H3.3-G34W in giant cell tumor of bone functionally aligns with the exon choice repressor hnRNPA1L2.

RNA processing is an essential post-transcriptional phenomenon that provides the necessary complexity of transcript diversity prior to translation. Aberrations in this process could contribute to tumourigenesis, and we have previously reported increased splicing alterations in giant cell tumor of bone (GCTB), which carries mutations in the histone variant H3.3 encoding glycine 34 substituted for tryptophan (H3.3-G34W). G34W interacts with several splicing factors, most notably the trans-acting splicing factor hnRNPA1L2. To gain a deeper understanding of RNA processing in GCTB and isogenic HeLa cells with H3.3-G34W, we generated RNA-immunoprecipitation sequencing data from hnRNPA1L2 and H3.3-G34W associated RNAs, which showed that 80% overlapped across genic regions and were frequently annotated as E2F transcription factor binding sites. Splicing aberrations in both GCTB and HeLa cells with H3.3-G34W were significantly enriched for known hnRNPA1L2 binding motifs (p value < 0.01). This splicing aberration differed from hnRNPA1L2 knockouts, which showed alterations independent of H3.3-G34W. Of functional significance, hnRNPA1L2 was redistributed to closely match the H3.3 pattern, likely driven by G34W, and to loci not occupied in normal parental cells. Taken together, our data reveal a functional overlap between hnRNPA1L2 and H3.3-G34W with likely significant consequences for RNA processing during GCTB pathogenesis. This provides novel opportunities for therapeutic intervention in future modus operandi.

Anti-obesity effect of butyrate links to modulation of gut microbiome and epigenetic regulation of muscular circadian clock.

The role of the muscle circadian clock in regulating oxidative metabolism exerts a significant influence on whole-body energy metabolism; however, research on the connection between the muscle circadian clock and obesity is limited. Moreover, there is a lack of studies demonstrating the regulatory effects of dietary butyrate on muscle circadian clock and the resulting antiobesity effects. This study aimed to investigate the impacts of dietary butyrate on metabolic and microbiome alterations and muscle circadian clock in a diet-induced obesity model. Male Sprague-Dawley rats were fed a high-fat diet with or without butyrate. Gut microbiota and serum metabolome were analyzed, and molecular changes were examined using tissues and a cell line. Further correlation analysis was performed on butyrate-induced results. Butyrate supplementation reduced weight gain, even with increased food intake. Gut microbiome analysis revealed an increased abundance of Firmicutes in butyrate group. Serum metabolite profile in butyrate group exhibited reduced amino acid and increased fatty acid content. Muscle circadian clock genes were upregulated, resulting in increased transcription of fatty acid oxidation-related genes. In myoblast cells, butyrate also enhanced pan-histone acetylation via histone deacetylase inhibition, particularly modulating acetylation at the promoter of circadian clock genes. Correlation analysis revealed potential links between Firmicutes phylum, including certain genera within it, and butyrate-induced molecular changes in muscle as well as phenotypic alterations. The butyrate-driven effects on diet-induced obesity were associated with alterations in gut microbiota and a muscle-specific increase in histone acetylation, leading to the transcriptional activation of circadian clock genes and their controlled genes.

Clarifying the carbon balance recovery time after clear-cutting.

Clear-cutting of forest is causing large emissions of carbon dioxide immediately after the clear-cutting and an important question is how fast the forest can achieve carbon balance (CB) again and how long time it takes until the initial losses are compensated for (CCP). This question is particularly important for the discussion on how to use forests most efficiently in the climate change mitigation work considering the short remaining time frame there is in order to achieve the targets according to the Paris agreement. Knowledge of how net ecosystem exchange (NEE) varies over time is also important for determination of time for optimal carbon sequestration (OCS).

Joint optimization of land carbon uptake and albedo can help achieve moderate instantaneous and long-term cooling effects.

Both carbon dioxide uptake and albedo of the land surface affect global climate. However, climate change mitigation by increasing carbon uptake can cause a warming trade-off by decreasing albedo, with most research focusing on afforestation and its interaction with snow. Here, we present carbon uptake and albedo observations from 176 globally distributed flux stations. We demonstrate a gradual decline in maximum achievable annual albedo as carbon uptake increases, even within subgroups of non-forest and snow-free ecosystems. Based on a paired-site permutation approach, we quantify the likely impact of land use on carbon uptake and albedo. Shifting to the maximum attainable carbon uptake at each site would likely cause moderate net global warming for the first approximately 20 years, followed by a strong cooling effect. A balanced policy co-optimizing carbon uptake and albedo is possible that avoids warming on any timescale, but results in a weaker long-term cooling effect.

