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Characterization of mRNA isoform diversity in a transgenic model of tau pathology using targeted long‐read sequencing
Author(s) -
Leung Szi Kay,
Jeffries Aaron,
Han Eilis,
Castanho Isabel,
Moore Karen,
Murray Tracey K.,
Ahmed Zeshan,
Collier David A.,
Mill Jonathan
Publication year - 2020
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1002/alz.046061
Subject(s) - alternative splicing , rna splicing , biology , gene isoform , computational biology , rna seq , transcriptome , gene , exon , entorhinal cortex , genetics , deep sequencing , rna , gene expression , genome , neuroscience , hippocampus
Background An increasing number of studies implicate a role for alternative splicing in the development and neuropathology of Alzheimer’s disease (AD). However, it has been historically challenging to characterise splicing events, due to the limitations of short‐read RNA‐sequencing (RNA‐Seq) for the capture full‐length transcripts critical for transcriptome assembly. In this study, we used Pacific Biosciences long‐read isoform sequencing (Iso‐Seq) to enrich and comprehensively characterise isoform diversity for AD‐associated genes in entorhinal cortex samples from a well‐validated AD transgenic mouse model. Method We used Pacific Biosciences Targeted Iso‐Seq to investigate splicing of 20 AD‐associated genes (including TREM2, BIN1 , and APOE ) in entorhinal cortex of a well‐characterised mouse model of tau pathology, rTg4510. Sequence data was processed using the Iso‐Seq3 pipeline, followed by downstream analysis using other publically available resources and customised scripts. The same samples have additionally been sequenced using whole transcriptome Iso‐Seq and RNA‐Seq as validation. Result We obtained deep sequencing coverage of full‐length transcripts for each of the AD genes, revealing complex usage of alternative start sites and splicing events, as well as many novel 5′starts and 3’ends not previously annotated in existing genomic datasets. We identified differential transcript expression and isoform usage between transgenic and wild‐type mice, with highly‐correlated gene expression between Iso‐Seq and short‐read RNA‐seq data. Conclusion This study highlights the application of long‐read sequencing approaches to assess splicing variation and isoform diversity in AD by selective gene enrichment. Results suggest differential splicing events associated with AD pathology, supporting a role for transcriptomic dysregulation in development of AD. Further work will be undertaken to characterise other AD‐associated genes, and to extend these analyses to human post‐mortem brain samples.