The Parkinson's disease protein alpha-synuclein is a modulator of processing bodies and mRNA stability.

Alpha-synuclein (αS) is a conformationally plastic protein that reversibly binds to cellular membranes. It aggregates and is genetically linked to Parkinson's disease (PD). Here, we show that αS directly modulates processing bodies (P-bodies), membraneless organelles that function in mRNA turnover and storage. The N terminus of αS, but not other synucleins, dictates mutually exclusive binding either to cellular membranes or to P-bodies in the cytosol. αS associates with multiple decapping proteins in close proximity on the Edc4 scaffold. As αS pathologically accumulates, aberrant interaction with Edc4 occurs at the expense of physiologic decapping-module interactions. mRNA decay kinetics within PD-relevant pathways are correspondingly disrupted in PD patient neurons and brain. Genetic modulation of P-body components alters αS toxicity, and human genetic analysis lends support to the disease-relevance of these interactions. Beyond revealing an unexpected aspect of αS function and pathology, our data highlight the versatility of conformationally plastic proteins with high intrinsic disorder.

Transcriptional programs mediating neuronal toxicity and altered glial-neuronal signaling in a Drosophila knock-in tauopathy model.

Missense mutations in the gene encoding the microtubule-associated protein TAU (current and approved symbol is MAPT) cause autosomal dominant forms of frontotemporal dementia. Multiple models of frontotemporal dementia based on transgenic expression of human TAU in experimental model organisms, including Drosophila, have been described. These models replicate key features of the human disease but do not faithfully recreate the genetic context of the human disorder. Here we use CRISPR-Cas-mediated gene editing to model frontotemporal dementia caused by the TAU P301L mutation by creating the orthologous mutation, P251L, in the endogenous Drosophila tau gene. Flies heterozygous or homozygous for Tau P251L display age-dependent neurodegeneration, display metabolic defects, and accumulate DNA damage in affected neurons. To understand the molecular events promoting neuronal dysfunction and death in knock-in flies, we performed single-cell RNA sequencing on approximately 130,000 cells from brains of Tau P251L mutant and control flies. We found that expression of disease-associated mutant tau altered gene expression cell autonomously in all neuronal cell types identified. Gene expression was also altered in glial cells, suggestive of non-cell-autonomous regulation. Cell signaling pathways, including glial-neuronal signaling, were broadly dysregulated as were brain region and cell type-specific protein interaction networks and gene regulatory programs. In summary, we present here a genetic model of tauopathy that faithfully recapitulates the genetic context and phenotypic features of the human disease, and use the results of comprehensive single-cell sequencing analysis to outline pathways of neurotoxicity and highlight the potential role of non-cell-autonomous changes in glia.

Mbd3/NURD complex regulates expression of 5-hydroxymethylcytosine marked genes in embryonic stem cells.

Numerous chromatin regulators are required for embryonic stem (ES) cell self-renewal and pluripotency, but few have been studied in detail. Here, we examine the roles of several chromatin regulators whose loss affects the pluripotent state of ES cells. We find that Mbd3 and Brg1 antagonistically regulate a common set of genes by regulating promoter nucleosome occupancy. Furthermore, both Mbd3 and Brg1 play key roles in the biology of 5-hydroxymethylcytosine (5hmC): Mbd3 colocalizes with Tet1 and 5hmC in vivo, Mbd3 knockdown preferentially affects expression of 5hmC-marked genes, Mbd3 localization is Tet1-dependent, and Mbd3 preferentially binds to 5hmC relative to 5-methylcytosine in vitro. Finally, both Mbd3 and Brg1 are themselves required for normal levels of 5hmC in vivo. Together, our results identify an effector for 5hmC, and reveal that control of gene expression by antagonistic chromatin regulators is a surprisingly common regulatory strategy in ES cells.

FLED: a full-length eccDNA detector for long-reads sequencing data.

