Human proteins that interact with RNA/DNA hybrids

RNA/DNA hybrids form when RNA hybridizes with its template DNA generating a three-stranded structure known as the R-loop. Knowledge of how they form and resolve, as well as their functional roles, is limited. Here, by pull-down assays followed by mass spectrometry, we identified 803 proteins that bind to RNA/DNA hybrids. Because these proteins were identified using in vitro assays, we confirmed that they bind to R-loops in vivo. They include proteins that are involved in a variety of functions, including most steps of RNA processing. The proteins are enriched for K homology (KH) and helicase domains. Among them, more than 300 proteins preferred binding to hybrids than double-stranded DNA. These proteins serve as starting points for mechanistic studies to elucidate what RNA/DNA hybrids regulate and how they are regulated.

[1]  N. Gromak,et al.  RNA/DNA Hybrid Interactome Identifies DXH9 as a Molecular Player in Transcriptional Termination and R-Loop-Associated DNA Damage , 2018, Cell reports.

[2]  Joshua T. Burdick,et al.  Senataxin Mutation Reveals How R-Loops Promote Transcription by Blocking DNA Methylation at Gene Promoters. , 2018, Molecular cell.

[3]  Matthias W. Hentze,et al.  A brave new world of RNA-binding proteins , 2018, Nature Reviews Molecular Cell Biology.

[4]  H. Ochs,et al.  R‐loops cause genomic instability in T helper lymphocytes from patients with Wiskott‐Aldrich syndrome , 2017, The Journal of allergy and clinical immunology.

[5]  Yvonne N Fondufe-Mittendorf,et al.  Transcriptome-wide identification of the RNA-binding landscape of the chromatin-associated protein PARP1 reveals functions in RNA biogenesis , 2017, Cell Discovery.

[6]  C. Shao,et al.  R-ChIP Using Inactive RNase H Reveals Dynamic Coupling of R-loops with Transcriptional Pausing at Gene Promoters. , 2017, Molecular cell.

[7]  Lorenzo Manti,et al.  The RNA processing factors THRAP3 and BCLAF1 promote the DNA damage response through selective mRNA splicing and nuclear export , 2017, Nucleic acids research.

[8]  Ian M. Carr,et al.  m6aViewer: software for the detection, analysis, and visualization of N6-methyladenosine peaks from m6A-seq/ME-RIP sequencing data , 2017, RNA.

[9]  T. Graubert,et al.  Functions of Replication Protein A as a Sensor of R Loops and a Regulator of RNaseH1. , 2017, Molecular cell.

[10]  Anjali D. Zimmer,et al.  S1-DRIP-seq identifies high expression and polyA tracts as major contributors to R-loop formation , 2016, Genes & development.

[11]  Jennifer A. Doudna,et al.  Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage , 2016, Science.

[12]  W. Baumeister,et al.  Structure of transcribing mammalian RNA polymerase II , 2016, Nature.

[13]  K. Cimprich,et al.  Transcription-coupled nucleotide excision repair factors promote R-loop-induced genome instability. , 2014, Molecular cell.

[14]  Eli Rothenberg,et al.  Mechanistic Insight into the Interaction of BLM helicase with Intra-strand G-quadruplex Structures , 2014, Nature Communications.

[15]  D. Tollervey,et al.  Genome-Wide Distribution of RNA-DNA Hybrids Identifies RNase H Targets in tRNA Genes, Retrotransposons and Mitochondria , 2014, PLoS genetics.

[16]  M. Nowotny,et al.  Crystal structure of RNase H3–substrate complex reveals parallel evolution of RNA/DNA hybrid recognition , 2014, Nucleic Acids Research.

[17]  A Keith Dunker,et al.  Intrinsically disordered proteins and intrinsically disordered protein regions. , 2014, Annual review of biochemistry.

[18]  E. Mercken,et al.  RNA-Binding Protein AUF1 Promotes Myogenesis by Regulating MEF2C Expression Levels , 2014, Molecular and Cellular Biology.

[19]  N. Gromak,et al.  R-loops Associated with Triplet Repeat Expansions Promote Gene Silencing in Friedreich Ataxia and Fragile X Syndrome , 2014, PLoS genetics.

[20]  H. Leonhardt,et al.  Poly(ADP-ribose) Polymerase 1 (PARP1) Associates with E3 Ubiquitin-Protein Ligase UHRF1 and Modulates UHRF1 Biological Functions* , 2014, The Journal of Biological Chemistry.

