Recognition of Histone Crotonylation by Taf14 Links Metabolic State to Gene Expression.

Metabolic signaling to chromatin often underlies how adaptive transcriptional responses are controlled. While intermediary metabolites serve as co-factors for histone-modifying enzymes during metabolic flux, how these modifications contribute to transcriptional responses is poorly understood. Here, we utilize the highly synchronized yeast metabolic cycle (YMC) and find that fatty acid ß-oxidation genes are periodically expressed coincident with the ß-oxidation byproduct histone crotonylation. Specifically, we found that H3K9 crotonylation peaks when H3K9 acetylation declines and energy resources become limited. During this metabolic state, pro-growth gene expression is dampened; however, mutation of the Taf14 YEATS domain, a H3K9 crotonylation reader, results in de-repression of these genes. Conversely, exogenous addition of crotonic acid results in increased histone crotonylation, constitutive repression of pro-growth genes, and disrupted YMC oscillations. Together, our findings expose an unexpected link between metabolic flux and transcription and demonstrate that histone crotonylation and Taf14 participate in the repression of energy-demanding gene expression.

Endocardial/endothelial angiocrines regulate cardiomyocyte development and maturation and induce features of ventricular non-compaction.

AIMS: Non-compaction cardiomyopathy is a devastating genetic disease caused by insufficient consolidation of ventricular wall muscle that can result in inadequate cardiac performance. Despite being the third most common cardiomyopathy, the mechanisms underlying the disease, including the cell types involved, are poorly understood. We have previously shown that endothelial cell-specific deletion of the chromatin remodeller gene Ino80 results in defective coronary vessel development that leads to ventricular non-compaction in embryonic mouse hearts. We aimed to identify candidate angiocrines expressed by endocardial and endothelial cells (ECs) in wildtype and LVNC conditions in Tie2Cre;Ino80fl/fltransgenic embryonic mouse hearts, and test the effect of these candidates on cardiomyocyte proliferation and maturation. METHODS AND RESULTS: We used single-cell RNA-sequencing to characterize endothelial and endocardial defects in Ino80-deficient hearts. We observed a pathological endocardial cell population in the non-compacted hearts and identified multiple dysregulated angiocrine factors that dramatically affected cardiomyocyte behaviour. We identified Col15a1 as a coronary vessel-secreted angiocrine factor, downregulated by Ino80-deficiency, that functioned to promote cardiomyocyte proliferation. Furthermore, mutant endocardial and endothelial cells up-regulated expression of secreted factors, such as Tgfbi, Igfbp3, Isg15, and Adm, which decreased cardiomyocyte proliferation and increased maturation. CONCLUSIONS: These findings support a model where coronary endothelial cells normally promote myocardial compaction through secreted factors, but that endocardial and endothelial cells can secrete factors that contribute to non-compaction under pathological conditions.

Carcinogen susceptibility is regulated by genome architecture and predicts cancer mutagenesis.

The development of many sporadic cancers is directly initiated by carcinogen exposure. Carcinogens induce malignancies by creating DNA lesions (i.e., adducts) that can result in mutations if left unrepaired. Despite this knowledge, there has been remarkably little investigation into the regulation of susceptibility to acquire DNA lesions. In this study, we present the first quantitative human genome-wide map of DNA lesions induced by ultraviolet (UV) radiation, the ubiquitous carcinogen in sunlight that causes skin cancer. Remarkably, the pattern of carcinogen susceptibility across the genome of primary cells significantly reflects mutation frequency in malignant melanoma. Surprisingly, DNase-accessible euchromatin is protected from UV, while lamina-associated heterochromatin at the nuclear periphery is vulnerable. Many cancer driver genes have an intrinsic increase in carcinogen susceptibility, including the BRAF oncogene that has the highest mutation frequency in melanoma. These findings provide a genome-wide snapshot of DNA injuries at the earliest stage of carcinogenesis. Furthermore, they identify carcinogen susceptibility as an origin of genome instability that is regulated by nuclear architecture and mirrors mutagenesis in cancer.