Reconstructing the full-length sequence of extrachromosomal circular DNA (eccDNA) from short sequencing reads has proved challenging given the similarity of eccDNAs and their corresponding linear DNAs. Previous sequencing methods were unable to achieve high-throughput detection of full-length eccDNAs. Herein, a novel algorithm was developed, called Full-Length eccDNA Detection (FLED), to reconstruct the sequence of eccDNAs based on the strategy that combined rolling circle amplification and nanopore long-reads sequencing technology. Seven human epithelial and cancer cell line samples were analyzed by FLED and over 5000 full-length eccDNAs were identified per sample. The structures of identified eccDNAs were validated by both Polymerase Chain Reaction (PCR) and Sanger sequencing. Compared to other published nanopore-based eccDNA detectors, FLED exhibited higher sensitivity. In cancer cell lines, the genes overlapped with eccDNA regions were enriched in cancer-related pathways and cis-regulatory elements can be predicted in the upstream or downstream of intact genes on eccDNA molecules, and the expressions of these cancer-related genes were dysregulated in tumor cell lines, indicating the regulatory potency of eccDNAs in biological processes. The proposed method takes advantage of nanopore long reads and enables unbiased reconstruction of full-length eccDNA sequences. FLED is implemented using Python3 which is freely available on GitHub (https://github.com/FuyuLi/FLED).

Exosome detection via the ultrafast-isolation system: EXODUS.

Exosomes have shown great potential in disease diagnostics and therapeutics. However, current isolation approaches are burdensome and suffer from low speed, yield and purity, limiting basic research and clinical applications. Here, we describe an efficient exosome detection method via the ultrafast-isolation system (EXODUS) that allows automated label-free purification of exosomes from varied biofluids. We obtained the ultra-efficient purification of exosomes by negative pressure oscillation and double coupled harmonic oscillator-enabled membrane vibration. Our two coupled oscillators generate dual-frequency transverse waves on the membranes, enabling EXODUS to outperform other isolation techniques in speed, purity and yield. We demonstrated EXODUS by purifying exosomes from urine samples of 113 patients and validated the practical relevance in exosomal RNA profiling with the high-resolution capability and high-throughput analysis.

WebCSEA: web-based cell-type-specific enrichment analysis of genes.

Human complex traits and common diseases show tissue- and cell-type- specificity. Recently, single-cell RNA sequencing (scRNA-seq) technology has successfully depicted cellular heterogeneity in human tissue, providing an unprecedented opportunity to understand the context-specific expression of complex trait-associated genes in human tissue-cell types (TCs). Here, we present the first web-based application to quickly assess the cell-type-specificity of genes, named Web-based Cell-type Specific Enrichment Analysis of Genes (WebCSEA, available at https://bioinfo.uth.edu/webcsea/). Specifically, we curated a total of 111 scRNA-seq panels of human tissues and 1,355 TCs from 61 different general tissues across 11 human organ systems. We adapted our previous decoding tissue-specificity (deTS) algorithm to measure the enrichment for each tissue-cell type (TC). To overcome the potential bias from the number of signature genes between different TCs, we further developed a permutation-based method that accurately estimates the TC-specificity of a given inquiry gene list. WebCSEA also provides an interactive heatmap that displays the cell-type specificity across 1355 human TCs, and other interactive and static visualizations of cell-type specificity by human organ system, developmental stage, and top-ranked tissues and cell types. In short, WebCSEA is a one-click application that provides a comprehensive exploration of the TC-specificity of genes among human major TC map.

The genetic source tracking of human urinary exosomes.

The genetic origins of nanoscale extracellular vesicles in our body fluids remains unclear. Here, we perform a tracking analysis of urinary exosomes via RNA sequencing, revealing that urine exosomes mostly express tissue-specific genes for the bladder and have close cell-genetic relationships to the endothelial cell, basal cell, monocyte, and dendritic cell. Tracking the differentially expressed genes of cancers and corresponding enrichment analysis show urine exosomes are intensively involved in immune activities, indicating that they may be harnessed as reliable biomarkers of noninvasive liquid biopsy in cancer genomic diagnostics and precision medicine.

Plasma and CSF biomarkers of aging and cognitive decline in Caribbean vervets.

INTRODUCTION: Vervets are non-human primates that share high genetic homology with humans and develop amyloid beta (Aß) pathology with aging. We expand current knowledge by examining Aß pathology, aging, cognition, and biomarker proteomics. METHODS: Amyloid immunoreactivity in the frontal cortex and temporal cortex/hippocampal regions from archived vervet brain samples ranging from young adulthood to old age was quantified. We also obtained cognitive scores, plasma samples, and cerebrospinal fluid (CSF) samples in additional animals. Plasma and CSF proteins were quantified with platforms utilizing human antibodies. RESULTS: We found age-related increases in Aß deposition in both brain regions. Bioinformatic analyses assessed associations between biomarkers and age, sex, cognition, and CSF Aß levels, revealing changes in proteins related to immune-related inflammation, metabolism, and cellular processes. DISCUSSION: Vervets are an effective model of aging and early-stage Alzheimer's disease, and we provide translational biomarker data that both align with previous results in humans and provide a basis for future investigations. HIGHLIGHTS: We found changes in immune and metabolic plasma biomarkers associated with age and cognition. Cerebrospinal fluid (CSF) biomarkers revealed changes in cell signaling indicative of adaptative processes. TNFRSF19 (TROY) and Artemin co-localize with Alzheimer's disease pathology. Vervets are a relevant model for translational studies of early-stage Alzheimer's disease.

powerEQTL: an R package and shiny application for sample size and power calculation of bulk tissue and single-cell eQTL analysis.

