A new view of transcriptome complexity and regulation through the lens of local splicing variations

Alternative splicing (AS) can critically affect gene function and disease, yet mapping splicing variations remains a challenge. Here, we propose a new approach to define and quantify mRNA splicing in units of local splicing variations (LSVs). LSVs capture previously defined types of alternative splicing as well as more complex transcript variations. Building the first genome wide map of LSVs from twelve mouse tissues, we find complex LSVs constitute over 30% of tissue dependent transcript variations and affect specific protein families. We show the prevalence of complex LSVs is conserved in humans and identify hundreds of LSVs that are specific to brain subregions or altered in Alzheimer's patients. Amongst those are novel isoforms in the Camk2 family and a novel poison exon in Ptbp1, a key splice factor in neurogenesis. We anticipate the approach presented here will advance the ability to relate tissue-specific splice variation to genetic variation, phenotype, and disease. DOI: http://dx.doi.org/10.7554/eLife.11752.001

[1]  S. Shen-Orr,et al.  Network motifs: simple building blocks of complex networks. , 2002, Science.

[2]  Douglas L Black,et al.  Neuronal regulation of pre-mRNA splicing by polypyrimidine tract binding proteins, PTBP1 and PTBP2 , 2012, Critical reviews in biochemistry and molecular biology.

[3]  D. Black,et al.  MicroRNAs regulate the expression of the alternative splicing factor nPTB during muscle development. , 2007, Genes & development.

[4]  Bo Torben Porse,et al.  Psd-95 is post-transcriptionally repressed during early neural development by PTBP1 and PTBP2 , 2011, Nature Neuroscience.

[5]  J. Tapia,et al.  hnRNP U protein is required for normal pre-mRNA splicing and postnatal heart development and function , 2015, Proceedings of the National Academy of Sciences.

[6]  Masanori Arita,et al.  Automated classification of alternative splicing and transcriptional initiation and construction of visual database of classified patterns , 2006, Bioinform..

[7]  Gene W. Yeo,et al.  Rbfox proteins regulate alternative mRNA splicing through evolutionarily conserved RNA bridges , 2013, Nature Structural &Molecular Biology.

[8]  Chadwick M. Hales,et al.  U1 small nuclear ribonucleoprotein complex and RNA splicing alterations in Alzheimer’s disease , 2013, Proceedings of the National Academy of Sciences.

[9]  Leslie C. Griffith.  Calcium/Calmodulin-Dependent Protein Kinase II: An Unforgettable Kinase , 2004, The Journal of Neuroscience.

[10]  Thomas R. Gingeras,et al.  STAR: ultrafast universal RNA-seq aligner , 2013, Bioinform..

[11]  T. Tsunoda,et al.  Transcriptome analysis of distinct mouse strains reveals kinesin light chain-1 splicing as an amyloid-β accumulation modifier , 2014, Proceedings of the National Academy of Sciences.

[12]  M. Kirschner,et al.  Phosphorylation of microtubule‐associated protein tau: identification of the site for Ca2(+)‐calmodulin dependent kinase and relationship with tau phosphorylation in Alzheimer tangles. , 1990, The EMBO journal.

[13]  Roderic Guigó,et al.  Intron-centric estimation of alternative splicing from RNA-seq data , 2012, Bioinform..

[14]  B. Frey,et al.  Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing , 2008, Nature Genetics.

[15]  Poethig Rs,et al.  Life with 25,000 genes. , 2001 .

[16]  B. Frey,et al.  The human splicing code reveals new insights into the genetic determinants of disease , 2015, Science.

[17]  Xinchen Wang,et al.  Tissue-specific alternative splicing remodels protein-protein interaction networks. , 2012, Molecular cell.

[18]  Thomas M. Keane,et al.  Mouse genomic variation and its effect on phenotypes and gene regulation , 2011, Nature.

[19]  A. Newman,et al.  RNA splicing , 1998, Current Biology.

[20]  M. Hughes,et al.  A circadian gene expression atlas in mammals: Implications for biology and medicine , 2014, Proceedings of the National Academy of Sciences.

[21]  Tom Maniatis,et al.  A role for exon sequences and splice-site proximity in splice-site selection , 1986, Cell.

[22]  Juw Won Park,et al.  MATS: a Bayesian framework for flexible detection of differential alternative splicing from RNA-Seq data , 2012, Nucleic acids research.

[23]  Lili Wan,et al.  RNA and Disease , 2009, Cell.

[24]  Eric T. Wang,et al.  Alternative Isoform Regulation in Human Tissue Transcriptomes , 2008, Nature.

[25]  Xiang-Dong Fu,et al.  Global analysis of physical and functional RNA targets of hnRNP L reveals distinct sequence and epigenetic features of repressed and enhanced exons , 2015, RNA.

[26]  H. Schulman,et al.  The multifunctional calcium/calmodulin-dependent protein kinase: from form to function. , 1995, Annual review of physiology.

[27]  Gael P. Alamancos,et al.  Methods to study splicing from high-throughput RNA sequencing data. , 2013, Methods in molecular biology.

