The tumor immune microenvironment is a main contributor to cancer progression and a promising therapeutic target for oncology. However, immune microenvironments vary profoundly between patients, and biomarkers for prognosis and treatment response lack precision. A comprehensive compendium of tumor immune cells is required to pinpoint predictive cellular states and their spatial localization. We generated

Direct comparison of bulk gene expression profiles is complicated by distinct cell type mixtures in each sample that obscure whether observed differences are actually caused by changes in the expression levels themselves or are simply a result of differing cell type compositions. Single-cell technology has made it possible to measure gene expression in individual cells, achieving higher resolution

The metabolic heterogeneity and metabolic interplay between cells are known as significant contributors to disease treatment resistance. However, with the lack of a mature high-throughput single-cell metabolomics technology, we are yet to establish systematic understanding of the intra-tissue metabolic heterogeneity and cooperative mechanisms. To mitigate this knowledge gap, we developed a novel computational

Organ function relies on the spatial organization and functional coordination of numerous cell types. The Drosophila ovary is a widely used model system to study the cellular activities underlying organ function, including stem cell regulation, cell signaling and epithelial morphogenesis. However, the relative paucity of cell type–specific reagents hinders investigation of molecular functions at the

Structural variants (SVs) are an important source of human genome diversity but their functional effects are not well understood. We mapped 61,668 SVs in 613 individuals with deep genome sequencing data from the GTEx project and measured their effects on gene expression. We estimate that common SVs are causal at 2.66% of eQTLs, which is a 10.5-fold enrichment relative to their abundance in the genome

More than 80% of the wheat genome consists of transposable elements (TEs), which act as one major driver of wheat genome evolution. However, their contributions to the regulatory evolution of wheat adaptations remain largely unclear. Here, we created genome-binding maps for 53 transcription factors (TFs) underlying environmental responses by leveraging DAP-seq in Triticum urartu, together with epigenomic

The number of publicly available microbiome samples is continually growing. As dataset size increases, bottlenecks arise in standard analytical pipelines. Faith’s phylogenetic diversity is a highly utilized phylogenetic alpha diversity metric that has thus far failed to effectively scale to trees with millions of vertices. Stacked Faith's Phylogenetic Diversity (SFPhD) enables calculation of this widely

Intronic polyadenylation (IpA) usually leads to changes in coding region of an mRNA, and its implication in diseases has been recognized, though at its very beginning status. Conveniently and accurately identifying IpA is of great importance for further evaluating its biological significance. Here, we developed IPAFinder, a bioinformatic method for the de novo identification of intronic poly(A) sites

The effects of DNASE1L3 or DNASE1 deficiency on cell-free DNA (cfDNA) methylation was explored in plasma of mice deficient in these nucleases and in DNASE1L3-deficient humans. Compared to wild-type cfDNA, cfDNA in Dnase1l3-deficient mice was significantly hypomethylated, while cfDNA in Dnase1-deficient mice was hypermethylated. The cfDNA hypomethylation in Dnase1l3-deficient mice was due to increased

We used capped analysis of gene expression with sequencing (CAGE-seq) to profile eRNA expression and enhancer activity during embryogenesis of a model echinoderm: the sea urchin, Strongylocentrotus purpuratus. We identified more than 18,000 enhancers that were active in mature oocytes and developing embryos and documented a burst of enhancer activation during cleavage and early blastula stages. We

Antibodies offer a powerful means to interrogate specific proteins in a complex milieu. However, antibody availability and reliability can be problematic, whereas epitope tagging can be impractical in many cases. To address these limitations, the Protein Capture Reagents Program (PCRP) generated over a thousand renewable monoclonal antibodies (mAbs) against human presumptive chromatin proteins. However

High-throughput sequencing-based assays measure different biochemical activities pertaining to gene regulation, genome-wide. These activities include transcription factor (TF)–DNA binding, enhancer activity, open chromatin, and more. A major goal is to understand underlying sequence components, or motifs, that can explain the measured activity. It is usually not one motif but a combination of motifs