SUMMARY: Genome-wide association studies (GWAS) have revealed thousands of genetic loci for common diseases. One of the main challenges in the post-GWAS era is to understand the causality of the genetic variants. Expression quantitative trait locus (eQTL) analysis is an effective way to address this question by examining the relationship between gene expression and genetic variation in a sufficiently powered cohort. However, it is frequently a challenge to determine the sample size at which a variant with a specific allele frequency will be detected to associate with gene expression with sufficient power. This is a particularly difficult task for single-cell RNAseq studies. Therefore, a user-friendly tool to estimate statistical power for eQTL analyses in both bulk tissue and single-cell data is needed. Here, we presented an R package called powerEQTL with flexible functions to estimate power, minimal sample size or detectable minor allele frequency for both bulk tissue and single-cell eQTL analysis. A user-friendly, program-free web application is also provided, allowing users to calculate and visualize the parameters interactively. AVAILABILITY AND IMPLEMENTATION: The powerEQTL R package source code and online tutorial are freely available at CRAN: https://cran.r-project.org/web/packages/powerEQTL/. The R shiny application is publicly hosted at https://bwhbioinfo.shinyapps.io/powerEQTL/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A pipeline for RNA-seq based eQTL analysis with automated quality control procedures.

BACKGROUND: Advances in the expression quantitative trait loci (eQTL) studies have provided valuable insights into the mechanism of diseases and traits-associated genetic variants. However, it remains challenging to evaluate and control the quality of multi-source heterogeneous eQTL raw data for researchers with limited computational background. There is an urgent need to develop a powerful and user-friendly tool to automatically process the raw datasets in various formats and perform the eQTL mapping afterward. RESULTS: In this work, we present a pipeline for eQTL analysis, termed eQTLQC, featured with automated data preprocessing for both genotype data and gene expression data. Our pipeline provides a set of quality control and normalization approaches, and utilizes automated techniques to reduce manual intervention. We demonstrate the utility and robustness of this pipeline by performing eQTL case studies using multiple independent real-world datasets with RNA-seq data and whole genome sequencing (WGS) based genotype data. CONCLUSIONS: eQTLQC provides a reliable computational workflow for eQTL analysis. It provides standard quality control and normalization as well as eQTL mapping procedures for eQTL raw data in multiple formats. The source code, demo data, and instructions are freely available at https://github.com/stormlovetao/eQTLQC .

Genetic Modifiers of Age at Onset for Parkinson's Disease in Asians: A Genome-Wide Association Study.

BACKGROUND: Age at onset (AAO) is an essential feature of Parkinson's disease (PD) and can help predict disease progression and mortality. Identification of genetic variants influencing AAO of PD could lead to a better understanding of the disease's biological mechanism and provide clinical guidance. However, genetic determinants for AAO of PD remain mostly unknown, especially in the Asian population. OBJECTIVES: To identify genetic determinants for AAO of PD in the Asian population. METHODS: We performed a genome-wide association meta-analysis on AAO of PD in 5166 Chinese patients with PD (Ndiscovery  = 3628, Nreplication  = 1538). We then conducted a further cross-ethnic meta-analysis using our results and summary statistics for the AAO of PD from the European population. RESULTS: The total heritability of AAO of PD was around 0.10 ~ 0.14, similar to that (~0.11) estimated in populations of European ancestry. One novel significant intergenic locus rs9783733 (NDN; PWRN4) was identified (P = 3.14E-09, beta = 2.30, SE = 0.39). Remarkably, this variant could delay AAO of PD by ~2.43 years, with a more considerable effect on males (~3.18 years) than females (~1.45 years). The variant was suggestively significant in the cross-ethnic meta-analysis and suggested a positive selection in the East Asian population. Additionally, cross-ethnic meta-analysis identified a significant locus rs356203 in SNCA (P = 2.35E-11, beta = -0.71, SE = 0.01). CONCLUSIONS: These findings improve the current understanding of the genetic etiology of AAO of PD in different ethnic groups, and provide a new target for further research on PD pathogenesis and potential therapeutic options. © 2021 International Parkinson and Movement Disorder Society.

