Long-term intermittent hypoxia in mice induces inflammatory pathways implicated in sleep apnea and steatohepatitis in humans.

Obstructive sleep apnea (OSA) induces intermittent hypoxia (IH), an independent risk factor for non-alcoholic fatty liver disease (NAFLD). While the molecular links between IH and NAFLD progression are unclear, immune cell-driven inflammation plays a crucial role in NAFLD pathogenesis. Using lean mice exposed to long-term IH and a cohort of lean OSA patients (n = 71), we conducted comprehensive hepatic transcriptomics, lipidomics, and targeted serum proteomics. Significantly, we demonstrated that long-term IH alone can induce NASH molecular signatures found in human steatohepatitis transcriptomic data. Biomarkers (PPARs, NRFs, arachidonic acid, IL16, IL20, IFNB, TNF-α) associated with early hepatic and systemic inflammation were identified. This molecular link between IH, sleep apnea, and steatohepatitis merits further exploration in clinical trials, advocating for integrating sleep apnea diagnosis in liver disease phenotyping. Our unique signatures offer potential diagnostic and treatment response markers, highlighting therapeutic targets in the comorbidity of NAFLD and OSA.

Astrocytic circadian clock control of energy expenditure by transcriptional stress responses in the ventromedial hypothalamus.

Hypothalamic circuits compute systemic information to control metabolism. Astrocytes residing within the hypothalamus directly sense nutrients and hormones, integrating metabolic information, and modulating neuronal responses. Nevertheless, the role of the astrocytic circadian clock on the control of energy balance remains unclear. We used mice with a targeted ablation of the core-clock gene Bmal1 within Gfap-expressing astrocytes to gain insight on the role played by this transcription factor in astrocytes. While this mutation does not substantially affect the phenotype in mice fed normo-caloric diet, under high-fat diet we unmasked a thermogenic phenotype consisting of increased energy expenditure, and catabolism in brown adipose and overall metabolic improvement consisting of better glycemia control, and body composition. Transcriptomic analysis in the ventromedial hypothalamus revealed an enhanced response to moderate cellular stress, including ER-stress response, unfolded protein response and autophagy. We identified Xbp1 and Atf1 as two key transcription factors enhancing cellular stress responses. Therefore, we unveiled a previously unknown role of the astrocytic circadian clock modulating energy balance through the regulation of cellular stress responses within the VMH.

Genotype-phenotype associations within the Li-Fraumeni spectrum: a report from the German Registry.

Li-Fraumeni syndrome (LFS) is a cancer predisposition syndrome caused by pathogenic TP53 variants. The condition represents one of the most relevant genetic causes of cancer in children and adults due to its frequency and high cancer risk. The term Li-Fraumeni spectrum reflects the evolving phenotypic variability of the condition. Within this spectrum, patients who meet specific LFS criteria are diagnosed with LFS, while patients who do not meet these criteria are diagnosed with attenuated LFS. To explore genotype-phenotype correlations we analyzed 141 individuals from 94 families with pathogenic TP53 variants registered in the German Cancer Predisposition Syndrome Registry. Twenty-one (22%) families had attenuated LFS and 73 (78%) families met the criteria of LFS. NULL variants occurred in 32 (44%) families with LFS and in two (9.5%) families with attenuated LFS (P value < 0.01). Kato partially functional variants were present in 10 out of 53 (19%) families without childhood cancer except adrenocortical carcinoma (ACC) versus 0 out of 41 families with childhood cancer other than ACC alone (P value < 0.01). Our study suggests genotype-phenotype correlations encouraging further analyses.

Intermittent Hypoxia Rewires the Liver Transcriptome and Fires up Fatty Acids Usage for Mitochondrial Respiration.

