title: Circular RNA enrichment in platelets is a signature of transcriptome degradation.
abstract: In platelets, splicing and translation occur in the absence of a nucleus. However, the integrity and stability of mRNAs derived from megakaryocyte progenitor cells remain poorly quantified on a transcriptome-wide level. As circular RNAs (circRNAs) are resistant to degradation by exonucleases, their abundance relative to linear RNAs can be used as a surrogate marker for mRNA stability in the absence of transcription. Here we show that circRNAs are enriched in human platelets 17- to 188-fold relative to nucleated tissues and 14- to 26-fold relative to samples digested with RNAse R to selectively remove linear RNA. We compare RNAseq read depths inside and outside circRNAs to provide in silico evidence of transcript circularity, show that exons within circRNAs are enriched on average 12.7 times in platelets relative to nucleated tissues and identify 3162 genes significantly enriched for circRNAs, including some where all RNAseq reads appear to be derived from circular molecules. We also confirm that this is a feature of other anucleate cells through transcriptome sequencing of mature erythrocytes, demonstrate that circRNAs are not enriched in cultured megakaryocytes, and demonstrate that linear RNAs decay more rapidly than circRNAs in platelet preparations. Collectively, these results suggest that circulating platelets have lost >90% of their progenitor mRNAs and that translation in platelets occurs against the backdrop of a highly degraded transcriptome. Finally, we find that transcripts previously classified as products of reverse transcriptase template switching are both enriched in platelets and resistant to decay, countering the recent suggestion that up to 50% of rearranged RNAs are artifacts.
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: Abd A
authors.initials: AA
authors.lastname: Alhasan
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: Osagie G
authors.identifier: http://orcid.org/0000-0003-3116-2558
authors.initials: OG
authors.lastname: Izuogu
authors.affiliation: Kuwait Medical Genetics Centre, Al-Sabah Medical Area, Kuwait City, Kuwait;
authors.forename: Haya H
authors.initials: HH
authors.lastname: Al-Balool
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom; Bioinformatics Support Unit, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: Jannetta S
authors.identifier: http://orcid.org/0000-0002-0231-9897
authors.initials: JS
authors.lastname: Steyn
authors.affiliation: National Health Service Blood and Transplant, Department of Haematology, University of Cambridge, Cambridge, United Kingdom;
authors.forename: Amanda
authors.initials: A
authors.lastname: Evans
authors.affiliation: National Health Service Blood and Transplant, Department of Haematology, University of Cambridge, Cambridge, United Kingdom;
authors.forename: Maria
authors.initials: M
authors.lastname: Colzani
authors.affiliation: National Health Service Blood and Transplant, Department of Haematology, University of Cambridge, Cambridge, United Kingdom;
authors.forename: Cedric
authors.initials: C
authors.lastname: Ghevaert
authors.affiliation: Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow, United Kingdom; and Tissue and Cellular Therapies, Research Development & Innovation, Scottish National Blood Transfusion Service, Glasgow, United Kingdom.
authors.forename: Joanne C
authors.identifier: http://orcid.org/0000-0003-2142-6572
authors.initials: JC
authors.lastname: Mountford
authors.affiliation: Tissue and Cellular Therapies, Research Development & Innovation, Scottish National Blood Transfusion Service, Glasgow, United Kingdom.
authors.forename: Lamin
authors.initials: L
authors.lastname: Marenah
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: David J
authors.initials: DJ
authors.lastname: Elliott
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: Mauro
authors.initials: M
authors.lastname: Santibanez-Koref
authors.affiliation: Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;
authors.forename: Michael S
authors.initials: MS
authors.lastname: Jackson
mesh_terms: D001792:Blood Platelets / Q000378:metabolism*; D005091:Exons / Q000235:genetics; D005095:Exoribonucleases / Q000378:metabolism; D006801:Humans; D008533:Megakaryocytes / Q000378:metabolism; D012313:RNA / Q000235:genetics*; D020871:RNA Stability / Q000235:genetics*; D000079962:RNA, Circular; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D059467:Transcriptome / Q000235:genetics*
chemical_list: D000079962:RNA, Circular; D012313:RNA; D005095:Exoribonucleases; C080868:ribonuclease R