SpliceTools – a suite of downstream RNA splicing analysis tools to investigate mechanisms and impact of alternative splicing

As a fundamental aspect of normal cell signaling and disease states, there is great interest in determining alternative splicing (AS) changes in physiologic, pathologic, and pharmacologic settings. High throughput RNA sequencing and specialized software to detect AS has greatly enhanced our ability to determine transcriptome-wide splicing changes. Despite the richness of this data, deriving meaning from sometimes thousands of AS events is a substantial bottleneck for most investigators.

Tulane University researchers have developed SpliceTools, a suite of data processing modules that arms investigators with the ability to quickly produce summary statistics, mechanistic insights, and functional significance of AS changes through command line or through an online user interface. Utilizing RNA-seq datasets for 186 RNA binding protein knockdowns, nonsense mediated RNA decay inhibition, and pharmacologic splicing inhibition, the researchers illustrate the utility of SpliceTools to distinguish splicing disruption from regulated transcript isoform changes, they show the broad transcriptome footprint of the pharmacologic splicing inhibitor, indisulam, they illustrate the utility in uncovering mechanistic underpinnings of splicing inhibition, they identify predicted neo-epitopes in pharmacologic splicing inhibition, and they show the impact of splicing alterations induced by indisulam on cell cycle progression. Together, SpliceTools puts rapid and easy downstream analysis at the fingertips of any investigator studying AS.

SpliceTools schematic

(A) The six core skipped exon analysis tools are shown with their respective outputs indicated to the right. Tools can either be used individually or all six can be called using one command with SEMedley. (B) Listing of core tools for retained intron analysis and their outputs are indicated. RIMedley can be used to run all three retained intron tools using a single command. (C) SpliceCompare analyzes any combination of A5SS, A3SS, MXE, RI, or SE files for comparative analyses of AS profiles across datasets. A parallel processing version of SpliceCompare is provided that utilizes 5 processors to simultaneously analyze A5SS, A3SS, MXE, RI and SE files if the user has installed the Parallel::Forkmanager module.

Availability – SpliceTools is available through an online web interface (splicetools.org) or for download at (https://github.com/flemingtonlab/SpliceTools).