EventPointer

EventPointer identifies and characterizes alternative splicing events from Human Transcriptome Array 2.0 (HTA 2.0) data to quantify splicing changes and assess their potential effects on encoded protein domains.

Key Features:

• R package implementation: Implemented as an R package within the aroma.affymetrix framework for analysis of HTA 2.0 array data.
• Linear-model framework: Employs linear models that accommodate complex experimental designs including time-course and paired-sample studies.
• Joint exon and junction analysis: Jointly analyzes probes for exons and junctions involved in splicing events to improve detection and characterization.
• Input data and matrices: Processes CEL files together with design and contrast matrices as analysis inputs.
• Event-level outputs: Reports splicing event type (e.g., exon cassette, alternative 3' splice site), fold change, and statistical significance for each detected event.
• Protein-domain impact: Identifies potential protein domains affected by alternative splicing and provides statistical significance for domain enrichment.
• Performance metrics: Delivers high sensitivity and specificity with a reported very low false discovery rate (one false positive among the top 200 detected events in testing).
• Visualization and validation files: Generates files compatible with the Integrative Genomics Viewer (IGV) and produces outputs to support PCR primer design for validation.

Scientific Applications:

• Alternative splicing research: Detection and quantification of alternative splicing events for studies of transcriptome variability and regulatory mechanisms.
• Disease-focused studies: Analysis of splicing alterations implicated in pathological conditions to prioritize events for biological follow-up.
• Array-based alternative to RNA-seq: Use of HTA 2.0 array data as a viable approach to detect alternative splicing events when RNA-seq is not employed.
• Complex experimental designs: Analysis of experiments with time-course, paired samples, or other multifactor designs enabled by the linear-model framework.
• Validation prioritization: Prioritization of candidate splicing events for experimental validation using event-level fold changes, p-values, and protein domain enrichment.

Methodology:

Implemented as an R package within aroma.affymetrix that processes CEL files with design and contrast matrices and applies linear models to jointly analyze exon and junction probe signals to detect and quantify alternative splicing events.