Histone deacetylase 1 maintains lineage integrity through histone acetylome refinement during early embryogenesis
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In this manuscript, the authors investigate the role of histone deacetylases in the spatial epigenetic control of zygotic gene expression in early gastrulation. They discover HDAC1 binding is maternally-controlled and that inhibition of histone acetylation blocks gastrulation and disrupts cell lineage integrity, tied to both positive and negative regulatory effects on gene transcription in space and time. The study contributes to a growing body of evidence that highlights a central role of histone acetylation-deacetylation dynamics in epigenetic regulation of gene expression and cell fating in early tissue patterning of the embryo.
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Abstract
Histone acetylation is a pivotal epigenetic modification that controls chromatin structure and regulates gene expression. It plays an essential role in modulating zygotic transcription and cell lineage specification of developing embryos. While the outcomes of many inductive signals have been described to require enzymatic activities of histone acetyltransferases and deacetylases (HDACs), the mechanisms by which HDACs confine the utilization of the zygotic genome remain to be elucidated. Here, we show that histone deacetylase 1 (Hdac1) progressively binds to the zygotic genome from mid-blastula and onward. The recruitment of Hdac1 to the genome at blastula is instructed maternally. Cis -regulatory modules (CRMs) bound by Hdac1 possess epigenetic signatures underlying distinct functions. We highlight a dual function model of Hdac1 where Hdac1 not only represses gene expression by sustaining a histone hypoacetylation state on inactive chromatin, but also maintains gene expression through participating in dynamic histone acetylation–deacetylation cycles on active chromatin. As a result, Hdac1 maintains differential histone acetylation states of bound CRMs between different germ layers and reinforces the transcriptional program underlying cell lineage identities, both in time and space. Taken together, our study reveals a comprehensive role for Hdac1 during early vertebrate embryogenesis.
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eLife assessment
In this manuscript, the authors investigate the role of histone deacetylases in the spatial epigenetic control of zygotic gene expression in early gastrulation. They discover HDAC1 binding is maternally-controlled and that inhibition of histone acetylation blocks gastrulation and disrupts cell lineage integrity, tied to both positive and negative regulatory effects on gene transcription in space and time. The study contributes to a growing body of evidence that highlights a central role of histone acetylation-deacetylation dynamics in epigenetic regulation of gene expression and cell fating in early tissue patterning of the embryo.
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Reviewer #1 (Public Review):
In this study, the authors characterize the impact of histone deacetylation on spatial regulation of gene expression in the early gastrula embryo. They utilize Xenopus tropicalis as a vertebrate model embryo and focus on maternal HDAC1 and HDAC2 deacetylases to characterize the regulatory role of histone acetylation on zygotic transcription. In particular, they are interested in whether this epigenetic mark positively or negatively regulates gene expression for the presumptive germ layer and contributes spatially to cell lineage integrity in gastrulation.
Using gene expression analysis, they find that HDAC1 and HDAC2 are present maternally in the egg and throughout blastula and gastrula stages. By performing HDAC1 ChIP-Seq, they find that the deacetylase is already bound as early as the Stage 8 blastula - …
Reviewer #1 (Public Review):
In this study, the authors characterize the impact of histone deacetylation on spatial regulation of gene expression in the early gastrula embryo. They utilize Xenopus tropicalis as a vertebrate model embryo and focus on maternal HDAC1 and HDAC2 deacetylases to characterize the regulatory role of histone acetylation on zygotic transcription. In particular, they are interested in whether this epigenetic mark positively or negatively regulates gene expression for the presumptive germ layer and contributes spatially to cell lineage integrity in gastrulation.
Using gene expression analysis, they find that HDAC1 and HDAC2 are present maternally in the egg and throughout blastula and gastrula stages. By performing HDAC1 ChIP-Seq, they find that the deacetylase is already bound as early as the Stage 8 blastula - time of genome activation - and that HDAC1 peaks located within promoter regions generally increase over time from blastula to early gastrula, Stage 10.5. Interestingly, the binding of HDAC1 is not dependent on the zygotic transcript, as HDAC1 ChIP-seq peaks show little difference upon alpha-amanitin treatment. Many of the HDAC1 peaks correlate with peaks of both FoxH1 and Sox3, suggesting their role in its deacetylase recruitment to the genome. Examination of epigenetic signatures of HDAC1 bound regions using previously published datasets identifies distinct chromatin binding categories: authors find a strong correlation with H3K27-Ac and pan-H3Kac, and that HDAC1 generally binds to regions free of repressive marks such as H3K9-me3. The authors find that a majority of HDAC1 peaks contain H3K27Ac but not H3K37me3 peaks and approximately ten percent of HDAC1 loci have both activating and repressive marks.
The authors investigate a functional role for histone deacetylation by inhibiting it, using the broad inhibitor TSA, and HDAC1 specific inhibitor VPA. Importantly, they spatially characterize pan-H3K acetylation and gene expression changes in animal cag (AC) and vegetal mass (VG) regions on the embryo. These are very useful datasets that provide new insights into how histone acetylation is tied to the maintenance of lineage integrity. At a global level, they find that TSA inhibition leads to gastrulation arrest and leads to widespread upregulation of H3K acetylation (pan-H3Kac); suggesting that proper regulation of histone acetylation is required for development. Further, they find that previously repressed regions, marked by H3K27me3 show the most upregulation of pan-H3Kac upon TSA treatment. Regionally, they find a number of interesting results upon inhibition of histone acetylation. First, TSA treatment causes dysregulation - upregulation - of the animal cap (AC) pan-H3Kac peaks in vegetal mass (VG), and upregulation of VG peaks in the animal cap. This suggests that lineage specifically is likely maintained in part by HDAC-mediated de-acetylation of germ layer genes. Gene expression characterization in AC and VG explants +/- TSA treatment supports this conclusion as inappropriate upregulation of VG gene expression is found in AC and inappropriate upregulation of AC genes is found in VG. Somewhat surprisingly, HDACs also appears to play a positive regulatory role in germ layer expression. Focusing on genes near HDAC1 peaks containing H3K27Ac, the authors show that genes downregulated upon TSA treatment tend to be spatially restricted; downregulated genes in AC tended to be AC genes and downregulated genes in VG tended to be VG genes. This suggests that HDACs play both positive and negative roles in regulating germ layer expression in the gastrula.