Massively parallel reporter assays (MPRAs) are a high-throughput method for evaluating in vitro activities of thousands of candidate cis-regulatory elements (CREs). In these assays, candidate sequences are cloned upstream or downstream from a reporter gene tagged by unique DNA sequences. However, tag sequences may themselves affect reporter gene expression and lead to major potential biases in the

The X Chromosome plays an important role in human development and disease. However, functional genomic and disease association studies of X genes greatly lag behind autosomal gene studies, in part owing to the unique biology of X-Chromosome inactivation (XCI). Because of XCI, most genes are only expressed from one allele. Yet, ∼30% of X genes “escape” XCI and are transcribed from both alleles, many

The hexaploid bread wheat genome comprises over 16 gigabases of sequence across 21 chromosomes. Meiotic crossovers are highly polarized along the chromosomes, with elevation in the gene-dense distal regions and suppression in the Gypsy retrotransposon-dense centromere-proximal regions. We profiled the genomic landscapes of the meiotic recombinase DMC1 and the chromosome axis protein ASY1 in wheat and

Important clues about natural selection can be gleaned from discrepancies between the properties of segregating genetic variants and of mutations accumulated experimentally under minimal selection, provided the mutational process is the same in the laboratory as in nature. The base-substitution spectrum differs between C. elegans laboratory mutation accumulation (MA) experiments and the standing site-frequency

High-resolution spatial and temporal maps of gene expression have facilitated a comprehensive understanding of animal development and evolution. In nematodes, the small body size represented a major challenge for such studies, but recent advancements have helped overcome this limitation. Here, we have implemented single worm transcriptomics (SWT) in the nematode model organism Pristionchus pacificus

Mutation is the source of genetic variation and the fundament of evolution. Temperature has long been suggested to have a direct impact on realized spontaneous mutation rates. If mutation rates vary in response to environmental conditions, such as the variation of the ambient temperature through space and time, they should no longer be described as species-specific constants. By combining mutation

Inter-species comparisons of both morphology and gene expression within a phylum have revealed a period in the middle of embryogenesis with more similarity between species compared with earlier and later time points. This “developmental hourglass” pattern has been observed in many phyla, yet the evolutionary constraints on gene expression, as well as the underlying mechanisms of how this is regulated

Recombination enables reciprocal exchange of genomic information between parental chromosomes and successful segregation of homologous chromosomes during meiosis. Errors in this process lead to negative health outcomes, whereas variability in recombination rate affects genome evolution. In mammals, most crossovers occur in hotspots defined by PRDM9 motifs, although PRDM9 binding peaks are not all equally

G-quadruplexes (G4s) are noncanonical DNA secondary structures formed through the self-association of guanines, and G4s are distributed widely across the genome. G4 participates in multiple biological processes including gene transcription, and G4-targeted ligands serve as potential therapeutic agents for DNA-targeted therapies. However, genome-wide studies of the exact roles of G4s in transcriptional

Transposable elements (TEs) account for more than 50% of the human genome and many have been co-opted throughout evolution to provide regulatory functions for gene expression networks. Several lines of evidence suggest that these networks are fine-tuned by the largest family of TE controllers, the KRAB-containing zinc finger proteins (KZFPs). One tissue permissive for TE transcriptional activation

Uniparental embryos derived from only the mother (gynogenetic [GG]) or the father (androgenetic [AG]) are unique models for studying genomic imprinting and parental contributions to embryonic development. Human parthenogenetic embryos can be obtained following artificial activation of unfertilized oocytes, but the production of AG embryos by injection of two sperm into one denucleated oocyte leads

The number of de novo mutations (DNMs) in the human germline is correlated with parental age at conception, but this explains only part of the observed variation. We investigated whether there is a family-specific contribution to the number of DNMs in offspring. The analysis of DNMs in 111 dizygotic twin pairs did not identify a substantial family-specific contribution. This result was corroborated