Sleep Apnea Syndrome (SAS) is one of the most common chronic diseases, affecting nearly one billion people worldwide. The repetitive occurrence of abnormal respiratory events generates cyclical desaturation-reoxygenation sequences known as intermittent hypoxia (IH). Among SAS metabolic sequelae, it has been established by experimental and clinical studies that SAS is an independent risk factor for the development and progression of non-alcoholic fatty liver disease (NAFLD). The principal goal of this study was to decrypt the molecular mechanisms at the onset of IH-mediated liver injury. To address this question, we used a unique mouse model of SAS exposed to IH, employed unbiased high-throughput transcriptomics and computed network analysis. This led us to examine hepatic mitochondrial ultrastructure and function using electron microscopy, high-resolution respirometry and flux analysis in isolated mitochondria. Transcriptomics and network analysis revealed that IH reprograms Nuclear Respiratory Factor- (NRF-) dependent gene expression and showed that mitochondria play a central role. We thus demonstrated that IH boosts the oxidative capacity from fatty acids of liver mitochondria. Lastly, the unbalance between oxidative stress and antioxidant defense is tied to an increase in hepatic ROS production and DNA damage during IH. We provide a comprehensive analysis of liver metabolism during IH and reveal the key role of the mitochondria at the origin of development of liver disease. These findings contribute to the understanding of the mechanisms underlying NAFLD development and progression during SAS and provide a rationale for novel therapeutic targets and biomarker discovery.

A non-pharmacological therapeutic approach in the gut triggers distal metabolic rewiring capable of ameliorating diet-induced dysfunctions encompassed by metabolic syndrome.

Metabolic syndrome has increased at a worrisome level. Lifestyle changes are not sufficient to prevent and improve the adverse effects of obesity, thus novel interventions are necessary. The aim of this study was to investigate the use and metabolic outcomes of a non-pharmacological intervention in a high-fat diet (HFD) fed mouse model, capable of recapitulating key aspects of metabolic syndrome. We show that Policaptil Gel Retard has remarkable, beneficial effects on metabolic dysfunction caused by consumption of HFD. We describe the mechanism by which such effects are obtained, highlighting the fact that the amelioration of metabolic function observed upon Policaptil Gel Retard administration is profound and of systemic nature, despite being originated by sequestering, therefore non-pharmacological events elicited in the gut lumen.

Multi-platform NMR Study of Pluripotent Stem Cells Unveils Complementary Metabolic Signatures towards Differentiation.

Stem cells, poised to revolutionize current medicine, stand as major workhorses for monitoring changes in cell fate. Characterizing metabolic phenotypes is key to monitor in differentiating cells transcriptional and epigenetic shifts at a functional level and provides a non-genetic means to control cell specification. Expanding the arsenal of analytical tools for metabolic profiling of cell differentiation is therefore of importance. Here, we describe the metabolome of whole pluripotent stem cells (PSCs) using high-resolution magic angle spinning (HR-MAS), a non-destructive approach for Nuclear Magnetic Resonance (NMR) analysis. The integrated 1H NMR analysis results in detection of metabolites of various groups, including energy metabolites, amino acids, choline derivatives and short chain fatty acids. It unveils new metabolites that discriminate PSCs from differentiated counterparts and directly measures substrates and co-factors of histone modifying enzymes, suggesting that NMR stands as a strategic technique for deciphering metabolic regulations of histone post-translational modifications. HR-MAS NMR analysis of whole PSCs complements the much used solution NMR of cell extracts. Altogether, our multi-platform NMR investigation provides a consolidated picture of PSC metabolic signatures and of metabolic pathways involved in differentiation.

Distinct metabolic adaptation of liver circadian pathways to acute and chronic patterns of alcohol intake.

Binge drinking and chronic exposure to ethanol contribute to alcoholic liver diseases (ALDs). A potential link between ALDs and circadian disruption has been observed, though how different patterns of alcohol consumption differentially impact hepatic circadian metabolism remains virtually unexplored. Using acute versus chronic ethanol feeding, we reveal differential reprogramming of the circadian transcriptome in the liver. Specifically, rewiring of diurnal SREBP transcriptional pathway leads to distinct hepatic signatures in acetyl-CoA metabolism that are translated into the subcellular patterns of protein acetylation. Thus, distinct drinking patterns of alcohol dictate differential adaptation of hepatic circadian metabolism.

Targeting the interplay between metabolism and epigenetics in cancer.