Adenosine derivatives from Cordyceps exert antitumor effects against ovarian cancer cells through ENT1-mediated transport, induction of AMPK signaling, and consequent autophagic cell death.

Cordyceps militaris is rich in adenosine derivatives, including 3'-deoxyadenosine, also known as cordycepin. It has been reported for antitumor effects, but its underlying molecular mechanism has yet to be elucidated. We investigated how adenosine derivatives exerted antitumor effects against ovarian cancer using human ovarian cancer cells and a xenograft mouse model. Treatment with adenosine derivatives effectively resulted in cell death of ovarian cancer cells through AMPK activation and subsequently mTOR-mediated autophagic induction. Intriguingly, the effect required membrane transport of adenosine derivatives via ENT1, rather than ADORA-mediated cellular signaling. Our data suggest that adenosine derivatives may be an effective therapeutic intervention in ovarian cancer through induction of ENT1-AMPK-mTOR-mediated autophagic cell death.

Boreal forest soil carbon fluxes one year after a wildfire: Effects of burn severity and management.

The extreme 2018 hot drought that affected central and northern Europe led to the worst wildfire season in Sweden in over a century. The Ljusdal fire complex, the largest area burnt that year (8995 ha), offered a rare opportunity to quantify the combined impacts of wildfire and post-fire management on Scandinavian boreal forests. We present chamber measurements of soil CO2 and CH4  fluxes, soil microclimate and nutrient content from five Pinus sylvestris sites for the first growing season after the fire. We analysed the effects of three factors on forest soils: burn severity, salvage-logging and stand age. None of these caused significant differences in soil CH4 uptake. Soil respiration, however, declined significantly after a high-severity fire (complete tree mortality) but not after a low-severity fire (no tree mortality), despite substantial losses of the organic layer. Tree root respiration is thus key in determining post-fire soil CO2 emissions and may benefit, along with heterotrophic respiration, from the nutrient pulse after a low-severity fire. Salvage-logging after a high-severity fire had no significant effects on soil carbon fluxes, microclimate or nutrient content compared with leaving the dead trees standing, although differences are expected to emerge in the long term. In contrast, the impact of stand age was substantial: a young burnt stand experienced more extreme microclimate, lower soil nutrient supply and significantly lower soil respiration than a mature burnt stand, due to a thinner organic layer and the decade-long effects of a previous clear-cut and soil scarification. Disturbance history and burn severity are, therefore, important factors for predicting changes in the boreal forest carbon sink after wildfires. The presented short-term effects and ongoing monitoring will provide essential information for sustainable management strategies in response to the increasing risk of wildfire.

Accounting for all territorial emissions and sinks is important for development of climate mitigation policies.

The Paris agreement identifies the importance of the conservation, or better, increase of the land carbon sink. In this respect, the mitigation policies of many forest rich countries rely heavily on products from forests as well as on the land sink. Here we demonstrate that Sweden's land sink, which is critical in order to achieve zero net emissions by 2045 and negative emissions thereafter, is reduced to less than half when accounting for emissions from wetlands, lakes and running waters. This should have implications for the development of Sweden's mitigation policy. National as well as the emerging global inventory of sources and sinks need to consider the entire territory to allow accurate guidance of future mitigation of climate change.

Globally altered epigenetic landscape and delayed osteogenic differentiation in H3.3-G34W-mutant giant cell tumor of bone.

The neoplastic stromal cells of giant cell tumor of bone (GCTB) carry a mutation in H3F3A, leading to a mutant histone variant, H3.3-G34W, as a sole recurrent genetic alteration. We show that in patient-derived stromal cells H3.3-G34W is incorporated into the chromatin and associates with massive epigenetic alterations on the DNA methylation, chromatin accessibility and histone modification level, that can be partially recapitulated in an orthogonal cell line system by the introduction of H3.3-G34W. These epigenetic alterations affect mainly heterochromatic and bivalent regions and provide possible explanations for the genomic instability, as well as the osteolytic phenotype of GCTB. The mutation occurs in differentiating mesenchymal stem cells and associates with an impaired osteogenic differentiation. We propose that the observed epigenetic alterations reflect distinct differentiation stages of H3.3 WT and H3.3 MUT stromal cells and add to H3.3-G34W-associated changes.