Coalescent methods are proven and powerful tools for population genetics, phylogenetics, epidemiology, and other fields. A promising avenue for the analysis of large genomic alignments, which are increasingly common, are coalescent hidden Markov model (coalHMM) methods, but these methods have lacked general usability and flexibility. We introduce a novel method for automatically learning a coalHMM

Mutation in the germline is the ultimate source of genetic variation, but little is known about the influence of germline chromatin structure on mutational processes. Using ATAC-seq, we profile the open chromatin landscape of human spermatogonia, the most proliferative cell type of the germline, identifying transcription factor binding sites (TFBSs) and PRDM9-binding sites, a subset of which will initiate

Nucleosomes composed of histones are the fundamental units around which DNA is wrapped to form chromatin. Transcriptionally active euchromatin or repressive heterochromatin is regulated in part by the addition or removal of histone post-translational modifications (PTMs) by ‘writer’ and ‘eraser’ enzymes, respectively. Nucleosomal PTMs are recognised by a variety of ‘reader’ proteins which alter gene

Targeted transcriptional activation or interference can be induced with the CRISPR-Cas9 system (CRISPRa/CRISPRi) using nuclease-deactivated Cas9 fused to transcriptional effector molecules. These technologies have been used in cancer cell lines, particularly for genome-wide functional genetic screens using lentiviral vectors. However, CRISPRa and CRISPRi have not yet been widely applied to ex vivo

The structure of the human ribosomal DNA (rDNA) cluster has traditionally been hard to analyze due to its highly repetitive nature. However, the recent development of long-read sequencing technology, such as Oxford Nanopore sequencing, has enabled us to approach the large-scale structure of the genome. Using this technology, we found that human cells have a quite regular rDNA structure. Although each

Dictyostelium development begins with single-cell starvation and ends with multicellular fruiting bodies. Developmental morphogenesis is accompanied by sweeping transcriptional changes, encompassing nearly half of the 13,000 genes in the genome. We performed time-series RNA-sequencing analyses of the wild type and 20 mutants to explore the relationships between transcription and morphogenesis. These

Alternate isoforms are important contributors to phenotypic diversity across eukaryotes. Although short-read RNA-sequencing has increased our understanding of isoform diversity, it is challenging to accurately detect full-length transcripts, preventing the identification of many alternate isoforms. Long-read sequencing technologies have made it possible to sequence full-length alternative transcripts

Small noncoding RNAs (sRNAs) play important roles during the oocyte-to-embryo transition (OET), when the maternal phenotype is reprogrammed and the embryo genome is gradually activated. The transcriptional program driving early human development has been studied with the focus mainly on protein-coding RNAs, and expression dynamics of sRNAs remain largely unexplored. We profiled sRNAs in human oocytes

Understanding how protein function has evolved and diversified is of great importance for human genetics and medicine. Here, we tackle the problem of describing the whole transcript variability observed in several species by generalizing the definition of splicing graph. We provide a practical solution to construct parsimonious evolutionary splicing graphs where each node is a minimal transcript building

A wealth of genetic information is available describing single-nucleotide variants in the human population that appear to be well-tolerated and in and of themselves do not confer disease. These variant data sets contain signatures about the protein structure-function relationships and provide an unbiased view of various protein functions in the context of human health. This information can be used

Gut microbial communities can respond to antibiotic perturbations by rapidly altering their taxonomic and functional composition. However, little is known about the strain-level processes that drive this collective response. Here, we characterize the gut microbiome of a single individual at high temporal and genetic resolution through a period of health, disease, antibiotic treatment, and recovery

Spontaneous mutations can alter tissue dynamics and lead to cancer initiation. Although large-scale sequencing projects have illuminated processes that influence somatic mutation and subsequent tumor evolution, the mutational dynamics operating in the very early stages of cancer development are currently not well understood. To explore mutational processes in the early stages of cancer evolution, we

