS) on cytoplasmic and nuclear fractions isolated from germ cells at sequential stages of spermatogenesis. Finally, we will functionally characterize the role of H3 variants in spermatogenesis by generating conditional mutant mice for H3.3 and H3t genes as well as for candidate chromatin assembly factors identified by the IP/MS experiments. 2.To determine the mechanism and function of histone clipping in nucleosomal eviction during spermiogenesis. Unpublished data indicate the existence of chromatin-bound H3 proteins cleaved around residue Glycine 33 in purified elongating spermatids. In vitro assays indicate that the proteolytic activity may reside in a serine or threonine protease. To investigate the function of histone cleavage in nucleosome eviction, we will first identify the precise cleavage site using Edman degradation and sophisticated MS approaches. We will subsequently generate antibodies recognizing cleaved histones. We will then perform biochemical fractionation studies combined with IP/MS detection methods to identify the protease involved. To elucidate the function of the protease in histone eviction, we will generate conditionally deficient mice and study histone cleavage in relation to turnover during spermatogenesis. Likewise, we will generate transgenic mice expressing uncleavable point mutants of H3 proteins in an inducible manner. 3.To elucidate the role of paternally inherited histone proteins in the regulation of early embryonic gene expression. We will use sperm from transgenic males heterozygous for the epitope tagged H3 transgenes (described in Aim 1) to produce pre-implantation embryos. Using microscopy approaches we will then track the presence of tagged H3 in genetically wild-type versus transgenic early embryos to determine whether and how long paternally inherited histones are retained on DNA during early embryogenesis. In addition, we will generate mice expressing H3 variants lacking crucial modified residues to examine the role of residue specific histone modifications in paternal chromatin inheritance. We will examine expression of genes associated with modified histones in sperm in two-cell stage embryos sired by these mice to determine if embryonic gene expression is affected. Expected value: The proposed study comprises a systematic genetic and biochemical analysis of nucleosome remodeling and turnover processes occurring during spermatogenesis. We expect to answer the highly relevant and open question whether nucleosomes in mouse spermatozoa mediate transmission of epigenetic information between generations. We further anticipate to obtain detailed insights into the molecular mechanism underlying nucleosome eviction during spermatid differentiation, leading to the production of functional spermatozoa.
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