Accelerating Medicines Partnership: Parkinson's Disease. Genetic Resource.

BACKGROUND: Whole-genome sequencing data are available from several large studies across a variety of diseases and traits. However, massive storage and computation resources are required to use these data, and to achieve sufficient power for discoveries, harmonization of multiple cohorts is critical. OBJECTIVES: The Accelerating Medicines Partnership Parkinson's Disease program has developed a research platform for Parkinson's disease (PD) that integrates the storage and analysis of whole-genome sequencing data, RNA expression data, and clinical data, harmonized across multiple cohort studies. METHODS: The version 1 release contains whole-genome sequencing data derived from 3941 participants from 4 cohorts. Samples underwent joint genotyping by the TOPMed Freeze 9 Variant Calling Pipeline. We performed descriptive analyses of these whole-genome sequencing data using the Accelerating Medicines Partnership Parkinson's Disease platform. RESULTS: The clinical diagnosis of participants in version 1 release includes 2005 idiopathic PD patients, 963 healthy controls, 64 prodromal subjects, 62 clinically diagnosed PD subjects without evidence of dopamine deficit, and 705 participants of genetically enriched cohorts carrying PD risk-associated GBA variants or LRRK2 variants, of whom 304 were affected. We did not observe significant enrichment of pathogenic variants in the idiopathic PD group, but the polygenic risk score was higher in PD both in nongenetically enriched cohorts and genetically enriched cohorts. The population analysis showed a correlation between genetically enriched cohorts and Ashkenazi Jewish ancestry. CONCLUSIONS: We describe the genetic component of the Accelerating Medicines Partnership Parkinson's Disease platform, a solution to democratize data access and analysis for the PD research community. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. This article is a U.S. Government work and is in the public domain in the USA.

An ancient transcription factor initiates the burst of piRNA production during early meiosis in mouse testes.

Animal germ cells produce PIWI-interacting RNAs (piRNAs), small silencing RNAs that suppress transposons and enable gamete maturation. Mammalian transposon-silencing piRNAs accumulate early in spermatogenesis, whereas pachytene piRNAs are produced later during postnatal spermatogenesis and account for >95% of all piRNAs in the adult mouse testis. Mutants defective for pachytene piRNA pathway proteins fail to produce mature sperm, but neither the piRNA precursor transcripts nor the trigger for pachytene piRNA production is known. Here, we show that the transcription factor A-MYB initiates pachytene piRNA production. A-MYB drives transcription of both pachytene piRNA precursor RNAs and the mRNAs for core piRNA biogenesis factors including MIWI, the protein through which pachytene piRNAs function. A-MYB regulation of piRNA pathway proteins and piRNA genes creates a coherent feedforward loop that ensures the robust accumulation of pachytene piRNAs. This regulatory circuit, which can be detected in rooster testes, likely predates the divergence of birds and mammals.

High-resolution analysis of differential gene expression during skeletal muscle atrophy and programmed cell death.

Skeletal muscles can undergo atrophy and/or programmed cell death (PCD) during development or in response to a wide range of insults, including immobility, cachexia, and spinal cord injury. However, the protracted nature of atrophy and the presence of multiple cell types within the tissue complicate molecular analyses. One model that does not suffer from these limitations is the intersegmental muscle (ISM) of the tobacco hawkmoth Manduca sexta. Three days before the adult eclosion (emergence) at the end of metamorphosis, the ISMs initiate a nonpathological program of atrophy that results in a 40% loss of mass. The ISMs then generate the eclosion behavior and initiate a nonapoptotic PCD during the next 30 h. We have performed a comprehensive transcriptomics analysis of all mRNAs and microRNAs throughout ISM development to better understand the molecular mechanisms that mediate atrophy and death. Atrophy involves enhanced protein catabolism and reduced expression of the genes involved in respiration, adhesion, and the contractile apparatus. In contrast, PCD involves the induction of numerous proteases, DNA methylases, membrane transporters, ribosomes, and anaerobic metabolism. These changes in gene expression are largely repressed when insects are injected with the insect steroid hormone 20-hydroxyecdysone, which delays death. The expression of the death-associated proteins may be greatly enhanced by reductions in specific microRNAs that function to repress translation. This study not only provides fundamental new insights into basic developmental processes, it may also represent a powerful resource for identifying potential diagnostic markers and molecular targets for therapeutic intervention.

Two independent transcription initiation codes overlap on vertebrate core promoters.

