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Loss of Function variants and Transcription

Loss of Function variants and Transcription. Manuel Rivas, University of Oxford Geuvadis RNA Sequencing Analysis Meeting 15/04/2012. Loss of Function variants. (Putative) LoF variants: Splice site variants Nonsense and Readthrough mutations Frameshift indels .

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Loss of Function variants and Transcription

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  1. Loss of Function variants and Transcription Manuel Rivas, University of Oxford Geuvadis RNA Sequencing Analysis Meeting 15/04/2012

  2. Loss of Function variants • (Putative) LoF variants: • Splice site variants • Nonsense and Readthrough mutations • Frameshiftindels. • Exon Deletion and Duplication • Gene Deletion and Duplication • Gene Fusion* 5’ AG GT[AG]AG AG G 3’

  3. What can we learn about LoF using RNA sequencing data • Effect of splice variants on transcripts • Exon Skipping • Intron Retention • mRNA Surveillance Pathways • Escape NMD • Trigger NMD What are the sequence properties that trigger NMD pathway or dictate escaping NMD pathway. 3. NMD efficiency in individuals. Do rates vary between individuals? • CNVs • Gene Fusion – impact of gene fusions in transcription? Do they properly get transcribed? • Exon/Gene Deletions and Duplications – impact of exon/gene deletions and duplications in transcription?

  4. Splice Site Variants Initial analysis of Exon Quantification Data using splice variant annotation successfully recapitulates splicing event. (IFIH1 Exon Skipping c.IVS14+1G>A) G ` ` ` A ` ` ` Annotation -> Evaluate Empirical Effect on Transcript

  5. mRNA surveillance – Nonsense Mediated Decay (NMD) IFIH1 Transcript Escapes Nonsense Mediated Decay. • Model 1: NMD is not triggered • Model 2: NMD is triggered G ` ` ` A ` ` ` Propose Statistic that detects Nonsense Mediated Decay EXAMPLE 1: ESCAPE NMD

  6. mRNA surveillance – Nonsense Mediated Decay (NMD) CIAO1 premature stop variant triggers Nonsense Mediated Decay. • Model 1: NMD is not triggered • Model 2: NMD is triggered Propose Statistic that detects Nonsense Mediated Decay EXAMPLE 2: TRIGGER NMD REDUCTION OBSERVED THROUGHOUT TRANSCRIPT

  7. Loss of Function variants and ASE Added Annotation Layer to VAT output from annotation program co-developed at Oxford. Fraction of LoF observations with allele specific expression detected. Crude measure at the moment based on Tuuli’s first run (n = 180 samples). Determines a two-sided confidence interval from the beta posterior.

  8. Loss of Function variants and eQTL Analysis • Dependent on frequency range. • ASE most powerful for rare variants. • eQTL suitable for common variants.

  9. Methods and Analytical Strategies for LoF variants • Splicing Effects based on Annotation • Exon Skipping • Intron Retention • mRNA Surveillance Pathways • Escape NMD • Trigger NMD • CNVs • Gene Fusion • Exon/Gene Deletions and Duplications

  10. Splice Site Variants • (Putative) LoF variants: • Splice site variants • Nonsense and Readthrough mutations • Frameshiftindels. • Exon Deletion and Duplication • Gene Deletion and Duplication • Gene Fusions* Identify additional splice variants based on transcript assembly M. Sammeth and other groups.

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