PURPOSE OF REVIEW: Metabolic perturbation is a hallmark of cancer favoring tumor progression. It is now demonstrated that cell metabolism has an impact on gene expression through epigenetic modifications. In this review, we expose recent evidences of metabolic-driven epigenetic perturbations in cancer and subsequent therapeutic opportunities. RECENT FINDINGS: The intimate link between metabolism and epigenetics and its rewiring in carcinogenesis is a hot topic. Chromatin-modifying enzymes involved in the dynamics of methylation or acetylation require small metabolites as cofactors or substrates, thus orchestrating the integration between epigenetic and transcriptional states. Mutations in metabolic enzymes such as isocitrate dehydrogenase 1 and 2 cause the accumulation of metabolites that upset the balance of histone and DNA methylation, thus generating widespread deregulation of epigenetically controlled gene expression. Additionally, modifications of catalytic activity and subcellular localization of metabolic enzymes in cancer can impact on epigenetic modifications and gene expression programs to favor tumor progression. SUMMARY: The interplay between metabolism and epigenetics and its molecular characterization in cancer cells identifies potential targets for the development of new therapies.

Epigenetic regulation of the circadian gene Per1 contributes to age-related changes in hippocampal memory.

Aging is accompanied by impairments in both circadian rhythmicity and long-term memory. Although it is clear that memory performance is affected by circadian cycling, it is unknown whether age-related disruption of the circadian clock causes impaired hippocampal memory. Here, we show that the repressive histone deacetylase HDAC3 restricts long-term memory, synaptic plasticity, and experience-induced expression of the circadian gene Per1 in the aging hippocampus without affecting rhythmic circadian activity patterns. We also demonstrate that hippocampal Per1 is critical for long-term memory formation. Together, our data challenge the traditional idea that alterations in the core circadian clock drive circadian-related changes in memory formation and instead argue for a more autonomous role for circadian clock gene function in hippocampal cells to gate the likelihood of long-term memory formation.

Molecular Cogs: Interplay between Circadian Clock and Cell Cycle.

The cell cycle and the circadian clock operate as biological oscillators whose timed functions are tightly regulated. Accumulating evidence illustrates the presence of molecular links between these two oscillators. This mutual interplay utilizes various coupling mechanisms, such as the use of common regulators. The connection between these two cyclic systems has unique interest in the context of aberrant cell proliferation since both of these oscillators are frequently misregulated in cancer cells. Further studies will provide deeper understanding of the detailed molecular connections between the cell cycle and the circadian clock and may also serve as a basis for the design of innovative therapeutic strategies.

A Circadian Genomic Signature Common to Ketamine and Sleep Deprivation in the Anterior Cingulate Cortex.

BACKGROUND: Conventional antidepressants usually require several weeks to achieve a full clinical response in patients with major depressive disorder, an illness associated with dysregulated circadian rhythms and a high incidence of suicidality. Two rapid-acting antidepressant strategies, low-dose ketamine (KT) and sleep deprivation (SD) therapies, dramatically reduce depressive symptoms within 24 hours in a subset of major depressive disorder patients. However, it is unknown whether they exert their actions through shared regulatory mechanisms. To address this question, we performed comparative transcriptomics analyses to identify candidate genes and relevant pathways common to KT and SD. METHODS: We used the forced swim test, a standardized behavioral approach to measure antidepressant-like activity of KT and SD. We investigated gene expression changes using high-density microarrays and pathway analyses (Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Gene Set Enrichment Analysis) in KT- and SD-treated mice compared with saline-treated control male mice. RESULTS: We show that KT and SD elicit common transcriptional responses implicating distinct elements of the circadian clock and processes involved in neuronal plasticity. There is an overlap of 64 genes whose expression is common in KT and SD. Specifically, there is downregulation of clock genes including Ciart, Per2, Npas4, Dbp, and Rorb in both KT- and SD-treated mice. CONCLUSIONS: We demonstrate a potential involvement of the circadian clock in rapid antidepressant responses. These findings could open new research avenues to help design chronopharmacological strategies to treat major depressive disorder.

Histone Variant H2A.L.2 Guides Transition Protein-Dependent Protamine Assembly in Male Germ Cells.

