Dnmt3a knockout in excitatory neurons impairs postnatal synapse maturation and increases the repressive histone modification H3K27me3
- University of California, San Diego, United States
- Salk Institute for Biological Studies, United States
- Howard Hughes Medical Institute, Salk Institute for Biological Studies, United States
Abstract
Two epigenetic pathways of transcriptional repression, DNA methylation and Polycomb repressive complex 2 (PRC2) are known to regulate neuronal development and function. However, their respective contributions to brain maturation are unknown. We found that conditional loss of the de novo DNA methyltransferase Dnmt3a in mouse excitatory neurons altered expression of synapse-related genes, stunted synapse maturation, and impaired working memory and social interest. At the genomic level, loss of Dnmt3a abolished postnatal accumulation of CG and non-CG DNA methylation, leaving adult neurons with an unmethylated, fetal-like epigenomic pattern at ~222,000 genomic regions. The PRC2-associated histone modification, H3K27me3, increased at many of these sites. Our data support a dynamic interaction between two fundamental modes of epigenetic repression during postnatal maturation of excitatory neurons, which together confer robustness on neuronal regulation.
Data availability
All sequencing data are available in the Gene Expression Omnibus under accession GSE141587. A genome browser displaying the sequencing data is available at https://brainome.ucsd.edu/annoj_private/mm_dnmt3a_ko/
-
Transcriptomic and epigenetic disruptions in excitatory neurons in Dnmt3a conditional knockout mouseNCBI Gene Expression Omnibus, GSE141587.
-
Global epigenomic reconfiguration during mammalian brain developmentNCBI Gene Expression Omnibus, GSE47966.
Article and author information
Author details
Antonio Pinto-Duarte
Terrence J Sejnowski
M Margarita Behrens
Funding
National Institute of Mental Health (R01MH112763)
- Joseph R Ecker
- Eran A Mukamel
- M Margarita Behrens
Kavli Foundation
- Antonio Pinto-Duarte
- Susan B Powell
- M Margarita Behrens
Howard Hughes Medical Institute
- Joseph R Ecker
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: All animal procedures were conducted in accordance with the guidelines of the American Association for the Accreditation of Laboratory Animal Care and were approved by the Salk Institute for Biological Studies Institutional Animal Care and Use Committee (Protocol number 18-00006).
Copyright
© 2022, Li et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,865
- views
-
- 568
- downloads
-
- 10
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Dnmt3a knockout in excitatory neurons impairs postnatal synapse maturation and increases the repressive histone modification H3K27me3
eLife 11:e66909.
https://doi.org/10.7554/eLife.66909
Further reading
-
- Genetics and Genomics
Due to proteostasis stress induced by aging or disease, misfolded proteins can form toxic intermediate species of aggregates and eventually mature into less toxic inclusion bodies (IBs). Here, using a yeast imaging-based screen, we identified 84 potential synphilin-1 (SY1) IB regulators and isolated the conserved sphingolipid metabolic components in the most enriched groups. Furthermore, we show that, in both yeast cells and mammalian cells, SY1 IBs are associated with mitochondria. Pharmacological inhibition of the sphingolipid metabolism pathway or knockout of its key genes results in a delayed IB maturation and increased SY1 cytotoxicity. We postulate that SY1 IB matures by association with the mitochondrion membrane, and that sphingolipids stimulate the maturation via their membrane-modulating function and thereby protecting cells from SY1 cytotoxicity. Our findings identify a conserved cellular component essential for IB maturation and suggest a mechanism by which cells may detoxify the pathogenic protein aggregates through forming mitochondrion-associated IBs.
-
- Genetics and Genomics
Advances in single-cell sequencing technologies have provided novel insights into the dynamics of gene expression and cellular heterogeneity within tissues and have enabled the construction of transcriptomic cell atlases. However, linking anatomical information to transcriptomic data and positively identifying the cell types that correspond to gene expression clusters in single-cell sequencing data sets remains a challenge. We describe a straightforward genetic barcoding approach that takes advantage of the powerful genetic tools in Drosophila to allow in vivo tagging of defined cell populations. This method, called Targeted Genetically-Encoded Multiplexing (TaG-EM), involves inserting a DNA barcode just upstream of the polyadenylation site in a Gal4-inducible UAS-GFP construct so that the barcode sequence can be read out during single-cell sequencing, labeling a cell population of interest. By creating many such independently barcoded fly strains, TaG-EM enables positive identification of cell types in cell atlas projects, identification of multiplet droplets, and barcoding of experimental timepoints, conditions, and replicates. Furthermore, we demonstrate that TaG-EM barcodes can be read out using next-generation sequencing to facilitate population-scale behavioral measurements. Thus, TaG-EM has the potential to enable large-scale behavioral screens in addition to improving the ability to multiplex and reliably annotate single-cell transcriptomic experiments.
