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Subcerebral projection neurons are controlled by polycomb mediated epigenetics
Various neuronal and glial cells found in the neocortex arise from the differentiation of multipotent neural precursor cells. However, the molecular mechanism controlling the differentiation of neural precursor cells into various mature neuronal subtypes, such as subcerebral projection neurons (SCPNs), is not well understood. This limits our understanding of normal and abnormal brain development.
In this paper Yukiko Gotoh and colleagues seek to understand the epigenetic mechanisms controlling the differentiation of neural precursor cells into SCPNs. To do this they performed H3K27me3 chromatin immunoprecipitation on normal, Ring1B-deficient (a component of the polycomb group), undifferentiated and differentiating neural precursor cells.
Doing so they found that:
They conclude that Ring1B epigenetically down-regulates Fezf2 expression thereby controlling SCPN number. Understanding this offers insight into normal brain development. Additionally, since SCPNs degrade in the case of some neurodegenerative diseases and spinal cord injuries, understanding how SCPNs are down-regulated may also aid the development of up-regulating therapeutics.
Read the full report in Development, October 2014.
RNA feedback loops create disease cell-specificity
A perplexing question in disease genetics has been why do mutations in ubiquitously expressed housekeeping genes affect only certain body systems? Spinocerebellar ataxia type 7 (SCA7) is a rare neurodegenerative disorder caused by a CAG-repeat expansion in ATXN7, an essential component of the STAGA transcription coactivation complex. SCA7 is characterized by the degeneration of the retinal macula and cerebellum, however, ATXN7 is expressed at high levels in many non-neural tissues.
Ana C. Marques and colleagues at the University of Oxford hypothesized that intercellular differences in non-coding RNAs might account for the disparity of affected cell types in SCA7. Antisense long non-coding RNA perturbation is key in the pathology of SCA8 and other neurodegenerative disorders.
Using both human and mouse paradigms, they found:
STAGA, miR-124, lnc-SCA7 and ATXN7 form a tissue-specific crosstalk network impacted in SCA7. The authors showed that in SCA7, the reduced transcriptional activity of the STAGA complex leads to a decrease in miR-124 expression. Lower miR-124 levels then contribute to the increased abundance of ATXN7 transcripts in a feedback loop. The authors propose that in SCA7, increased mutant ATXN7 protein levels result from cell-specific post-transcriptional derepression caused by decreased miR-124.
Read the full report in Nature Structural and Molecular Biology, October 2014.
Register for our upcoming event, Non-Coding RNA: New Mechanisms and Approaches, May 18–19, Boston.
Novel histone acetylation reader mediates methylation
A key mechanism involved in chromatin regulation of gene expression is the recognition of modified histones by reader proteins. Histone acetylation decreases the electrostatic interaction between DNA and histone proteins in the nucleosome and can also serve as a docking site for such readers. Whereas histone methylation marks are read by a variety of proteins, few are capable of recognizing acetylated lysine residues.
Scientists from Tsinghua University, Baylor College of Medicine and the University of Texas MD Anderson Cancer Center, led by Dr. Xiaobing Shi investigated the role of an evolutionarily conserved AF9 YEATS domain that functions as a novel acetyl lysine-binding module with a mechanism distinct from other histone acetylation readers. AF9 binds to several histone acetylation marks, most strongly to H3K9 and to a lesser extent to H3K27 and H3K18 acetylation.
The authors found a genome-wide association between AF9 and H3K9 acetylation using reciprocal AF9 and H3K9ac ChIP-Seq and demonstrated a direct link between histone acetylation and the recruitment of methyltransferase DOT1L that targets H3K79 for methylation and activation of gene transcription.
Here are the important results from the study:
The mode of reading histone acetylation used by the YEATS domain represents a novel mechanism of acetyl lysine recognition that is distinct from other readers. Furthermore, the ability of this domain to regulate gene expression via the recruitment of DOT1L methyltransferase makes YEATS domains potential therapeutic targets for cancer treatment.
Read full report in Cell, October 2014.
DNA methylation during the life of cardiac cells
Heart development is a highly regulated, critical process in development and disease. Cardiomyocytes undergo few cell divisions during development and have limited regenerative capacity. They are reliant on gene expression programs to guide their development to mature cardiac tissue. These programs appear to be influenced by epigenetic marks such as DNA methylation, however, their precise role remains unclear.
Lutz Hein and colleges from the University of Freiburg, Germany sought to investigate the role of DNA methylation in heart development and disease. They investigated whole-genome DNA methylation in mouse cardiomyocytes from neonatal, healthy adult and failing adult hearts.
Using whole-genome bisulfite sequencing on cardiomyocytes isolated by flow cytometry, the authors found:
The results of this study establish DNA methylation as a key regulator of cardiomyocyte development. The comprehensive data reported here can serve as a valuable resource for studying heart development and disease. The study also includes several novel findings, for example, the wave of demethylation occurring at gene bodies during development.
Read the full report in Nature Communications, October 2014.
Characterizing genes by targeted knockdown in the brain
The clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated endonuclease (Cas)9 from Streptococcus pyogenes (SpCas9) is a powerful system for genome editing in mammalian cells. It can be used to induce insertion/deletion frame shift mutations and protein depletion. Such experiments can elucidate gene function in vivo, however, developing technologies to deliver SpCas9 complexes into specific cells in live animals has been challenging.