Effects of drought and meteorological forcing on carbon and water fluxes in Nordic forests during the dry summer of 2018.

The Nordic region was subjected to severe drought in 2018 with a particularly long-lasting and large soil water deficit in Denmark, Southern Sweden and Estonia. Here, we analyse the impact of the drought on carbon and water fluxes in 11 forest ecosystems of different composition: spruce, pine, mixed and deciduous. We assess the impact of drought on fluxes by estimating the difference (anomaly) between year 2018 and a reference year without drought. Unexpectedly, the evaporation was only slightly reduced during 2018 compared to the reference year at two sites while it increased or was nearly unchanged at all other sites. This occurred under a 40 to 60% reduction in mean surface conductance and the concurrent increase in evaporative demand due to the warm and dry weather. The anomaly in the net ecosystem productivity (NEP) was 93% explained by a multilinear regression with the anomaly in heterotrophic respiration and the relative precipitation deficit as independent variables. Most of the variation (77%) was explained by the heterotrophic component. Six out of 11 forests reduced their annual NEP with more than 50 g C m-2 yr-1 during 2018 as compared to the reference year. The NEP anomaly ranged between -389 and +74 g C m-2 yr-1 with a median value of -59 g C m-2 yr-1. This article is part of the theme issue 'Impacts of the 2018 severe drought and heatwave in Europe: from site to continental scale'.

The mechanistic GEMMs of oncogenic histones.

The last decade's progress unraveling the mutational landscape of all age groups of cancer has uncovered mutations in histones as vital contributors of tumorigenesis. Here we review three new aspects of oncogenic histones: first, the identification of additional histone mutations potentially contributing to cancer formation; second, tumors expressing histone mutations to study the crosstalk of post-translational modifications, and; third, development of sophisticated biological model systems to reproduce tumorigenesis. At the outset, we recapitulate the firstly discovered histone mutations in pediatric and adolescent tumors of the brain and bone, which still remain the most pronounced histone alterations in cancer. We branch out to discuss the ramifications of histone mutations, including novel ones, that stem from altered protein-protein interactions of cognate histone modifiers as well as the stability of the nucleosome. We close by discussing animal models of oncogenic histones that reproduce tumor formation molecularly and morphologically and the prospect of utilizing them for drug testing, leading to efficient treatment and cure of deadly cancers with histone mutations.

EHMT2 Inhibition Induces Cell Death in Human Non-Small Cell Lung Cancer by Altering the Cholesterol Biosynthesis Pathway.

Non-small cell lung cancer (NSCLC) is a major subtype of lung cancer. Besides genetic and environmental factors, epigenetic alterations contribute to the tumorigenesis of NSCLC. Epigenetic changes are considered key drivers of cancer initiation and progression, and altered expression and activity of epigenetic modifiers reshape the epigenetic landscape in cancer cells. Euchromatic histone-lysine N-methyltransferase 2 (EHMT2) is a histone methyltransferase and catalyzes mono- and di-methylation at histone H3 lysine 9 (H3K9me1 and H3K9me2, respectively), leading to gene silencing. EHMT2 overexpression has been reported in various types of cancer, including ovarian cancer and neuroblastoma, in relation to cell proliferation and metastasis. However, its role in NSCLC is not fully understood. In this study, we showed that EHMT2 gene expression was higher in NSCLC than normal lung tissue based on publicly available data. Inhibition of EHMT2 by BIX01294 (BIX) reduced cell viability of NSCLC cell lines via induction of autophagy. Through RNA sequencing analysis, we found that EHMT2 inhibition significantly affected the cholesterol biosynthesis pathway. BIX treatment directly induced the expression of SREBF2, which is a master regulator of cholesterol biosynthesis, by lowering H3K9me1 and H3K9me2 at the promoter. Treatment of a cholesterol biosynthesis inhibitor, 25-hydroxycholesterol (25-HC), partially recovered BIX-induced cell death by attenuating autophagy. Our data demonstrated that EHMT2 inhibition effectively induced cell death in NSCLC cells through altering cholesterol metabolism-dependent autophagy.

