title: Global transcript structure resolution of high gene density genomes through multi-platform data integration.
abstract: Annotation of herpesvirus genomes has traditionally been undertaken through the detection of open reading frames and other genomic motifs, supplemented with sequencing of individual cDNAs. Second generation sequencing and high-density microarray studies have revealed vastly greater herpesvirus transcriptome complexity than is captured by existing annotation. The pervasive nature of overlapping transcription throughout herpesvirus genomes, however, poses substantial problems in resolving transcript structures using these methods alone. We present an approach that combines the unique attributes of Pacific Biosciences Iso-Seq long-read, Illumina short-read and deepCAGE (Cap Analysis of Gene Expression) sequencing to globally resolve polyadenylated isoform structures in replicating Epstein-Barr virus (EBV). Our method, Transcriptome Resolution through Integration of Multi-platform Data (TRIMD), identifies nearly 300 novel EBV transcripts, quadrupling the size of the annotated viral transcriptome. These findings illustrate an array of mechanisms through which EBV achieves functional diversity in its relatively small, compact genome including programmed alternative splicing (e.g. across the IR1 repeats), alternative promoter usage by LMP2 and other latency-associated transcripts, intergenic splicing at the BZLF2 locus, and antisense transcription and pervasive readthrough transcription throughout the genome.
authors.affiliation: Department of Pathology, Tulane University School of Medicine, New Orleans, LA 70112, USA.
authors.forename: Tina
authors.identifier: http://orcid.org/0000-0002-1283-0293
authors.initials: T
authors.lastname: O'Grady
authors.affiliation: Department of Pathology, Tulane University School of Medicine, New Orleans, LA 70112, USA.
authors.forename: Xia
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Microbiology and Immunology, Tulane University School of Medicine, New Orleans, LA 70112, USA.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Höner Zu Bentrup
authors.affiliation: Department of Pathology, Tulane University School of Medicine, New Orleans, LA 70112, USA.
authors.forename: Melody
authors.initials: M
authors.lastname: Baddoo
authors.affiliation: Department of Pathology, Tulane University School of Medicine, New Orleans, LA 70112, USA.
authors.forename: Monica
authors.initials: M
authors.lastname: Concha
authors.affiliation: Department of Pathology, Tulane University School of Medicine, New Orleans, LA 70112, USA Tulane Cancer Center, New Orleans, LA 70112, USA eflemin@tulane.edu.
authors.forename: Erik K
authors.initials: EK
authors.lastname: Flemington
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D002460:Cell Line; D021901:DNA, Intergenic / Q000235:genetics; D005091:Exons / Q000235:genetics; D016679:Genome, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D058977:Molecular Sequence Annotation; D026723:Polyadenylation / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D015203:Reproducibility of Results; D017423:Sequence Analysis, RNA; D013223:Statistics as Topic; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D059467:Transcriptome / Q000235:genetics
chemical_list: D021901:DNA, Intergenic; D012333:RNA, Messenger