The INO80 chromatin remodeler sustains metabolic stability by promoting TOR signaling and regulating histone acetylation.

Chromatin remodeling complexes are essential for gene expression programs that coordinate cell function with metabolic status. However, how these remodelers are integrated in metabolic stability pathways is not well known. Here, we report an expansive genetic screen with chromatin remodelers and metabolic regulators in Saccharomyces cerevisiae. We found that, unlike the SWR1 remodeler, the INO80 chromatin remodeling complex is composed of multiple distinct functional subunit modules. We identified a strikingly divergent genetic signature for the Ies6 subunit module that links the INO80 complex to metabolic homeostasis. In particular, mitochondrial maintenance is disrupted in ies6 mutants. INO80 is also needed to communicate TORC1-mediated signaling to chromatin, as ino80 mutants exhibit defective transcriptional profiles and altered histone acetylation of TORC1-responsive genes. Furthermore, comparative analysis reveals subunits of INO80 and mTORC1 have high co-occurrence of alterations in human cancers. Collectively, these results demonstrate that the INO80 complex is a central component of metabolic homeostasis that influences histone acetylation and may contribute to disease when disrupted.

A multimodal approach in total hip arthroplasty preoperative templating.

OBJECTIVES: Accurate preoperative planning is mandatory for a successful total hip arthroplasty (THA). Templating helps to achieve multiple technical goals, including offset restoration, leg-length equality, and components positioning. This study aims to measure how successful a new stepwise method for preoperative radiographic templating is in achieving postoperative restoration of anatomy. METHODS: We measured the relationship of several radiographic landmarks on the affected limb in comparison to the unaffected limb to define the appropriate position of the components. One hundred consecutive patients with unilateral disease undergoing THA were retrospectively analyzed. The preoperative templates were compared with the postoperative radiographs. RESULTS: Accuracy in achieving the desired offset, leg-length discrepancy within 5 mm, was 93 % and 100 % respectively. Acetabular component positioning within 2° was replicated in 92 %. CONCLUSION: We conclude that this technique can help the surgeon to accurately reproduce the anatomy of the contralateral unaffected hip.

Retinoblastoma protein regulates carcinogen susceptibility at heterochromatic cancer driver loci.

Carcinogenic insult, such as UV light exposure, creates DNA lesions that evolve into mutations if left unrepaired. These resulting mutations can contribute to carcinogenesis and drive malignant phenotypes. Susceptibility to carcinogens (i.e., the propensity to form a carcinogen-induced DNA lesion) is regulated by both genetic and epigenetic factors. Importantly, carcinogen susceptibility is a critical contributor to cancer mutagenesis. It is known that mutations can be prevented by tumor suppressor regulation of DNA damage response pathways; however, their roles carcinogen susceptibility have not yet been reported. In this study, we reveal that the retinoblastoma (RB1) tumor suppressor regulates UV susceptibility across broad regions of the genome. In particular, centromere and telomere-proximal regions exhibit significant increases in UV lesion susceptibility when RB1 is deleted. Several cancer-related genes are located within genomic regions of increased susceptibility, including telomerase reverse transcriptase, TERT, thereby accelerating mutagenic potential in cancers with RB1 pathway alterations. These findings reveal novel genome stability mechanisms of a tumor suppressor and uncover new pathways to accumulate mutations during cancer evolution.

Genome-wide profiles of UV lesion susceptibility, repair, and mutagenic potential in melanoma.