A core promoter is a stretch of DNA surrounding the transcription start site (TSS) that integrates regulatory inputs and recruits general transcription factors to initiate transcription. The nature and causative relationship of the DNA sequence and chromatin signals that govern the selection of most TSSs by RNA polymerase II remain unresolved. Maternal to zygotic transition represents the most marked change of the transcriptome repertoire in the vertebrate life cycle. Early embryonic development in zebrafish is characterized by a series of transcriptionally silent cell cycles regulated by inherited maternal gene products: zygotic genome activation commences at the tenth cell cycle, marking the mid-blastula transition. This transition provides a unique opportunity to study the rules of TSS selection and the hierarchy of events linking transcription initiation with key chromatin modifications. We analysed TSS usage during zebrafish early embryonic development at high resolution using cap analysis of gene expression, and determined the positions of H3K4me3-marked promoter-associated nucleosomes. Here we show that the transition from the maternal to zygotic transcriptome is characterized by a switch between two fundamentally different modes of defining transcription initiation, which drive the dynamic change of TSS usage and promoter shape. A maternal-specific TSS selection, which requires an A/T-rich (W-box) motif, is replaced with a zygotic TSS selection grammar characterized by broader patterns of dinucleotide enrichments, precisely aligned with the first downstream (+1) nucleosome. The developmental dynamics of the H3K4me3-marked nucleosomes reveal their DNA-sequence-associated positioning at promoters before zygotic transcription and subsequent transcription-independent adjustment to the final position downstream of the zygotic TSS. The two TSS-defining grammars coexist, often physically overlapping, in core promoters of constitutively expressed genes to enable their expression in the two regulatory environments. The dissection of overlapping core promoter determinants represents a framework for future studies of promoter structure and function across different regulatory contexts.

Epigenetic dysregulation of hairy and enhancer of split 4 (HES4) is associated with striatal degeneration in postmortem Huntington brains.

To investigate epigenetic contributions to Huntington's disease (HD) pathogenesis, we carried out genome-wide mapping of the transcriptional mark, trimethyl-histone H3-lysine 4 (H3K4me3) in neuronal nuclei extracted from prefrontal cortex of HD cases and controls using chromatin immunoprecipitation followed by deep-sequencing. Neuron-specific mapping of the genome-wide distribution of H3K4me3 revealed 136 differentially enriched loci associated with genes implicated in neuronal development and neurodegeneration, including GPR3, TMEM106B, PDIA6 and the Notch signaling genes hairy and enhancer of split 4 (HES4) and JAGGED2, supporting the view that the neuronal epigenome is affected in HD. Importantly, loss of H3K4me3 at CpG-rich sequences on the HES4 promoter was associated with excessive DNA methylation, reduced binding of nuclear proteins to the methylated region and altered expression of HES4 and HES4 targeted genes MASH1 and P21 involved in striatal development. Moreover, hypermethylation of HES4 promoter sequences was strikingly correlated with measures of striatal degeneration and age-of-onset in a cohort of 25 HD brains (r = 0.56, P = 0.006). Lastly, shRNA knockdown of HES4 in human neuroblastoma cells altered MASH1 and P21 mRNA expression and markedly increased mutated HTT-induced aggregates and cell death. These findings, taken together, suggest that epigenetic dysregulation of HES4 could play a critical role in modifying HD disease pathogenesis and severity.

MicroRNAs located in the Hox gene clusters are implicated in huntington's disease pathogenesis.

Transcriptional dysregulation has long been recognized as central to the pathogenesis of Huntington's disease (HD). MicroRNAs (miRNAs) represent a major system of post-transcriptional regulation, by either preventing translational initiation or by targeting transcripts for storage or for degradation. Using next-generation miRNA sequencing in prefrontal cortex (Brodmann Area 9) of twelve HD and nine controls, we identified five miRNAs (miR-10b-5p, miR-196a-5p, miR-196b-5p, miR-615-3p and miR-1247-5p) up-regulated in HD at genome-wide significance (FDR q-value<0.05). Three of these, miR-196a-5p, miR-196b-5p and miR-615-3p, were expressed at near zero levels in control brains. Expression was verified for all five miRNAs using reverse transcription quantitative PCR and all but miR-1247-5p were replicated in an independent sample (8HD/8C). Ectopic miR-10b-5p expression in PC12 HTT-Q73 cells increased survival by MTT assay and cell viability staining suggesting increased expression may be a protective response. All of the miRNAs but miR-1247-5p are located in intergenic regions of Hox clusters. Total mRNA sequencing in the same samples identified fifteen of 55 genes within the Hox cluster gene regions as differentially expressed in HD, and the Hox genes immediately adjacent to the four Hox cluster miRNAs as up-regulated. Pathway analysis of mRNA targets of these miRNAs implicated functions for neuronal differentiation, neurite outgrowth, cell death and survival. In regression models among the HD brains, huntingtin CAG repeat size, onset age and age at death were independently found to be inversely related to miR-10b-5p levels. CAG repeat size and onset age were independently inversely related to miR-196a-5p, onset age was inversely related to miR-196b-5p and age at death was inversely related to miR-615-3p expression. These results suggest these Hox-related miRNAs may be involved in neuroprotective response in HD. Recently, miRNAs have shown promise as biomarkers for human diseases and given their relationship to disease expression, these miRNAs are biomarker candidates in HD.