The Net Landscape Carbon Balance-Integrating terrestrial and aquatic carbon fluxes in a managed boreal forest landscape in Sweden.

The boreal biome exchanges large amounts of carbon (C) and greenhouse gases (GHGs) with the atmosphere and thus significantly affects the global climate. A managed boreal landscape consists of various sinks and sources of carbon dioxide (CO2 ), methane (CH4 ), and dissolved organic and inorganic carbon (DOC and DIC) across forests, mires, lakes, and streams. Due to the spatial heterogeneity, large uncertainties exist regarding the net landscape carbon balance (NLCB). In this study, we compiled terrestrial and aquatic fluxes of CO2 , CH4 , DOC, DIC, and harvested C obtained from tall-tower eddy covariance measurements, stream monitoring, and remote sensing of biomass stocks for an entire boreal catchment (~68 km2 ) in Sweden to estimate the NLCB across the land-water-atmosphere continuum. Our results showed that this managed boreal forest landscape was a net C sink (NLCB = 39 g C m-2  year-1 ) with the landscape-atmosphere CO2 exchange being the dominant component, followed by the C export via harvest and streams. Accounting for the global warming potential of CH4 , the landscape was a GHG sink of 237 g CO2 -eq m-2  year-1 , thus providing a climate-cooling effect. The CH4 flux contribution to the annual GHG budget increased from 0.6% during spring to 3.2% during winter. The aquatic C loss was most significant during spring contributing 8% to the annual NLCB. We further found that abiotic controls (e.g., air temperature and incoming radiation) regulated the temporal variability of the NLCB whereas land cover types (e.g., mire vs. forest) and management practices (e.g., clear-cutting) determined their spatial variability. Our study advocates the need for integrating terrestrial and aquatic fluxes at the landscape scale based on tall-tower eddy covariance measurements combined with biomass stock and stream monitoring to develop a holistic understanding of the NLCB of managed boreal forest landscapes and to better evaluate their potential for mitigating climate change.

Transcriptome and protein interaction profiling in cancer cells with mutations in histone H3.3.

Mutations of histone variant H3.3 are highly recurrent in childhood glioblastoma and in young adults with Giant Cell Tumor of the Bone (GCTB). The heterozygotic representation of the mutations in the tumors, and with potential histone H3 and H3.3 redundancy, suggest that the mutations are gain-of-function by nature. To address common H3.3 point mutations, we have generated data from GCTB patient samples with H3.3 G34W substitutions and engineered human GFP-tagged H3.3-mutated isogenic cell lines for high throughput data comparisons. First, a total of thirty-six patient samples and cell lines were used to acquire gene expression transcriptome data using microarray and RNA-sequencing. The expression data were validated with the orthogonal nCounter assay. Second, to uncover the H3.3-GFP interaction proteomes from the isogenic cell lines, immunoprecipitation of unmutated wild type, K27M, G34R, and G34W substitutions were performed. The RNA-sequencing data and the H3.3 interaction proteome enable potentially important functional insight into the tumorigenic process and should spur further detailed analysis.

Somatic PRDM2 c.4467delA mutations in colorectal cancers control histone methylation and tumor growth.

The chromatin modifier PRDM2/RIZ1 is inactivated by mutation in several forms of cancer and is a putative tumor suppressor gene. Frameshift mutations in the C-terminal region of PRDM2, affecting (A)8 or (A)9 repeats within exon 8, are found in one third of colorectal cancers with microsatellite instability, but the contribution of these mutations to colorectal tumorigenesis is unknown. To model somatic mutations in microsatellite unstable tumors, we devised a general approach to perform genome editing while stabilizing the mutated nucleotide repeat. We then engineered isogenic cell systems where the PRDM2 c.4467delA mutation in human HCT116 colorectal cancer cells was corrected to wild-type by genome editing. Restored PRDM2 increased global histone 3 lysine 9 dimethylation and reduced migration, anchorage-independent growth and tumor growth in vivo. Gene set enrichment analysis revealed regulation of several hallmark cancer pathways, particularly of epithelial-to-mesenchymal transition (EMT), with VIM being the most significantly regulated gene. These observations provide direct evidence that PRDM2 c.4467delA is a driver mutation in colorectal cancer and confirms PRDM2 as a cancer gene, pointing to regulation of EMT as a central aspect of its tumor suppressive action.