[21]  Zoltán Konthur,et al.  Identification and characterization of RNA guanine-quadruplex binding proteins , 2014, Nucleic acids research.

[22]  A. Hamza,et al.  Genome-Wide Profiling of Yeast DNA:RNA Hybrid Prone Sites with DRIP-Chip , 2014, PLoS genetics.

[23]  Patrick G. Shaw,et al.  C9orf72 Nucleotide Repeat Structures Initiate Molecular Cascades of Disease , 2014, Nature.

[24]  Feng Zhang,et al.  Crystal Structure of Cas9 in Complex with Guide RNA and Target DNA , 2014, Cell.

[25]  Gene W. Yeo,et al.  Genome-wide analysis reveals SR protein cooperation and competition in regulated splicing. , 2013, Molecular cell.

[26]  R. Kurokawa,et al.  Regulation of telomere length by G-quadruplex telomere DNA- and TERRA-binding protein TLS/FUS. , 2013, Chemistry & biology.

[27]  Bronwen L. Aken,et al.  GENCODE: The reference human genome annotation for The ENCODE Project , 2012, Genome research.

[28]  Richard Bonneau,et al.  The mRNA-bound proteome and its global occupancy profile on protein-coding transcripts. , 2012, Molecular cell.

[29]  I. Korf,et al.  R-loop formation is a distinctive characteristic of unmethylated human CpG island promoters. , 2012, Molecular cell.

[30]  C. Bond,et al.  Structure of the heterodimer of human NONO and paraspeckle protein component 1 and analysis of its role in subnuclear body formation , 2012, Proceedings of the National Academy of Sciences.

[31]  D. Koshland,et al.  RNase H and multiple RNA biogenesis factors cooperate to prevent RNA:DNA hybrids from generating genome instability. , 2011, Molecular cell.

[32]  C. Bond,et al.  Crystallization of a paraspeckle protein PSPC1-NONO heterodimer. , 2011, Acta crystallographica. Section F, Structural biology and crystallization communications.

[33]  Uwe Ohler,et al.  PARalyzer: definition of RNA binding sites from PAR-CLIP short-read sequence data , 2011, Genome Biology.

[34]  Konstantina Skourti-Stathaki,et al.  Human Senataxin Resolves RNA/DNA Hybrids Formed at Transcriptional Pause Sites to Promote Xrn2-Dependent Termination , 2011, Molecular cell.

[35]  L. Steinmetz,et al.  Yeast Sen1 Helicase Protects the Genome from Transcription-Associated Instability , 2011, Molecular cell.

[36]  Toshiro K. Ohsumi,et al.  Genome-wide identification of polycomb-associated RNAs by RIP-seq. , 2010, Molecular cell.

[37]  H. Chon,et al.  Crystal Structures of RNase H2 in Complex with Nucleic Acid Reveal the Mechanism of RNA-DNA Junction Recognition and Cleavage , 2010, Molecular cell.

[38]  Scott B. Dewell,et al.  Transcriptome-wide Identification of RNA-Binding Protein and MicroRNA Target Sites by PAR-CLIP , 2010, Cell.

[39]  Serban Nacu,et al.  Fast and SNP-tolerant detection of complex variants and splicing in short reads , 2010, Bioinform..

[40]  Daekyu Sun,et al.  Identification and Characterization of Nucleolin as a c-myc G-quadruplex-binding Protein* , 2009, The Journal of Biological Chemistry.

[41]  M. Lieber,et al.  G Clustering Is Important for the Initiation of Transcription-Induced R-Loops In Vitro, whereas High G Density without Clustering Is Sufficient Thereafter , 2009, Molecular and Cellular Biology.

[42]  R. Crouch,et al.  Ribonuclease H: the enzymes in eukaryotes , 2009, The FEBS journal.

[43]  M. Mann,et al.  MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification , 2008, Nature Biotechnology.

[44]  Clifford A. Meyer,et al.  Model-based Analysis of ChIP-Seq (MACS) , 2008, Genome Biology.

[45]  W. Kraus,et al.  Reciprocal Binding of PARP-1 and Histone H1 at Promoters Specifies Transcriptional Outcomes , 2008, Science.

[46]  K. Mitsuya,et al.  The SRA protein Np95 mediates epigenetic inheritance by recruiting Dnmt1 to methylated DNA , 2007, Nature.