[28]  Haixu Tang,et al.  Splicing graphs and EST assembly problem , 2002, ISMB.

[29]  Tyson A. Clark,et al.  Ultraconserved elements are associated with homeostatic control of splicing regulators by alternative splicing and nonsense-mediated decay. , 2007, Genes & development.

[30]  K. J. Hertel,et al.  Spliceosomal Pre-mRNA Splicing , 2014, Methods in Molecular Biology.

[31]  A. Weiss,et al.  A Model System for Activation-Induced Alternative Splicing of CD45 Pre-mRNA in T Cells Implicates Protein Kinase C and Ras , 2000, Molecular and Cellular Biology.

[32]  Derek Y. Chiang,et al.  DiffSplice: the genome-wide detection of differential splicing events with RNA-seq , 2012, Nucleic acids research.

[33]  B. Frey,et al.  Widespread intron retention in mammals functionally tunes transcriptomes , 2014, Genome research.

[34]  V. Vingtdeux,et al.  Calcium signaling in neurodegeneration , 2009, Molecular Neurodegeneration.

[35]  Sylvain Foissac,et al.  A General Definition and Nomenclature for Alternative Splicing Events , 2008, PLoS Comput. Biol..

[36]  Brad T. Sherman,et al.  Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources , 2008, Nature Protocols.

[37]  Peter J. Bickel,et al.  Measuring reproducibility of high-throughput experiments , 2011, 1110.4705.

[38]  P. Sharp,et al.  Rbfox2 controls autoregulation in RNA-binding protein networks , 2014, Genes & development.

[39]  Brendan J. Frey,et al.  Deciphering the splicing code , 2010, Nature.

[40]  D. Haussler,et al.  Evolutionarily conserved elements in vertebrate, insect, worm, and yeast genomes. , 2005, Genome research.

[41]  Jie Wu,et al.  SpliceTrap: a method to quantify alternative splicing under single cellular conditions , 2011, Bioinform..

[42]  T. Maniatis,et al.  The MicroRNA miR-124 promotes neuronal differentiation by triggering brain-specific alternative pre-mRNA splicing. , 2007, Molecular cell.

[43]  Christopher A. Yarosh,et al.  Alternative splicing networks regulated by signaling in human T cells. , 2012, RNA.

[44]  David G Hendrickson,et al.  Differential analysis of gene regulation at transcript resolution with RNA-seq , 2012, Nature Biotechnology.

[45]  K. Lynch,et al.  A conserved signal-responsive sequence mediates activation-induced alternative splicing of CD45. , 2003, Molecular cell.

[46]  Sebastien M. Weyn-Vanhentenryck,et al.  Systematic discovery of regulated and conserved alternative exons in the mammalian brain reveals NMD modulating chromatin regulators , 2015, Proceedings of the National Academy of Sciences.

[47]  C. Burge,et al.  Evolutionary Dynamics of Gene and Isoform Regulation in Mammalian Tissues , 2012, Science.

[48]  C. Gooding,et al.  Autoregulation of polypyrimidine tract binding protein by alternative splicing leading to nonsense-mediated decay. , 2004, Molecular cell.

[49]  Eric T. Wang,et al.  Alternative splicing regulates vesicular trafficking genes in cardiomyocytes during postnatal heart development , 2014, Nature Communications.

[50]  M. J. Mallory,et al.  Transcriptome-Wide RNA Interaction Profiling Reveals Physical and Functional Targets of hnRNP L in Human T Cells , 2013, Molecular and Cellular Biology.

[51]  Sandrine Dudoit,et al.  GC-Content Normalization for RNA-Seq Data , 2011, BMC Bioinformatics.

[52]  Lan Lin,et al.  rMATS: Robust and flexible detection of differential alternative splicing from replicate RNA-Seq data , 2014, Proceedings of the National Academy of Sciences.

[53]  Masato Yano,et al.  Ptbp2 represses adult-specific splicing to regulate the generation of neuronal precursors in the embryonic brain. , 2012, Genes & development.

[54]  D. Black,et al.  The splicing regulator PTBP2 controls a program of embryonic splicing required for neuronal maturation , 2014, eLife.

[55]  Michael D. Wilson,et al.  The Evolutionary Landscape of Alternative Splicing in Vertebrate Species , 2012, Science.

[56]  E. Wang,et al.  Analysis and design of RNA sequencing experiments for identifying isoform regulation , 2010, Nature Methods.

[57]  Weijun Gao,et al.  AVISPA: a web tool for the prediction and analysis of alternative splicing , 2013, Genome Biology.

[58]  Xiang-Dong Fu,et al.  ASF/SF2-Regulated CaMKIIδ Alternative Splicing Temporally Reprograms Excitation-Contraction Coupling in Cardiac Muscle , 2005, Cell.

[59]  Cole Trapnell,et al.  Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. , 2010, Nature biotechnology.

[60]  S. Bergmann,et al.  The evolution of gene expression levels in mammalian organs , 2011, Nature.