Exposure to the ultraviolet (UV) radiation in sunlight creates DNA lesions, which if left unrepaired can induce mutations and contribute to skin cancer. The two most common UV-induced DNA lesions are the cis-syn cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts (6-4PPs), both of which can initiate mutations. Interestingly, mutation frequency across the genomes of many cancers is heterogenous with significant increases in heterochromatin. Corresponding increases in UV lesion susceptibility and decreases in repair are observed in heterochromatin versus euchromatin. However, the individual contributions of CPDs and 6-4PPs to mutagenesis have not been systematically examined in specific genomic and epigenomic contexts. In this study, we compared genome-wide maps of 6-4PP and CPD lesion abundances in primary cells and conducted comprehensive analyses to determine the genetic and epigenetic features associated with susceptibility. Overall, we found a high degree of similarity between 6-4PP and CPD formation, with an enrichment of both in heterochromatin regions. However, when examining the relative levels of the two UV lesions, we found that bivalent and Polycomb-repressed chromatin states were uniquely more susceptible to 6-4PPs. Interestingly, when comparing UV susceptibility and repair with melanoma mutation frequency in these regions, disparate patterns were observed in that susceptibility was not always inversely associated with repair and mutation frequency. Functional enrichment analysis hint at mechanisms of negative selection for these regions that are essential for cell viability, immune function and induce cell death when mutated. Ultimately, these results reveal both the similarities and differences between UV-induced lesions that contribute to melanoma.

Somatic mutation distributions in cancer genomes vary with three-dimensional chromatin structure.

Somatic mutations in driver genes may ultimately lead to the development of cancer. Understanding how somatic mutations accumulate in cancer genomes and the underlying factors that generate somatic mutations is therefore crucial for developing novel therapeutic strategies. To understand the interplay between spatial genome organization and specific mutational processes, we studied 3,000 tumor-normal-pair whole-genome datasets from 42 different human cancer types. Our analyses reveal that the change in somatic mutational load in cancer genomes is co-localized with topologically-associating-domain boundaries. Domain boundaries constitute a better proxy to track mutational load change than replication timing measurements. We show that different mutational processes lead to distinct somatic mutation distributions where certain processes generate mutations in active domains, and others generate mutations in inactive domains. Overall, the interplay between three-dimensional genome organization and active mutational processes has a substantial influence on the large-scale mutation-rate variations observed in human cancers.

INO80 Chromatin Remodeling Coordinates Metabolic Homeostasis with Cell Division.

Adaptive survival requires the coordination of nutrient availability with expenditure of cellular resources. For example, in nutrient-limited environments, 50% of all S. cerevisiae genes synchronize and exhibit periodic bursts of expression in coordination with respiration and cell division in the yeast metabolic cycle (YMC). Despite the importance of metabolic and proliferative synchrony, the majority of YMC regulators are currently unknown. Here, we demonstrate that the INO80 chromatin-remodeling complex is required to coordinate respiration and cell division with periodic gene expression. Specifically, INO80 mutants have severe defects in oxygen consumption and promiscuous cell division that is no longer coupled with metabolic status. In mutant cells, chromatin accessibility of periodic genes, including TORC1-responsive genes, is relatively static, concomitant with severely attenuated gene expression. Collectively, these results reveal that the INO80 complex mediates metabolic signaling to chromatin to restrict proliferation to metabolically optimal states.

Endothelial deletion of Ino80 disrupts coronary angiogenesis and causes congenital heart disease.

During development, the formation of a mature, well-functioning heart requires transformation of the ventricular wall from a loose trabecular network into a dense compact myocardium at mid-gestation. Failure to compact is associated in humans with congenital diseases such as left ventricular non-compaction (LVNC). The mechanisms regulating myocardial compaction are however still poorly understood. Here, we show that deletion of the Ino80 chromatin remodeler in vascular endothelial cells prevents ventricular compaction in the developing mouse heart. This correlates with defective coronary vascularization, and specific deletion of Ino80 in the two major coronary progenitor tissues-sinus venosus and endocardium-causes intermediate phenotypes. In vitro, endothelial cells promote myocardial expansion independently of blood flow in an Ino80-dependent manner. Ino80 deletion increases the expression of E2F-activated genes and endothelial cell S-phase occupancy. Thus, Ino80 is essential for coronary angiogenesis and allows coronary vessels to support proper compaction of the heart wall.