Histone replacement by transition proteins (TPs) and protamines (Prms) constitutes an essential step for the successful production of functional male gametes, yet nothing is known on the underlying functional interplay between histones, TPs, and Prms. Here, by studying spermatogenesis in the absence of a spermatid-specific histone variant, H2A.L.2, we discover a fundamental mechanism involved in the transformation of nucleosomes into nucleoprotamines. H2A.L.2 is synthesized at the same time as TPs and enables their loading onto the nucleosomes. TPs do not displace histones but rather drive the recruitment and processing of Prms, which are themselves responsible for histone eviction. Altogether, the incorporation of H2A.L.2 initiates and orchestrates a series of successive transitional states that ultimately shift to the fully compacted genome of the mature spermatozoa. Hence, the current view of histone-to-nucleoprotamine transition should be revisited and include an additional step with H2A.L.2 assembly prior to the action of TPs and Prms.

Dynamic Competing Histone H4 K5K8 Acetylation and Butyrylation Are Hallmarks of Highly Active Gene Promoters.

Recently discovered histone lysine acylation marks increase the functional diversity of nucleosomes well beyond acetylation. Here, we focus on histone butyrylation in the context of sperm cell differentiation. Specifically, we investigate the butyrylation of histone H4 lysine 5 and 8 at gene promoters where acetylation guides the binding of Brdt, a bromodomain-containing protein, thereby mediating stage-specific gene expression programs and post-meiotic chromatin reorganization. Genome-wide mapping data show that highly active Brdt-bound gene promoters systematically harbor competing histone acetylation and butyrylation marks at H4 K5 and H4 K8. Despite acting as a direct stimulator of transcription, histone butyrylation competes with acetylation, especially at H4 K5, to prevent Brdt binding. Additionally, H4 K5K8 butyrylation also marks retarded histone removal during late spermatogenesis. Hence, alternating H4 acetylation and butyrylation, while sustaining direct gene activation and dynamic bromodomain binding, could impact the final male epigenome features.

Lysine 2-hydroxyisobutyrylation is a widely distributed active histone mark.

We report the identification of a new type of histone mark, lysine 2-hydroxyisobutyrylation (Khib), and identify the mark at 63 human and mouse histone Khib sites, including 27 unique lysine sites that are not known to be modified by lysine acetylation (Kac) and lysine crotonylation (Kcr). This histone mark was initially identified by MS and then validated by chemical and biochemical methods. Histone Khib shows distinct genomic distributions from histone Kac or histone Kcr during male germ cell differentiation. Using chromatin immunoprecipitation sequencing, gene expression analysis and immunodetection, we show that in male germ cells, H4K8hib is associated with active gene transcription in meiotic and post-meiotic cells. In addition, H4K8ac-associated genes are included in and constitute only a subfraction of H4K8hib-labeled genes. The histone Khib mark is conserved and widely distributed, has high stoichiometry and induces a large structural change. These findings suggest its critical role on the regulation of chromatin functions.

Acetylation of histone H3 at lysine 64 regulates nucleosome dynamics and facilitates transcription.

Post-translational modifications of proteins have emerged as a major mechanism for regulating gene expression. However, our understanding of how histone modifications directly affect chromatin function remains limited. In this study, we investigate acetylation of histone H3 at lysine 64 (H3K64ac), a previously uncharacterized acetylation on the lateral surface of the histone octamer. We show that H3K64ac regulates nucleosome stability and facilitates nucleosome eviction and hence gene expression in vivo. In line with this, we demonstrate that H3K64ac is enriched in vivo at the transcriptional start sites of active genes and it defines transcriptionally active chromatin. Moreover, we find that the p300 co-activator acetylates H3K64, and consistent with a transcriptional activation function, H3K64ac opposes its repressive counterpart H3K64me3. Our findings reveal an important role for a histone modification within the nucleosome core as a regulator of chromatin function and they demonstrate that lateral surface modifications can define functionally opposing chromatin states. DOI: http://dx.doi.org/10.7554/eLife.01632.001.

Chromatin-to-nucleoprotamine transition is controlled by the histone H2B variant TH2B.

