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Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi. Uppsala universitet, Science for Life Laboratory, SciLifeLab.ORCID-id: 0000-0001-6085-6749
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Genomik. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för immunologi, genetik och patologi, Genomik. Uppsala universitet, Science for Life Laboratory, SciLifeLab.
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2011 (Engelska)Ingår i: Nature Structural & Molecular Biology, ISSN 1545-9993, E-ISSN 1545-9985, Vol. 18, nr 12, s. 1435-1440Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Transcriptome sequencing allows for analysis of mature RNAs at base pair resolution. Here we show that RNA-seq can also be used for studying nascent RNAs undergoing transcription. We sequenced total RNA from human brain and liver and found a large fraction of reads (up to 40%) within introns. Intronic RNAs were abundant in brain tissue, particularly for genes involved in axonal growth and synaptic transmission. Moreover, we detected significant differences in intronic RNA levels between fetal and adult brains. We show that the pattern of intronic sequence read coverage is explained by nascent transcription in combination with co-transcriptional splicing. Further analysis of co-transcriptional splicing indicates a correlation between slowly removed introns and alternative splicing. Our data show that sequencing of total RNA provides unique insight into the transcriptional processes in the cell, with particular importance for normal brain development.

Ort, förlag, år, upplaga, sidor
2011. Vol. 18, nr 12, s. 1435-1440
Nationell ämneskategori
Medicin och hälsovetenskap
Identifikatorer
URN: urn:nbn:se:uu:diva-166861DOI: 10.1038/nsmb.2143ISI: 000298011600036OAI: oai:DiVA.org:uu-166861DiVA, id: diva2:478253
Anmärkning

De två sista författarna delar sistaförfattarskapet

Tillgänglig från: 2012-01-16 Skapad: 2012-01-16 Senast uppdaterad: 2017-12-08Bibliografiskt granskad
Ingår i avhandling
1. Genome-wide Characterization of RNA Expression and Processing
Öppna denna publikation i ny flik eller fönster >>Genome-wide Characterization of RNA Expression and Processing
2013 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

The production of fully mature protein-coding transcripts is an intricate process that involves numerous regulation steps. The complexity of these steps provides the means for multilayered control of gene expression. Comprehensive understanding of gene expression regulation is essential for interpreting the role of gene expression programs in tissue specificity, development and disease. In this thesis, we aim to provide a better global view of the human transcriptome, focusing on its content, synthesis, processing and regulation using next-generation sequencing as a read-out.

In Paper I, we show that sequencing of total RNA provides unique insights into RNA processing. Our results revealed that co-transcriptional splicing is a widespread mechanism in human and chimpanzee brain tissues. We also found a correlation between slowly removed introns and alternative splicing. In Paper II, we explore the benefits of exome capture approaches in combination with RNA-sequencing to detect transcripts expressed at low-levels. Based on our results, we demonstrate that this approach increases the sensitivity for detecting low level transcripts and leads to the identification of novel exons and splice isoforms. In Paper III, we highlight the advantages of performing RNA-sequencing on separate cytoplasmic and nuclear RNA fractions. In comparison with conventional poly(A) RNA, cytoplasmic RNA contained a significantly higher fraction of exonic sequence, providing increased sensitivity for splice junction detection and for improved de novo assembly. Conversely, the nuclear fraction showed an enrichment of unprocessed RNA compared to when sequencing total RNA, making it suitable for analysis of RNA processing dynamics. In Paper IV, we used exome sequencing to sequence the DNA of a patient with unexplained intellectual disability and identified a de novo mutation in BAZ1A, which encodes the chromatin-remodeling factor ACF1. Functional studies indicated that the mutation influences the expression of genes involved in extracellular matrix organization, synaptic function and vitamin D3 metabolism. The differential expression of CYP24A, SYNGAP1 and COL1A2 correlated with the patient’s clinical diagnosis.

The findings presented in this thesis contribute towards an improved understanding of the human transcriptome in health and disease, and highlight the advantages of developing novel methods to obtain global and comprehensive views of the transcriptome.

Ort, förlag, år, upplaga, sidor
Uppsala: Acta Universitatis Upsaliensis, 2013. s. 61
Serie
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 939
Nyckelord
RNA sequencing, RNA splicing, RNA processing, Gene expression
Nationell ämneskategori
Medicinsk genetik
Forskningsämne
Molekylär genetik
Identifikatorer
urn:nbn:se:uu:diva-209390 (URN)978-91-554-8784-3 (ISBN)
Disputation
2013-11-29, Rudbeck Salen, Dag Hammarskjölds väg 20,, Uppsala, 09:15 (Engelska)
Opponent
Handledare
Tillgänglig från: 2013-11-08 Skapad: 2013-10-18 Senast uppdaterad: 2018-01-11

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Ameur, AdamZaghlool, AmmarHalvardson, JonatanGyllensten, UlfCavelier, LuciaFeuk, Lars

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