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DNA methylation contributes to the maintenance of genomic integrity in somatic cells, in part through the silencing of transposable elements. In this study, we use CRISPR-Cas9 technology to delete DNMT1 , the DNA methyltransferase key for DNA methylation maintenance, in human neural progenitor cells (hNPCs). We observe that inactivation of DNMT1 in hNPCs results in viable, proliferating cells despite a global loss of DNA CpG-methylation. DNA demethylation leads to specific transcriptional activation and chromatin remodeling of evolutionarily young, hominoid-specific LINE-1 elements (L1s), while older L1s and other classes of transposable elements remain silent. The activated L1s act as alternative promoters for many protein-coding genes involved in neuronal functions, revealing a hominoid-specific L1-based transcriptional network controlled by DNA methylation that influences neuronal protein-coding genes. Our results provide mechanistic insight into the role of DNA methylation in silencing transposable elements in somatic human cells, as well as further implicating L1s in human brain development and disease. DNA methylation plays an important role in silencing transposable elements. Here the authors find that loss of DNMT1 and DNA methylation leads to transcriptional activation and chromatin remodelling of evolutionarily young—hominoid-specific —LINE-1 elements which then act as alternative promoters for neuronal genes.

Age is a risk factor for hematologic malignancies. Attributes of the aging hematopoietic system include increased myelopoiesis, impaired adaptive immunity, and a functional decline of the hematopoietic stem cells (HSCs) that maintain hematopoiesis. Changes in the composition of diverse HSC subsets have been suggested to be responsible for age-related alterations, however, the underlying regulatory mechanisms are incompletely understood in the context of HSC heterogeneity. In this study, we investigated how distinct HSC subsets, separated by CD49b, functionally and molecularly change their behavior with age. We demonstrate that the lineage differentiation of both lymphoid-biased and myeloid-biased HSC subsets progressively shifts to a higher myeloid cellular output during aging. In parallel, we show that HSCs selectively undergo age-dependent gene expression and gene regulatory changes in a progressive manner, which is initiated already in the juvenile stage. Overall, our studies suggest that aging intrinsically alters both cellular and molecular properties of HSCs. Aging is a risk factor for blood malignancies, leading to increased myelopoiesis and impaired immunity. Here, the authors show that aging alters cellular and molecular properties of HSC subsets, with changes starting as early as the juvenile stage.

Background RNA sequencing has become the mainstay for studies of gene expression. Still, analysis of rare cells with random hexamer priming – to allow analysis of a broader range of transcripts – remains challenging. Results We here describe a tagmentation-based, rRNA blocked, random hexamer primed RNAseq approach (T-RHEX-RNAseq) for generating stranded RNAseq libraries from very low numbers of FACS sorted cells without RNA purification steps. Conclusion T-RHEX-RNAseq provides an easy-to-use, time efficient and automation compatible method for generating stranded RNAseq libraries from rare cells.

Cohesin is a major structural component of mammalian genomes and is required to maintain loop structures. While acute depletion in short-term culture models suggests a limited importance of cohesin for steady-state transcriptional circuits, long-term studies are hampered by essential functions of cohesin during replication. Here, we study genome architecture in a postmitotic differentiation setting, the differentiation of human blood monocytes (MO). We profile and compare epigenetic, transcriptome and 3D conformation landscapes during MO differentiation (either into dendritic cells or macrophages) across the genome and detect numerous architectural changes, ranging from higher level compartments down to chromatin loops. Changes in loop structures correlate with cohesin-binding, as well as epigenetic and transcriptional changes during differentiation. Functional studies show that the siRNA-mediated depletion of cohesin (and to a lesser extent also CTCF) markedly disturbs loop structures and dysregulates genes and enhancers that are primarily regulated during normal MO differentiation. In addition, gene activation programs in cohesin-depleted MO-derived macrophages are disturbed. Our findings implicate an essential function of cohesin in controlling long-term, differentiation- and activation-associated gene expression programs. How chromatin structure and gene accessibility changes during monocyte differentiation is not clearly defined. Here the authors characterize the chromatin changes during macrophage or dendritic cell maturation from monocytes and the dependence of this upon cohesin and CTCF.

