title: PAT-seq: a method to study the integration of 3'-UTR dynamics with gene expression in the eukaryotic transcriptome.
abstract: A major objective of systems biology is to quantitatively integrate multiple parameters from genome-wide measurements. To integrate gene expression with dynamics in poly(A) tail length and adenylation site, we developed a targeted next-generation sequencing approach, Poly(A)-Test RNA-sequencing. PAT-seq returns (i) digital gene expression, (ii) polyadenylation site/s, and (iii) the polyadenylation-state within and between eukaryotic transcriptomes. PAT-seq differs from previous 3' focused RNA-seq methods in that it depends strictly on 3' adenylation within total RNA samples and that the full-native poly(A) tail is included in the sequencing libraries. Here, total RNA samples from budding yeast cells were analyzed to identify the intersect between adenylation state and gene expression in response to loss of the major cytoplasmic deadenylase Ccr4. Furthermore, concordant changes to gene expression and adenylation-state were demonstrated in the classic Crabtree-Warburg metabolic shift. Because all polyadenylated RNA is interrogated by the approach, alternative adenylation sites, noncoding RNA and RNA-decay intermediates were also identified. Most important, the PAT-seq approach uses standard sequencing procedures, supports significant multiplexing, and thus replication and rigorous statistical analyses can for the first time be brought to the measure of 3'-UTR dynamics genome wide.
authors.affiliation: Victorian Bioinformatics Consortium, Monash University, Clayton 3800, Australia Life Sciences Computation Centre, Victorian Life Sciences Computation Initiative, Carlton 3053, Australia Monash Bioinformatics Platform, Monash University, Clayton 3800, Australia.
authors.forename: Paul F
authors.initials: PF
authors.lastname: Harrison
authors.affiliation: Victorian Bioinformatics Consortium, Monash University, Clayton 3800, Australia Life Sciences Computation Centre, Victorian Life Sciences Computation Initiative, Carlton 3053, Australia Monash Bioinformatics Platform, Monash University, Clayton 3800, Australia.
authors.forename: David R
authors.initials: DR
authors.lastname: Powell
authors.affiliation: EMBL-Australia Collaborating Laboratory, Genome Biology Department, The John Curtin School of Medical Research (JCSMR), The Australian National University, Acton (Canberra) 2601, Australian Capital Territory, Australia.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Clancy
authors.affiliation: EMBL-Australia Collaborating Laboratory, Genome Biology Department, The John Curtin School of Medical Research (JCSMR), The Australian National University, Acton (Canberra) 2601, Australian Capital Territory, Australia Victor Chang Cardiac Research Institute, Darlinghurst (Sydney), New South Wales 2010, Australia.
authors.forename: Thomas
authors.initials: T
authors.lastname: Preiss
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Australia.
authors.forename: Peter R
authors.initials: PR
authors.lastname: Boag
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Australia.
authors.forename: Ana
authors.initials: A
authors.lastname: Traven
authors.affiliation: Victorian Bioinformatics Consortium, Monash University, Clayton 3800, Australia Life Sciences Computation Centre, Victorian Life Sciences Computation Initiative, Carlton 3053, Australia.
authors.forename: Torsten
authors.initials: T
authors.lastname: Seemann
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Australia.
authors.forename: Traude H
authors.initials: TH
authors.lastname: Beilharz
mesh_terms: D020413:3' Untranslated Regions; D015966:Gene Expression Regulation, Fungal; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D020871:RNA Stability; D012331:RNA, Fungal / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis*; D012260:Ribonucleases / Q000172:deficiency / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D017423:Sequence Analysis, RNA / Q000379:methods*; D059467:Transcriptome
chemical_list: D020413:3' Untranslated Regions; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; C078657:CCR4 protein, S cerevisiae; D012260:Ribonucleases
keywords: Ccr4; RNA-seq; alternative polyadenylation; ePAT; gene expression; polyadenylation; translational control