extractSplicingEnrichmentComparison: Compare enrichment of specific type of alternative splicing...

In IsoformSwitchAnalyzeR: Identify, Annotate and Visualize Alternative Splicing and Isoform Switches with Functional Consequences from both short- and long-read RNA-seq data.

Description

This function compares the enrichment of alternative splicing (f.x. exon skipping) between two comparisons (ctrl vs ko1 compared to ctrl vs ko2) and reports whether there is a significant difference between the comparisons. It other words it compares the output of extractSplicingEnrichment.

Usage

extractSplicingEnrichmentComparison(
    switchAnalyzeRlist,
    splicingToAnalyze = 'all',
    alpha = 0.05,
    dIFcutoff = 0.1,
    onlySigIsoforms = FALSE,
    countGenes = TRUE,
    plot = TRUE,
    localTheme = theme_bw(base_size = 14),
    minEventsForPlotting = 10,
    returnResult=TRUE
)

Arguments

switchAnalyzeRlist	A switchAnalyzeRlist object where analyzeSwitchConsequences() have been run to identify consequences of isoform switches
splicingToAnalyze	A string indicating which consequences should be considered. See details for description. Default is all.
alpha	The cutoff which the FDR correct p-values must be smaller than for calling significant switches. Default is 0.05.
dIFcutoff	The cutoff which the changes in (absolute) isoform usage must be larger than before an isoform is considered switching. This cutoff can remove cases where isoforms with (very) low dIF values are deemed significant and thereby included in the downstream analysis. This cutoff is analogous to having a cutoff on log2 fold change in a normal differential expression analysis of genes to ensure the genes have a certain effect size. Default is 0.1 (10%).
onlySigIsoforms	A logic indicating whether to only consider significant isoforms, meaning only analyzing genes where at least two isoforms which both have significant usage changes in opposite direction (quite strict) Naturally this only works if the isoform switch test used have isoform resolution (which the build in isoformSwitchTestDEXSeq has). If FALSE all isoforms with an absolute dIF value larger than dIFcutoff in a gene with significant switches (defined by alpha and dIFcutoff) are included in the pairwise comparison. Default is FALSE (non significant isoforms are also considered based on the logic that if one isoform changes it contribution - there must be an equivalent opposite change in usage in the other isoforms from that gene).
countGenes	A logic indicating whether it is the number of genes (if TRUE) or isoform switches (if FALSE) which primary result in gain/loss that are counted. Default is TRUE.
plot	A logic indicting whether the analysis should be plotted. If TRUE and returnResult = FALSE the ggplot2 object will be returned instead. Default is TRUE.
localTheme	General ggplo2 theme with which the plot is made, see ?ggplot2::theme for more info. Default is theme_bw(base_size = 14).
minEventsForPlotting	The minimum number of events (total gain/loss) must be present before the result is visualized. Default is 10.
returnResult	A logic indicating whether the analysis should be returned as a data.frame. If FALSE (and plot=TRUE) the ggplot2 object will be returned instead. Default is FLASE.

Details

The classification of alternative splicing is always compared to the hypothetical pre-mRNA constructed by concatenating all exons from isoforms of the same gene.

The alternative splicing types, which can be passed to splicingToAnalyze must be a combination of:

• all : All of the alternative splicing types indicated below.

• IR : Intron Retention.

• A5 : Alternative 5' donor site (changes in the 5'end of the upstream exon).

• A3 : Alternative 3' acceptor site (changes in the 3'end of the downstream exon).

• ATSS : Alternative Transcription Start Site.

• ATTS : Alternative Transcription Termination Site.

• ES : Exon Skipping.

• MES : Multiple Exon Skipping. Skipping of >1 consecutive exons.

• MEE : Mutually Exclusive Exons.

For details of how to interpret the splice events see the details section of analyzeAlternativeSplicing.

The significance test is performed with R's build in fisher.test() with default parameters and resulting p-values are corrected via p.adjust() using FDR (Benjamini-Hochberg).

Value

If returnResult=TRUE a data.frame with the statistical summary for each opposing consequences in each comparison. If plot=TRUE a plot summarizing the proportions is also created of switches with specific consequences is created.

Author(s)

Kristoffer Vitting-Seerup

References

• Vitting-Seerup et al. The Landscape of Isoform Switches in Human Cancers. Mol. Cancer Res. (2017).

• Vitting-Seerup et al. IsoformSwitchAnalyzeR: Analysis of changes in genome-wide patterns of alternative splicing and its functional consequences. Bioinformatics (2019).

See Also

isoformSwitchTestDEXSeq
isoformSwitchTestDRIMSeq
analyzeAlternativeSplicing
extractSplicingSummary
extractSplicingEnrichment
extractSplicingGenomeWide

Examples

### Load example data
data("exampleSwitchListAnalyzed")

extractSplicingEnrichmentComparison( exampleSwitchListAnalyzed )

IsoformSwitchAnalyzeR documentation built on Nov. 8, 2020, 5:36 p.m.