The conversion of male germ cell chromatin to a nucleoprotamine structure is fundamental to the life cycle, yet the underlying molecular details remain obscure. Here we show that an essential step is the genome-wide incorporation of TH2B, a histone H2B variant of hitherto unknown function. Using mouse models in which TH2B is depleted or C-terminally modified, we show that TH2B directs the final transformation of dissociating nucleosomes into protamine-packed structures. Depletion of TH2B induces compensatory mechanisms that permit histone removal by up-regulating H2B and programming nucleosome instability through targeted histone modifications, including lysine crotonylation and arginine methylation. Furthermore, after fertilization, TH2B reassembles onto the male genome during protamine-to-histone exchange. Thus, TH2B is a unique histone variant that plays a key role in the histone-to-protamine packing of the male genome and guides genome-wide chromatin transitions that both precede and follow transmission of the male genome to the egg.

Bromodomain-dependent stage-specific male genome programming by Brdt.

Male germ cell differentiation is a highly regulated multistep process initiated by the commitment of progenitor cells into meiosis and characterized by major chromatin reorganizations in haploid spermatids. We report here that a single member of the double bromodomain BET factors, Brdt, is a master regulator of both meiotic divisions and post-meiotic genome repackaging. Upon its activation at the onset of meiosis, Brdt drives and determines the developmental timing of a testis-specific gene expression program. In meiotic and post-meiotic cells, Brdt initiates a genuine histone acetylation-guided programming of the genome by activating essential genes and repressing a 'progenitor cells' gene expression program. At post-meiotic stages, a global chromatin hyperacetylation gives the signal for Brdt's first bromodomain to direct the genome-wide replacement of histones by transition proteins. Brdt is therefore a unique and essential regulator of male germ cell differentiation, which, by using various domains in a developmentally controlled manner, first drives a specific spermatogenic gene expression program, and later controls the tight packaging of the male genome.

[Cross-fire over the nucleosome: molecular basis of post-meiotic male haploid genome compaction]. / Feux croisés sur le nucléosome: bases moléculaires de la compaction du génome mâle haploïde.

In mammals, the post-meiotic phases of spermatogenesis direct a drastic compaction of the genome characterized by the replacement of the majority of histones by small basic non-histone proteins, transition proteins followed by protamines. Although the mechanisms controlling this process are poorly understood, available data suggest that the nucleosome, which represents the universal unit of genome organization in eukaryotes, is the target of specific events cooperating to globally transform DNA organization at a genomic scale. Indeed, after meiosis, nearly all nucleosomes associated with the male genome undergo successive modifications increasing their instability, finally leading to their dissociation and re-organization. The characterization of the underlying mechanisms is a real challenge and appears as essential for the understanding of male gametogenesis and associated pathologies.

Histone crotonylation specifically marks the haploid male germ cell gene expression program: post-meiotic male-specific gene expression.

The haploid male germ cell differentiation program controls essential steps of male gametogenesis and relies partly on a significant number of sex chromosome-linked genes. These genes need to escape chromosome-wide transcriptional repression of sex chromosomes, which occurs during meiosis and is largely maintained in post-meiotic cells. A newly discovered histone lysine modification, crotonylation (Kcr), marks X/Y-linked genes that are active in post-meiotic male germ cells. Histone Kcr, by conferring resistance to transcriptional repressors, could be a dominant element in maintaining these genes active in the globally repressive environment of haploid cell sex chromosomes. Furthermore, the same mark was found associated with post-meiotically activated genes on autosomes. Histone Kcr therefore appears to be an indicator of the male haploid cell gene expression program and a notable element of genome programming in the post-meiotic phases of spermatogenesis.

Identification of 67 histone marks and histone lysine crotonylation as a new type of histone modification.

We report the identification of 67 previously undescribed histone modifications, increasing the current number of known histone marks by about 70%. We further investigated one of the marks, lysine crotonylation (Kcr), confirming that it represents an evolutionarily-conserved histone posttranslational modification. The unique structure and genomic localization of histone Kcr suggest that it is mechanistically and functionally different from histone lysine acetylation (Kac). Specifically, in both human somatic and mouse male germ cell genomes, histone Kcr marks either active promoters or potential enhancers. In male germinal cells immediately following meiosis, Kcr is enriched on sex chromosomes and specifically marks testis-specific genes, including a significant proportion of X-linked genes that escape sex chromosome inactivation in haploid cells. These results therefore dramatically extend the repertoire of histone PTM sites and designate Kcr as a specific mark of active sex chromosome-linked genes in postmeiotic male germ cells.