R-loops are stable chromatin structures comprising a DNA:RNA hybrid and a displaced single-stranded DNA. R-loops have been implicated in gene expression and chromatin structure, as well as in replication blocks and genome instability. Here, we conducted a genome-wide identification of R-loops and identified more than 700,000 R-loop peaks in the maize (Zea mays) genome. We found that sense R-loops were

RNA N6-methyladenosine (m6A) modification plays important roles in multiple aspects of RNA regulation. m6A is installed cotranscriptionally by the METTL3/14 complex, but its direct roles in RNA processing remain unclear. Here, we investigate the presence of m6A in nascent RNA of mouse embryonic stem cells. We find that around 10% of m6A peaks are located in alternative introns/exons, often close to

Hydroxycarbamide (HC, hydroxyurea) is a cytoreductive drug inducing cell cycle blockade. However, emerging evidence suggests that HC plays a role in the modulation of transcription through the activity of transcription factors and DNA methylation. Examining the global mechanism of action of HC in the context of myeloproliferative neoplasms (MPNs), for which HC is the first-line treatment, will provide

Concepts of evolutionary biology suggest that morphological change may occur by rare punctual but rather large changes, or by more steady and gradual transformations. It can therefore be asked whether genetic changes underlying morphological, physiological, and/or behavioral innovations during evolution occur in a punctual manner, whereby a single mutational event has prominent phenotypic consequences

Susumu Ohno proposed that the gene content of the mammalian X Chromosome should remain highly conserved due to dosage compensation. X Chromosome linkage (gene order) conservation is widespread in placental mammals but does not fall within the scope of Ohno's prediction and may be an indirect result of selection on gene content or selection against rearrangements that might disrupt X-Chromosome inactivation

Mammalian sex chromosomes carry large palindromes that harbor protein-coding gene families with testis-biased expression. However, there are few known examples of sex-chromosome palindromes conserved between species. We identified 26 palindromes on the human X Chromosome, constituting more than 2% of its sequence, and characterized orthologous palindromes in the chimpanzee and the rhesus macaque using

Tissue function and homeostasis reflect the gene expression signature by which the combination of ubiquitous and tissue-specific genes contribute to the tissue maintenance and stimuli-responsive function. Enhancers are central to control this tissue-specific gene expression pattern. Here, we explore the correlation between the genomic location of enhancers and their role in tissue-specific gene expression

There are more than 55,000 variable number tandem repeats (VNTRs) in the human genome, notable for both their striking polymorphism and mutability. Despite their role in human evolution and genomic variation, they have yet to be studied collectively and in detail, partially owing to their large size, variability, and predominant location in noncoding regions. Here, we examine 467 VNTRs that are human-specific

Archived formalin-fixed paraffin-embedded (FFPE) samples are the global standard format for preservation of the majority of biopsies in both basic research and translational cancer studies, and profiling chromatin accessibility in the archived FFPE tissues is fundamental to understanding gene regulation. Accurate mapping of chromatin accessibility from FFPE specimens is challenging because of the high

Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful experimental approach to study cellular heterogeneity. One of the challenges in scRNA-seq data analysis is integrating different types of biological data to consistently recognize discrete biological functions and regulatory mechanisms of cells, such as transcription factor activities and gene regulatory networks in distinct cell populations

Sequencing technologies using nucleotide conversion techniques such as cytosine to thymine in bisulfite-seq and thymine to cytosine in SLAM seq are powerful tools to explore the chemical intricacies of cellular processes. To date, no one has developed a unified methodology for aligning converted sequences and consolidating alignment of these technologies in one package. In this paper, we describe hierarchical

Bisulfite sequencing detects 5mC and 5hmC at single-base resolution. However, bisulfite treatment damages DNA, which results in fragmentation, DNA loss, and biased sequencing data. To overcome these problems, enzymatic methyl-seq (EM-seq) was developed. This method detects 5mC and 5hmC using two sets of enzymatic reactions. In the first reaction, TET2 and T4-BGT convert 5mC and 5hmC into products that