The histone variant H3.3 G34W substitution in giant cell tumor of the bone link chromatin and RNA processing.

While transcription as regulated by histones and their post-translational modifications has been well described, the function of histone variants in this process remains poorly characterized. Potentially important insight into this process pertain to the frequently occurring mutations of H3.3, leading to G34 substitutions in childhood glioblastoma and giant cell tumor of the bone (GCTB). In this study, we have established primary cell lines from GCTB patients and used them to uncover the influence of H3.3 G34W substitutions on cellular growth behavior, gene expression, and chromatin compaction. Primary cell lines with H3.3 G34W showed increased colony formation, infiltration and proliferation, known hallmarks of tumor development. Isogenic cell lines with H3.3 G34W recapitulated the increased proliferation observed in primary cells. Transcriptomic analysis of primary cells and tumor biopsies revealed slightly more downregulated gene expression, perhaps by increased chromatin compaction. We identified components related to splicing, most prominently hnRNPs, by immunoprecipitation and mass spectrometry that specifically interact with H3.3 G34W in the isogenic cell lines. RNA-sequencing analysis and hybridization-based validations further enforced splicing aberrations. Our data uncover a role for H3.3 in RNA processing and chromatin modulation that is blocked by the G34W substitution, potentially driving the tumorigenic process in GCTB.

Early snowmelt significantly enhances boreal springtime carbon uptake.

We determine the annual timing of spring recovery from space-borne microwave radiometer observations across northern hemisphere boreal evergreen forests for 1979-2014. We find a trend of advanced spring recovery of carbon uptake for this period, with a total average shift of 8.1 d (2.3 d/decade). We use this trend to estimate the corresponding changes in gross primary production (GPP) by applying in situ carbon flux observations. Micrometeorological CO2 measurements at four sites in northern Europe and North America indicate that such an advance in spring recovery would have increased the January-June GPP sum by 29 g⋅C⋅m-2 [8.4 g⋅C⋅m-2 (3.7%)/decade]. We find this sensitivity of the measured springtime GPP to the spring recovery to be in accordance with the corresponding sensitivity derived from simulations with a land ecosystem model coupled to a global circulation model. The model-predicted increase in springtime cumulative GPP was 0.035 Pg/decade [15.5 g⋅C⋅m-2 (6.8%)/decade] for Eurasian forests and 0.017 Pg/decade for forests in North America [9.8 g⋅C⋅m-2 (4.4%)/decade]. This change in the springtime sum of GPP related to the timing of spring snowmelt is quantified here for boreal evergreen forests.

Erratum: DNMT and HDAC inhibitors induce cryptic transcription start sites encoded in long terminal repeats.

This corrects the article DOI: 10.1038/ng.3889.

Loss of DIP2C in RKO cells stimulates changes in DNA methylation and epithelial-mesenchymal transition.

BACKGROUND: The disco-interacting protein 2 homolog C (DIP2C) gene is an uncharacterized gene found mutated in a subset of breast and lung cancers. To understand the role of DIP2C in tumour development we studied the gene in human cancer cells. METHODS: We engineered human DIP2C knockout cells by genome editing in cancer cells. The growth properties of the engineered cells were characterised and transcriptome and methylation analyses were carried out to identify pathways deregulated by inactivation of DIP2C. Effects on cell death pathways and epithelial-mesenchymal transition traits were studied based on the results from expression profiling. RESULTS: Knockout of DIP2C in RKO cells resulted in cell enlargement and growth retardation. Expression profiling revealed 780 genes for which the expression level was affected by the loss of DIP2C, including the tumour-suppressor encoding CDKN2A gene, the epithelial-mesenchymal transition (EMT) regulator-encoding ZEB1, and CD44 and CD24 that encode breast cancer stem cell markers. Analysis of DNA methylation showed more than 30,000 sites affected by differential methylation, the majority of which were hypomethylated following loss of DIP2C. Changes in DNA methylation at promoter regions were strongly correlated to changes in gene expression, and genes involved with EMT and cell death were enriched among the differentially regulated genes. The DIP2C knockout cells had higher wound closing capacity and showed an increase in the proportion of cells positive for cellular senescence markers. CONCLUSIONS: Loss of DIP2C triggers substantial DNA methylation and gene expression changes, cellular senescence and epithelial-mesenchymal transition in cancer cells.