An integrative ENCODE resource for cancer genomics.

ENCODE comprises thousands of functional genomics datasets, and the encyclopedia covers hundreds of cell types, providing a universal annotation for genome interpretation. However, for particular applications, it may be advantageous to use a customized annotation. Here, we develop such a custom annotation by leveraging advanced assays, such as eCLIP, Hi-C, and whole-genome STARR-seq on a number of data-rich ENCODE cell types. A key aspect of this annotation is comprehensive and experimentally derived networks of both transcription factors and RNA-binding proteins (TFs and RBPs). Cancer, a disease of system-wide dysregulation, is an ideal application for such a network-based annotation. Specifically, for cancer-associated cell types, we put regulators into hierarchies and measure their network change (rewiring) during oncogenesis. We also extensively survey TF-RBP crosstalk, highlighting how SUB1, a previously uncharacterized RBP, drives aberrant tumor expression and amplifies the effect of MYC, a well-known oncogenic TF. Furthermore, we show how our annotation allows us to place oncogenic transformations in the context of a broad cell space; here, many normal-to-tumor transitions move towards a stem-like state, while oncogene knockdowns show an opposing trend. Finally, we organize the resource into a coherent workflow to prioritize key elements and variants, in addition to regulators. We showcase the application of this prioritization to somatic burdening, cancer differential expression and GWAS. Targeted validations of the prioritized regulators, elements and variants using siRNA knockdowns, CRISPR-based editing, and luciferase assays demonstrate the value of the ENCODE resource.

Dynamic RNA-protein interactions underlie the zebrafish maternal-to-zygotic transition.

During the maternal-to-zygotic transition (MZT), transcriptionally silent embryos rely on post-transcriptional regulation of maternal mRNAs until zygotic genome activation (ZGA). RNA-binding proteins (RBPs) are important regulators of post-transcriptional RNA processing events, yet their identities and functions during developmental transitions in vertebrates remain largely unexplored. Using mRNA interactome capture, we identified 227 RBPs in zebrafish embryos before and during ZGA, hereby named the zebrafish MZT mRNA-bound proteome. This protein constellation consists of many conserved RBPs, some of which are potential stage-specific mRNA interactors that likely reflect the dynamics of RNA-protein interactions during MZT. The enrichment of numerous splicing factors like hnRNP proteins before ZGA was surprising, because maternal mRNAs were found to be fully spliced. To address potentially unique roles of these RBPs in embryogenesis, we focused on Hnrnpa1. iCLIP and subsequent mRNA reporter assays revealed a function for Hnrnpa1 in the regulation of poly(A) tail length and translation of maternal mRNAs through sequence-specific association with 3' UTRs before ZGA. Comparison of iCLIP data from two developmental stages revealed that Hnrnpa1 dissociates from maternal mRNAs at ZGA and instead regulates the nuclear processing of pri-mir-430 transcripts, which we validated experimentally. The shift from cytoplasmic to nuclear RNA targets was accompanied by a dramatic translocation of Hnrnpa1 and other pre-mRNA splicing factors to the nucleus in a transcription-dependent manner. Thus, our study identifies global changes in RNA-protein interactions during vertebrate MZT and shows that Hnrnpa1 RNA-binding activities are spatially and temporally coordinated to regulate RNA metabolism during early development.

ChEA: transcription factor regulation inferred from integrating genome-wide ChIP-X experiments.

MOTIVATION: Experiments such as ChIP-chip, ChIP-seq, ChIP-PET and DamID (the four methods referred herein as ChIP-X) are used to profile the binding of transcription factors to DNA at a genome-wide scale. Such experiments provide hundreds to thousands of potential binding sites for a given transcription factor in proximity to gene coding regions. RESULTS: In order to integrate data from such studies and utilize it for further biological discovery, we collected interactions from such experiments to construct a mammalian ChIP-X database. The database contains 189,933 interactions, manually extracted from 87 publications, describing the binding of 92 transcription factors to 31,932 target genes. We used the database to analyze mRNA expression data where we perform gene-list enrichment analysis using the ChIP-X database as the prior biological knowledge gene-list library. The system is delivered as a web-based interactive application called ChIP Enrichment Analysis (ChEA). With ChEA, users can input lists of mammalian gene symbols for which the program computes over-representation of transcription factor targets from the ChIP-X database. The ChEA database allowed us to reconstruct an initial network of transcription factors connected based on shared overlapping targets and binding site proximity. To demonstrate the utility of ChEA we present three case studies. We show how by combining the Connectivity Map (CMAP) with ChEA, we can rank pairs of compounds to be used to target specific transcription factor activity in cancer cells. AVAILABILITY: The ChEA software and ChIP-X database is freely available online at: http://amp.pharm.mssm.edu/lib/chea.jsp.