Background Chromatin immunoprecipitation coupled to sequencing (ChIP-seq) is widely used to map histone modifications and transcription factor binding on a genome-wide level. Results We present high-throughput ChIPmentation (HT-ChIPmentation) that eliminates the need for DNA purification prior to library amplification and reduces reverse-crosslinking time from hours to minutes. Conclusions The resulting workflow is easily established, extremely rapid, and compatible with requirements for very low numbers of FACS sorted cells, high-throughput applications and single day data generation.

Earlier work has shown that pre-B cells can be converted into macrophages by the transcription factor CCAAT/enhancer binding protein α at very high frequencies. Using this system, we performed a systematic analysis of whether during transdifferentiation the cells transiently reactivate progenitor-restricted genes or even retrodifferentiate. A transcriptome analysis of transdifferentiating cells showed that most genes are up- or down-regulated continuously, acquiring a macrophage phenotype within 5 d. In addition, we observed the transient reactivation of a subset of immature myeloid markers, as well as low levels of the progenitor markers Kit and FMS-like tyrosine kinase 3 and a few lineage-inappropriate genes. Importantly, however, we were unable to observe the reexpression of cell-surface marker combinations that characterize hematopoietic stem and progenitor cells, including c-Kit and FMS-like tyrosine kinase 3, even when CAAT/enhancer binding protein α was activated in pre-B cells under culture conditions that favor growth of hematopoietic stem and progenitor cells or when the transcription factor was activated in a time-limited fashion. Together, our findings are consistent with the notion that the conversion from pre-B cells to macrophages is mostly direct and does not involve overt retrodifferentiation.

PAIRED (PRD)-like homeobox genes belong to a class of predicted transcription factor genes. Several of these PRD-like homeobox genes have been predicted in silico from genomic sequence but until recently had no evidence of transcript expression. We found recently that nine PRD-like homeobox genes, ARGFX , CPHX1 , CPHX2 , DPRX , DUXA , DUXB , NOBOX, TPRX1 and TPRX2 , were expressed in human preimplantation embryos. In the current study we characterized these PRD-like homeobox genes in depth and studied their functions as transcription factors. We cloned multiple transcript variants from human embryos and showed that the expression of these genes is specific to embryos and pluripotent stem cells. Overexpression of the genes in human embryonic stem cells confirmed their roles as transcription factors as either activators (CPHX1, CPHX2, ARGFX) or repressors (DPRX, DUXA, TPRX2) with distinct targets that could be explained by the amino acid sequence in homeodomain. Some PRD-like homeodomain transcription factors had high concordance of target genes and showed enrichment for both developmentally important gene sets and a 36 bp DNA recognition motif implicated in Embryo Genome Activation (EGA). Our data implicate a role for these previously uncharacterized PRD-like homeodomain proteins in the regulation of human embryo genome activation and preimplantation embryo development.