[47]  Ralf P. Richter,et al.  FG-Rich Repeats of Nuclear Pore Proteins Form a Three-Dimensional Meshwork with Hydrogel-Like Properties , 2006, Science.

[48]  Zsuzsanna Dosztányi,et al.  IUPred: web server for the prediction of intrinsically unstructured regions of proteins based on estimated energy content , 2005, Bioinform..

[49]  J. Manley,et al.  Inactivation of the SR Protein Splicing Factor ASF/SF2 Results in Genomic Instability , 2005, Cell.

[50]  Andrés Aguilera,et al.  Cotranscriptionally formed DNA:RNA hybrids mediate transcription elongation impairment and transcription-associated recombination. , 2003, Molecular cell.

[51]  E. Wahle,et al.  The RNA Binding Domains of the Nuclear poly(A)-binding Protein* 210 , 2003, The Journal of Biological Chemistry.

[52]  M. Lieber,et al.  R-loops at immunoglobulin class switch regions in the chromosomes of stimulated B cells , 2003, Nature Immunology.

[53]  M. Deutscher,et al.  The Physiological Role of RNase T Can Be Explained by Its Unusual Substrate Specificity* , 2002, The Journal of Biological Chemistry.

[54]  I. Hickson,et al.  Inhibition of the Bloom's and Werner's syndrome helicases by G-quadruplex interacting ligands. , 2001, Biochemistry.

[55]  E. Nudler,et al.  The RNA–DNA Hybrid Maintains the Register of Transcription by Preventing Backtracking of RNA Polymerase , 1997, Cell.

[56]  D. A. Clayton,et al.  RNA‐DNA hybrid formation at the human mitochondrial heavy‐strand origin ceases at replication start sites: an implication for RNA‐DNA hybrids serving as primers. , 1996, The EMBO journal.

[57]  K. Murthy,et al.  The 160-kD subunit of human cleavage-polyadenylation specificity factor coordinates pre-mRNA 3'-end formation. , 1995, Genes & development.

[58]  R. Crouch,et al.  The non-RNase H domain of Saccharomyces cerevisiae RNase H1 binds double-stranded RNA: magnesium modulates the switch between double-stranded RNA binding and RNase H activity. , 1995, RNA.

[59]  B. Reid,et al.  Structure of a DNA:RNA hybrid duplex. Why RNase H does not cleave pure RNA. , 1993, Journal of molecular biology.

[60]  A. Kumar,et al.  Mammalian heterogeneous ribonucleoprotein A1 and its constituent domains. Nucleic acid interaction, structural stability and self-association. , 1993, Journal of molecular biology.

[61]  D. Crothers,et al.  Stability and properties of double and triple helices: dramatic effects of RNA or DNA backbone composition. , 1992, Science.

[62]  D. Lilley,et al.  RNA bulges and the helical periodicity of double-stranded RNA , 1990, Nature.

[63]  T. Baker,et al.  Transcriptional activation of initiation of replication from the E. coli chromosomal origin: An RNA-DNA hybrid near oriC , 1988, Cell.

[64]  A. Rich,et al.  Raman spectra of single crystals of r(GCG)d(CGC) and d(CCCCGGGG) as models for A DNA, their structure transitions in aqueous solution, and comparison with double-helical poly(dG).poly(dC). , 1986, Biochemistry.

[65]  C. Meares,et al.  Topography of transcription: path of the leading end of nascent RNA through the Escherichia coli transcription complex. , 1983, Proceedings of the National Academy of Sciences of the United States of America.

[66]  W. Keller,et al.  Degradation of DNA RNA hybrids by ribonuclease H and DNA polymerases of cellular and viral origin. , 1972, Proceedings of the National Academy of Sciences of the United States of America.

[67]  P. Hausen,et al.  Enzyme from Calf Thymus Degrading the RNA Moiety of DNA-RNA Hybrids: Effect on DNA-Dependent RNA Polymerase , 1969, Science.

[68]  Lesley M. Berghuis,et al.  DNA/RNA hybrid substrates modulate the catalytic activity of purified AID , 2018, Molecular immunology.

[69]  The UniProt Consortium.  UniProt: the universal protein knowledgebase , 2016, Nucleic Acids Res..

[70]  Joshua E. Elias,et al.  Target-Decoy Search Strategy for Mass Spectrometry-Based Proteomics , 2010, Proteome Bioinformatics.

[71]  Cathy H. Wu,et al.  UniProt: the Universal Protein knowledgebase , 2004, Nucleic Acids Res..