Telomeres are regions of repetitive nucleotide sequences capping the ends of eukaryotic chromosomes that protect against deterioration, and whose lengths can be correlated with age and adverse health risk factors. Yet, given their length and repetitive nature, telomeric regions are not easily reconstructed from short-read sequencing, thus making telomere sequencing, mapping, and variant resolution

Neisseria meningitidis (the meningococcus) is a major human pathogen with a history of high invasive disease burden, particularly in sub-Saharan Africa. Our current understanding of the evolution of meningococcal genomes is limited by the rarity of large-scale genomic population studies and lack of in-depth investigation of the genomic events associated with routine pathogen transmission. Here, we

Thousands of species will be sequenced in the next few years; however, understanding how their genomes work, without an unlimited budget, requires both molecular and novel evolutionary approaches. We developed a sensitive sequence alignment pipeline to identify conserved noncoding sequences (CNSs) in the Andropogoneae tribe (multiple crop species descended from a common ancestor ∼18 million years ago)

In animals, distant H3K27me3-marked Polycomb targets can establish physical interactions forming repressive chromatin hubs. In plants, growing evidence suggests that H3K27me3 acts directly or indirectly to regulate chromatin interactions, although how this histone modification modulates 3D chromatin architecture remains elusive. To decipher the impact of the dynamic deposition of H3K27me3 on the Arabidopsis

Most eukaryotic transcription factors (TFs) are part of large protein families, with members of the same family (i.e., paralogous TFs) recognizing similar DNA-binding motifs but performing different regulatory functions. Many TF paralogs are coexpressed in the cell and thus can compete for target sites across the genome. However, this competition is rarely taken into account when studying the in vivo

In contrast to the western honey bee, Apis mellifera, other honey bee species have been largely neglected despite their importance and diversity. The genetic basis of the evolutionary diversification of honey bees remains largely unknown. Here, we provide a genome-wide comparison of three honey bee species, each representing one of the three subgenera of honey bees, namely the dwarf (Apis florea),

DNA topology and alternative DNA structures are implicated in regulating diverse biological processes. Although biomechanical properties of these structures have been studied extensively in vitro, characterization in vivo, particularly in multicellular organisms, is limited. We devised new methods to map DNA supercoiling and single-stranded DNA in Caenorhabditis elegans embryos and diapause larvae

The testis transcriptome is highly complex and includes RNAs that potentially hybridize to form double-stranded RNA (dsRNA). We isolated dsRNA using the monoclonal J2 antibody and deep-sequenced the enriched samples from testes of juvenile Dicer1 knockout mice, age-matched controls, and adult animals. Comparison of our data set with recently published data from mouse liver revealed that the dsRNA transcriptome

Regulatory interactions mediated by transcription factors (TFs) make up complex networks that control cellular behavior. Fully understanding these gene regulatory networks (GRNs) offers greater insight into the consequences of disease-causing perturbations than can be achieved by studying single TF binding events in isolation. Chromosomal translocations of the lysine methyltransferase 2A (KMT2A) gene

Protein-truncating variants (PTVs) have important impacts on phenotype diversity and disease. However, their population genetics characteristics in more globally diverse populations are not well defined. Here, we describe patterns of PTVs in 1320 genes sequenced in 10,539 healthy controls and 9434 patients with psoriasis, all of Han Chinese ancestry. We identify 8720 PTVs, of which 77% are novel, and

Approximately 1% of the human genome has the ability to fold into G-quadruplexes (G4s)—noncanonical strand-specific DNA structures forming at G-rich motifs. G4s regulate several key cellular processes (e.g., transcription) and have been hypothesized to participate in others (e.g., firing of replication origins). Moreover, G4s differ in their thermostability, and this may affect their function. Yet

Heterochromatin remodeling is critical for various cell processes. In particular, the “loss of heterochromatin” phenotype in cellular senescence is associated with the process of aging and age-related disorders. Although biological processes of senescent cells, including senescence-associated heterochromatin foci (SAHF) formation, chromosome compaction, and redistribution of key proteins, have been