Strengths of the work include the demonstration that histone deacetylase HDAC1 binds to the genome by the onset of genome activation, accumulates in promoters as the embryo develops through early gastrula, and that inhibition of histone deacetylation disrupts germ layer lineage integrity. New datasets include ChIP-seq of HDAC1 from blastula to gastrula, panH3Kac ChIP-seq within animal and vegetal regions of the embryo, and regional RNA-seq of embryos with and without TSA inhibition of histone acetylation. This study helps demonstrate and clarify that HDAC enzymes play both a positive and negative role in gene expression regulation, and that histone acetylation is required to maintain spatial specificity of germ layer expression in gastrula. Some of the weaknesses of the work include the correlative nature of the experiments and missing analysis. Overall, the research is interesting and impactful, contributing to a growing body of work about the role of histone acetylation in the spatial regulation of earliest cell fate decisions in the embryo.
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Reviewer #2 (Public Review):
This manuscript dives deeply into the localized binding and potential function of the Histone deacetylase Hdac1, the major HDAC expressed in early frog development. The stage-specific binding of Hdac1 changes during early development, correlating with the binding due to maternal factors, then zygotically generally activated or generally repressed genes, and also genes that can be either activated or repressed depending on their context. The protein appears not to bind to constitutive heterochromatin.
The study pursues how the binding changes on Animal Cap versus Vegetal mass expressed genes, and studies how inhibition of Hdac1 with TSA or VPA affects the degree of acetylation and expression. Perhaps the most interesting finding is that inhibition of Hdac1 has large effects on the acetylation and expression …
Reviewer #2 (Public Review):
This manuscript dives deeply into the localized binding and potential function of the Histone deacetylase Hdac1, the major HDAC expressed in early frog development. The stage-specific binding of Hdac1 changes during early development, correlating with the binding due to maternal factors, then zygotically generally activated or generally repressed genes, and also genes that can be either activated or repressed depending on their context. The protein appears not to bind to constitutive heterochromatin.
The study pursues how the binding changes on Animal Cap versus Vegetal mass expressed genes, and studies how inhibition of Hdac1 with TSA or VPA affects the degree of acetylation and expression. Perhaps the most interesting finding is that inhibition of Hdac1 has large effects on the acetylation and expression of inactive, but facultatively expressed genes, while it has smaller hyperacetylation effects on already active facultatively expressed genes; despite a modest stimulation of the already stimulatory effects of acetylation, the additional acetylation correlates with inhibition of expression of this subset of genes. This result is clearly documented with embryonic region-specific effects on facultatively expressed genes. The effect on inactive genes fits with the general idea that Hdac1 is repressive, but the effect on already acetylated genes is not so easily explained, though some models are proposed.
The overall findings are important background for developmental and chromatin biologists because they add to the documentation of the correlations between acetylation, deacetylation, and expression of genes in development. The correlations allow the inference of potential functions, though these are not tested other than by pharmacological inhibition of Hdac1.
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Reviewer #3 (Public Review):
This paper investigates how the epigenetic landscape is set up during early frog embryogenesis focusing on the role of the histone deacetylase, HDAC1, in the regulation of histone acetylation around the period of Zygotic gene activation. The authors document the progressive binding of HDAC1 to the embryonic genome around the time of ZGA and on genomic sites harbouring binding motifs for maternally provided transcription factors. The authors classify HDAC1 binding sites based on their association with different epigenetic markings on H3K27 (acetylation and/or methylation) in embryonic chromatin. They infer from the observed co-occurrence of "incompatible" acetylation and methylation marks on H3K27 residue on a subset of HDAC1 binding sites, that these H3K27 modifications occur in different parts of the …
Reviewer #3 (Public Review):
This paper investigates how the epigenetic landscape is set up during early frog embryogenesis focusing on the role of the histone deacetylase, HDAC1, in the regulation of histone acetylation around the period of Zygotic gene activation. The authors document the progressive binding of HDAC1 to the embryonic genome around the time of ZGA and on genomic sites harbouring binding motifs for maternally provided transcription factors. The authors classify HDAC1 binding sites based on their association with different epigenetic markings on H3K27 (acetylation and/or methylation) in embryonic chromatin. They infer from the observed co-occurrence of "incompatible" acetylation and methylation marks on H3K27 residue on a subset of HDAC1 binding sites, that these H3K27 modifications occur in different parts of the embryos. Subsequently, they inhibit histone deacetylase activity by TSA and document its impact on the genomic distribution of acetylated histones as well as transcriptional deregulation in explant from different parts of the embryos. By cross comparing these data to the different classes of HDAC1-associated genomic regions, they conclude that HDAC1 is involved in the spatial regulation of embryonic gene expression. Altogether this work reveals how maternally provided transcription factors could direct chromatin modifiers to shape the epigenome of the developing embryos. The work however relies mostly on indirect evidence and it would be important in particular to confirm (i) that maternal factors are indeed required for HDAC1 targeting to chromatin and (ii) that the documented effect of TSA treatment is mediated through its inhibition of HDAC1.
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