Feng Zhang and colleagues from the Broad Institute of MIT and Harvard sought to deliver SpCas9 and guide RNAs into the brain using adeno-associated viral (AAV) vectors. Such a system would allow for unprecedented control over gene-knock down studies in time, space, and cell-type and allow greater insight into brain function.
Using adult mouse brain, the authors found:
The diversity of proof-of-concept experiments presented in this paper illustrate the versatility and utility of SpCas9 system in determining gene function the mammalian brain.
See the full report in Nature Biotechnology, October 2014.
The work used Abcam’s anti-GFAP antibody (ab4674).
EZH2 targets microRNAs to avoid apoptosis
The polycomb protein EZH2 acts as a promoter of cell proliferation and metastasis, and is significantly overexpressed in prostate cancers. While the oncogenic role of EZH2 is thoroughly studied, EZH2’s influence on the suppression of apoptosis is a recently uncovered mechanism that has yet to be fully explored. In this recent study, Dr. Bin Guo and a research team at North Dakota State University demonstrated that EZH2 directs the epigenetic silencing of miR-205 and miR-31 (known proapoptotic microRNAs) thereby, avoiding apoptosis in prostate cancer.
The scientists conducted a series of experiments using microRNA mimics and inhibitors, in addition to chromatin immunoprecipitation (ChIP) to investigate how EZH2 affects apoptosis.
Here is what they found:
The authors conclude that EZH2 coordinates the epigenetic silencing of miR-205 and miR-31, which protects cancer cells from chemotherapy-induced apoptosis. The scientists theorize that it would be possible to take advantage of this new data by targeting EZH2 with compounds, such as GSK126, which could potentially induce apoptosis directly or sensitize cancer cells to traditional chemotherapy treatments.
Read the full report in Cell Death and Disease, October 2014.
For more information on the role of polycombs in cancer, sign up for our free webinar with guest speaker Adrian Bracken.
PRC2 activity is essential for the maintenance of cell fate during differentiation
Loss of polycomb repressive complex 2 (PRC2) and its catalyzed mark H3K26me3 has been shown to silence developmental programs in embryonic stem cells (ESCs). However the inability of PRC2 ESC mutants to properly differentiate has meant that the role of PCR2 during lineage commitment and differentiation remains unexplored.
To address this, Laurie Boyer and colleagues at Massachusetts Institute of Technology used ChIP-seq, qRT-PCR and western blotting to analyze several mutant ESC lines that maintain varying levels of H3K26me3.
The researchers found that:
The authors provide novel insights into the role of PRC2 in mammalian development and its effect on gene expression during lineage commitment. They suggest that the low-level seeding of inappropriate DNA methylation may lead to further epigenetic instability in differentiated cells, explaining the molecular underpinning of PRC2 in cancer.
DNA loops control repressive gene terminator marks
R-loops are single-stranded DNA loops that form when the DNA double helix is invaded by a nascent RNA transcript. Their role in the cell is controversial. It has long been believed that they are damaging by exposing single-stranded DNA to mutation. However, they may also serve a functional role; they form over CpG islands and terminators and appear to be necessary for heterochromatin formation.
Nicholas J. Proudfoot and colleagues at the University of Oxford sought to investigate the relationship between R-loops and epigenetic marks in RNA interference-mediated termination. RNAi is a key mechanism of transcriptional silencing, which recruits methyltransferases (G9a) heterochromatin proteins (HP1γ) to facilitate transcriptional termination. This group previously showed that R-loops are enriched over G-rich terminator elements and facilitate RNA polymerase pausing during transcriptional termination.
Working at the beta-actin promoter in mammalian (human HeLa and mouse MEF) cell lines, the authors found that:
These data indicate that R-loops formed at the beta-actin pause element trigger antisense transcription, assembly of the RNAi apparatus, deposition of H3K9me2, and HP1γ recruitment. Further, the whole-genome data suggest that this mechanism likely applies to a large-subset of genes. Thus R-loops appear to be a key functional structure in the epigenetic regulation of transcription.
Read the full report in Letters to Nature, October 2014.
Effects of prenatal stress on DNA methylation are dependent on 5-Htt genotype
Exposure to prenatal stress (PS) affects fetal brain development and increases the risk of psychopathology later in life. The behavioral effects of PS have previously been shown to be partly dependent on a serotonin transporter gene (5-HTT/SLC6A4)-linked polymorphism.
To understand the molecular mechanisms behind this gene-by-environment interaction, researchers from the University of Wuerzburg in Germany and Maastricht University in the Netherlands sought to examine the role of DNA methylation in mediating differential gene expression in the offspring of 5-HTT-deficient mice subjected to PS.
Performing genome-wide hippocampal DNA methylation screening, the authors found that:
The authors conclude that effects of PS on DNA methylation in the hippocampus are partially dependent on 5-Htt genotype.These new data form the basis for future research investigating the interaction between prenatal stress, 5-Htt genotype and developmental epigenetic programming.
Read the full report in Translational Psychiatry, October 2014