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Sequence based analysis of neurodevelopmental disorders
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis the main focus is the use of methods and applications of next generation sequencing in order to study three of the most common neurodevelopmental disorders: intellectual disability, epilepsy and schizophrenia. A large fraction of the genes in our genome produce several distinct transcript isoforms through the process of splicing and there is an increasing amount of evidence pinpointing mutations affecting splicing as a mechanism of disease.  In Paper I we used exome capture of RNA in combination with sequencing in order to enrich for coding sequences. We show that this approach enables us to detect lowly expressed transcript and splice events that would have been missed in regular RNA sequencing using the same coverage.  In Paper II we selectively depleted the different transcripts of Quaking (QKI), a gene previously associated to schizophrenia. Using RNA sequencing we show that the effects of depletion differ between transcripts and that the QKI gene is a potential regulator of the Glial Fibrillary Acidic Protein (GFAP), a gene implicated in several diseases in the central nervous system.

De-novo mutations are frequently reported to be causative in neurodevelopmental disorders with a strong genetic component, such as epilepsy and intellectual disability. In Paper III we used exome sequencing in family trios where the child was diagnosed with both intellectual disability and epilepsy, focusing on finding de-novo mutations. We identified several previously unknown disease causing mutations in genes previously known to cause disease and used previously published interaction and mutation data to prioritize novel candidate genes. The most interesting result from this study are the implication of the HECW2 gene as a candidate gene in intellectual disability and epilepsy. In Paper IV we used RNA sequencing of post mortem brain tissue in a large cohort of schizophrenics and controls.  In this study we could show that the immune system and more specifically the complement system was dysregulated in a large fraction of patients. Further, using co-expression network we also found some evidence suggesting genes involved in axon development and maintenance.

 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. , 62 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1231
National Category
Medical Genetics
Research subject
Medical Genetics
Identifiers
URN: urn:nbn:se:uu:diva-287407ISBN: 978-91-554-9597-8 (print)OAI: oai:DiVA.org:uu-287407DiVA: diva2:922752
Public defence
2016-06-14, C2:305, BMC, Husargatan 3, Uppsala, 09:30 (English)
Opponent
Supervisors
Available from: 2016-05-20 Created: 2016-04-24 Last updated: 2016-06-15
List of papers
1. Exome RNA sequencing reveals rare and novel alternative transcripts
Open this publication in new window or tab >>Exome RNA sequencing reveals rare and novel alternative transcripts
2013 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 41, no 1, e6- p.Article in journal (Refereed) Published
Abstract [en]

RNA sequencing has become an important method to perform hypothesis-free characterization of global gene expression. One of the limitations of RNA sequencing is that most sequence reads represent highly expressed transcripts, whereas low level transcripts are challenging to detect. To combine the benefits of traditional expression arrays with the advantages of RNA sequencing, we have used whole exome enrichment prior to sequencing of total RNA. We show that whole exome capture can be successfully applied to cDNA to study the transcriptional landscape in human tissues. By introducing the exome enrichment step, we are able to identify transcripts present at very low levels, which are below the level of detection in conventional RNA sequencing. Although the enrichment increases the ability to detect presence of transcripts, it also lowers the accuracy of quantification of expression levels. Our results yield a large number of novel exons and splice isoforms, suggesting that conventional RNA sequencing methods only detect a small fraction of the full transcript diversity. We propose that whole exome enrichment of RNA is a suitable strategy for genome-wide discovery of novel transcripts, alternative splice variants and fusion genes.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-180350 (URN)10.1093/nar/gks816 (DOI)000312889900006 ()22941640 (PubMedID)
Available from: 2012-09-04 Created: 2012-09-04 Last updated: 2017-12-07Bibliographically approved
2. RNA-binding protein QKI regulates Glial fibrillary acidic protein expression in human astrocytes
Open this publication in new window or tab >>RNA-binding protein QKI regulates Glial fibrillary acidic protein expression in human astrocytes
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2013 (English)In: Human Molecular Genetics, ISSN 0964-6906, E-ISSN 1460-2083, Vol. 22, no 7, 1373-1382 p.Article in journal (Refereed) Published
Abstract [en]

Linkage, association and expression studies previously pointed to the human QKI, KH domain containing, RNA-binding (QKI) as a candidate gene for schizophrenia. Functional studies of the mouse orthologue Qk focused mainly on its role in oligodendrocyte development and myelination, while its function in astroglia remained unexplored. Here, we show that QKI is highly expressed in human primary astrocytes and that its splice forms encode proteins targeting different subcellular localizations. Uncovering the role of QKI in astrocytes is of interest in light of growing evidence implicating astrocyte dysfunction in the pathogenesis of several disorders of the central nervous system. We selectively silenced QKI splice variants in human primary astrocytes and used RNA sequencing to identify differential expression and splice variant composition at the genome-wide level. We found that an mRNA expression of Glial fibrillary acidic protein (GFAP), encoding a major component of astrocyte intermediate filaments, was down-regulated after QKI7 splice variant silencing. Moreover, we identified a potential QKI-binding site within the 3 untranslated region of human GFAP. This sequence was not conserved between mice and humans, raising the possibility that GFAP is a target for QKI in humans but not rodents. Haloperidol treatment of primary astrocytes resulted in coordinated increases in QKI7 and GFAP expression. Taken together, our results provide the first link between QKI and GFAP, two genes with alterations previously observed independently in schizophrenic patients. Our findings for QKI, together with its well-known role in myelination, suggest that QKI is a hub regulator of glia function in humans.

National Category
Natural Sciences Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-198376 (URN)10.1093/hmg/dds553 (DOI)000316297000009 ()
Available from: 2013-04-15 Created: 2013-04-15 Last updated: 2017-12-06Bibliographically approved
3. Identification of new candidate genes in intellectual disability and epilepsy
Open this publication in new window or tab >>Identification of new candidate genes in intellectual disability and epilepsy
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(English)Manuscript (preprint) (Other academic)
National Category
Basic Medicine
Identifiers
urn:nbn:se:uu:diva-287406 (URN)
Available from: 2016-04-24 Created: 2016-04-24 Last updated: 2016-06-15
4. Transcriptome sequencing implicate alterations of complement factor levels in schizophrenia.
Open this publication in new window or tab >>Transcriptome sequencing implicate alterations of complement factor levels in schizophrenia.
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(English)Manuscript (preprint) (Other academic)
National Category
Basic Medicine
Research subject
Medical Science
Identifiers
urn:nbn:se:uu:diva-287405 (URN)
Available from: 2016-04-24 Created: 2016-04-24 Last updated: 2016-06-15

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