The INO80 Complex Requires the Arp5-Ies6 Subcomplex for Chromatin Remodeling and Metabolic Regulation.

ATP-dependent chromatin remodeling complexes are essential for transcription regulation, and yet it is unclear how these multisubunit complexes coordinate their activities to facilitate diverse transcriptional responses. In this study, we found that the conserved Arp5 and Ies6 subunits of the Saccharomyces cerevisiae INO80 chromatin-remodeler form an abundant and distinct subcomplex in vivo and stimulate INO80-mediated activity in vitro. Moreover, our genomic studies reveal that the relative occupancy of Arp5-Ies6 correlates with nucleosome positioning at transcriptional start sites and expression levels of >1,000 INO80-regulated genes. Notably, these genes are significantly enriched in energy metabolism pathways. Specifically, arp5Δ, ies6Δ, and ino80Δ mutants demonstrate decreased expression of genes involved in glycolysis and increased expression of genes in the oxidative phosphorylation pathway. Deregulation of these metabolic pathways results in constitutively elevated mitochondrial potential and oxygen consumption. Our results illustrate the dynamic nature of the INO80 complex assembly and demonstrate for the first time that a chromatin remodeler regulates glycolytic and respiratory capacity, thereby maintaining metabolic stability.

Transcriptome profiling of Set5 and Set1 methyltransferases: Tools for visualization of gene expression.

Cells regulate transcription by coordinating the activities of multiple histone modifying complexes. We recently identified the yeast histone H4 methyltransferase Set5 and discovered functional overlap with the histone H3 methyltransferase Set1 in gene expression. Specifically, using next-generation RNA sequencing (RNA-Seq), we found that Set5 and Set1 function synergistically to regulate specific transcriptional programs at subtelomeres and transposable elements [1]. Here we provide a comprehensive description of the methodology and analysis tools corresponding to the data deposited in NCBI's Gene Expression Omnibus (GEO) under the accession number GSE52086. This data complements the experimental methods described in Mas Martín G et al., 2014, and provides the means to explore the cooperative functions of histone H3 and H4 methyltransferases in the regulation of transcription. Furthermore, a fully annotated R code is included to enable researchers to use the following computational tools: comparison of significant differential expression (SDE) profiles; gene ontology enrichment of SDE; and enrichment of SDE relative to chromosomal features, such as centromeres, telomeres, and transposable elements. Overall, we present a bioinformatics platform that can be generally implemented for similar analyses with different datasets and in different organisms.

Set5 and Set1 cooperate to repress gene expression at telomeres and retrotransposons.

A complex interplay between multiple chromatin modifiers is critical for cells to regulate chromatin structure and accessibility during essential DNA-templated processes such as transcription. However, the coordinated activities of these chromatin modifiers in the regulation of gene expression are not fully understood. We previously determined that the budding yeast histone H4 methyltransferase Set5 functions together with Set1, the H3K4 methyltransferase, in specific cellular contexts. Here, we sought to understand the relationship between these evolutionarily conserved enzymes in the regulation of gene expression. We generated a comprehensive genetic interaction map of the functionally uncharacterized Set5 methyltransferase and expanded the existing genetic interactome of the global chromatin modifier Set1, revealing functional overlap of the two enzymes in chromatin-related networks, such as transcription. Furthermore, gene expression profiling via RNA-Seq revealed an unexpected synergistic role of Set1 and Set5 in repressing transcription of Ty transposable elements and genes located in subtelomeric regions. This study uncovers novel pathways in which the methyltransferase Set5 participates and, more importantly, reveals a partnership between Set1 and Set5 in transcriptional repression near repetitive DNA elements in budding yeast. Together, our results define a new functional relationship between histone H3 and H4 methyltransferases, whose combined activity may be implicated in preserving genomic integrity.