Association between α-synuclein blood transcripts and early, neuroimaging-supported Parkinson's disease.

There are no cures for neurodegenerative diseases and this is partially due to the difficulty of monitoring pathogenic molecules in patients during life. The Parkinson's disease gene α-synuclein (SNCA) is selectively expressed in blood cells and neurons. Here we show that SNCA transcripts in circulating blood cells are paradoxically reduced in early stage, untreated and dopamine transporter neuroimaging-supported Parkinson's disease in three independent regional, national, and international populations representing 500 cases and 363 controls and on three analogue and digital platforms with P < 0.0001 in meta-analysis. Individuals with SNCA transcripts in the lowest quartile of counts had an odds ratio for Parkinson's disease of 2.45 compared to individuals in the highest quartile. Disease-relevant transcript isoforms were low even near disease onset. Importantly, low SNCA transcript abundance predicted cognitive decline in patients with Parkinson's disease during up to 5 years of longitudinal follow-up. This study reveals a consistent association of reduced SNCA transcripts in accessible peripheral blood and early-stage Parkinson's disease in 863 participants and suggests a clinical role as potential predictor of cognitive decline. Moreover, the three independent biobank cohorts provide a generally useful platform for rapidly validating any biological marker of this common disease.

Sequence features and chromatin structure around the genomic regions bound by 119 human transcription factors.

Chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq) has become the dominant technique for mapping transcription factor (TF) binding regions genome-wide. We performed an integrative analysis centered around 457 ChIP-seq data sets on 119 human TFs generated by the ENCODE Consortium. We identified highly enriched sequence motifs in most data sets, revealing new motifs and validating known ones. The motif sites (TF binding sites) are highly conserved evolutionarily and show distinct footprints upon DNase I digestion. We frequently detected secondary motifs in addition to the canonical motifs of the TFs, indicating tethered binding and cobinding between multiple TFs. We observed significant position and orientation preferences between many cobinding TFs. Genes specifically expressed in a cell line are often associated with a greater occurrence of nearby TF binding in that cell line. We observed cell-line-specific secondary motifs that mediate the binding of the histone deacetylase HDAC2 and the enhancer-binding protein EP300. TF binding sites are located in GC-rich, nucleosome-depleted, and DNase I sensitive regions, flanked by well-positioned nucleosomes, and many of these features show cell type specificity. The GC-richness may be beneficial for regulating TF binding because, when unoccupied by a TF, these regions are occupied by nucleosomes in vivo. We present the results of our analysis in a TF-centric web repository Factorbook (http://factorbook.org) and will continually update this repository as more ENCODE data are generated.

Understanding transcriptional regulation by integrative analysis of transcription factor binding data.

Statistical models have been used to quantify the relationship between gene expression and transcription factor (TF) binding signals. Here we apply the models to the large-scale data generated by the ENCODE project to study transcriptional regulation by TFs. Our results reveal a notable difference in the prediction accuracy of expression levels of transcription start sites (TSSs) captured by different technologies and RNA extraction protocols. In general, the expression levels of TSSs with high CpG content are more predictable than those with low CpG content. For genes with alternative TSSs, the expression levels of downstream TSSs are more predictable than those of the upstream ones. Different TF categories and specific TFs vary substantially in their contributions to predicting expression. Between two cell lines, the differential expression of TSS can be precisely reflected by the difference of TF-binding signals in a quantitative manner, arguing against the conventional on-and-off model of TF binding. Finally, we explore the relationships between TF-binding signals and other chromatin features such as histone modifications and DNase hypersensitivity for determining expression. The models imply that these features regulate transcription in a highly coordinated manner.