The gene desert upstream of the MYC oncogene on chromosome 8q24 contains susceptibility loci for several major forms of human cancer. The region shows high conservation between human and mouse and contains multiple MYC enhancers that are activated in tumor cells. However, the role of this region in normal development has not been addressed. Here we show that a 538 kb deletion of the entire MYC upstream super-enhancer region in mice results in 50% to 80% decrease in Myc expression in multiple tissues. The mice are viable and show no overt phenotype. However, they are resistant to tumorigenesis, and most normal cells isolated from them grow slowly in culture. These results reveal that only cells whose MYC activity is increased by serum or oncogenic driver mutations depend on the 8q24 super-enhancer region, and indicate that targeting the activity of this element is a promising strategy of cancer chemoprevention and therapy. Our cells each contain close to 20,000 genes, which provide the instructions needed to build our bodies and keep us alive. At any one time in the life of the cell, only some of these genes are active. The activity of each gene is constantly regulated to allow the cell to respond to changes in its environment. Enhancers are sections of DNA, outside of the genes, that act as molecular switches controlling the activity of genes. A gene can have many such enhancers; each enhancer is linked to a particular set of signals and having multiple enhancers allows the same gene to be activated by different signals in different tissues in the body. Changes to enhancers can have serious consequences. By altering the activity of genes, an enhancer can have widespread effects on the health and behavior of a cell, including transforming it from healthy to cancerous. The small differences in enhancers also make some people more susceptible to cancers than others. If we can identify enhancers whose activity is commonly altered in cancers, it could be possible to target them through treatment. Yet, it is not clear whether targeting enhancers in this way could be effectively used to treat cancer without damaging healthy cells. Now, Dave, Sur et al. have examined a large enhancer region with known links to several different cancers – including prostate, breast and colon cancers – to uncover whether it also plays a critical role in healthy cells and if it could be safely targeted for treatment. The region has multiple enhancers for a cancer-linked gene called MYC and is implicated in many cancer-associated deaths every year. This particular enhancer region is found in both humans and mice, which share many genes in common. Using genetic engineering, Dave, Sur et al. removed this enhancer region from the genetic information of a group of mice. The experiment showed that mice without the enhancer region were completely healthy. Also, when tested for cancer development, these mice were much less susceptible to several major types of cancer. This investigation reveals that it may be possible to create drugs to shut down or inhibit certain enhancers to prevent or treat cancer without damaging healthy cells. However, this is currently just one example in mice under laboratory conditions. Further research is needed to determine if a similar approach can be developed to treat patients in the clinic.

Nucleosome assembly proteins (NAPs) are histone chaperones with an important role in chromatin structure and epigenetic regulation of gene expression. We find that high gene expression levels of mouse Nap1l3 are restricted to haematopoietic stem cells (HSCs) in mice. Importantly, with shRNA or CRISPR-Cas9 mediated loss of function of mouse Nap1l3 and with overexpression of the gene, the number of colony-forming cells and myeloid progenitor cells in vitro are reduced. This manifests as a striking decrease in the number of HSCs, which reduces their reconstituting activities in vivo . Downregulation of human NAP1L3 in umbilical cord blood (UCB) HSCs impairs the maintenance and proliferation of HSCs both in vitro and in vivo . NAP1L3 downregulation in UCB HSCs causes an arrest in the G0 phase of cell cycle progression and induces gene expression signatures that significantly correlate with downregulation of gene sets involved in cell cycle regulation, including E2F and MYC target genes. Moreover, we demonstrate that HOXA3 and HOXA5 genes are markedly upregulated when NAP1L3 is suppressed in UCB HSCs. Taken together, our findings establish an important role for NAP1L3 in HSC homeostasis and haematopoietic differentiation.

Multiple myeloma (MM) is characterized by an uninhibited, clonal growth of plasma cells. While first-degree relatives of patients with MM show an increased risk of MM, the genetic basis of inherited MM susceptibility is incompletely understood. Here we report a genome-wide association study in the Nordic region identifying a novel MM risk locus at ELL2 (rs56219066T; odds ratio (OR)=1.25; P =9.6 × 10 −10 ). This gene encodes a stoichiometrically limiting component of the super-elongation complex that drives secretory-specific immunoglobulin mRNA production and transcriptional regulation in plasma cells. We find that the MM risk allele harbours a Thr298Ala missense variant in an ELL2 domain required for transcription elongation. Consistent with a hypomorphic effect, we find that the MM risk allele also associates with reduced levels of immunoglobulin A (IgA) and G (IgG) in healthy subjects ( P =8.6 × 10 −9 and P =6.4 × 10 −3 , respectively) and, potentially, with an increased risk of bacterial meningitis (OR=1.30; P =0.0024). Multiple myeloma is an incurable and fatal disease characterized by uninhibited growth of plasma cells in the bone marrow. Here, Swaminathan et al. conduct a genome-wide association study and identify a novel risk locus at ELL2 , which encodes a key component of the super-elongation complex.