GESS

GESS identifies exon-skipping events and determines dominant isoforms from raw RNA-seq reads without requiring prior gene annotations.

Key Features:

• De novo detection: Detects exon-skipping sites and both inclusion and skipping isoforms directly from raw RNA-seq reads without using existing gene annotations.
• Isoform dominance determination: Determines the dominant isoform produced at each exon-skipping site.
• Graph-based approach: Employs a graph-based methodology to map and analyze RNA-seq data and identify complex exon-skipping patterns.
• Integration with genomic data: Integrates sequencing-based genomic data to analyze effects of splicing activities, transcription factors (TFs), and epigenetic histone modifications on splicing outcomes.
• Splice-site strength analysis: Uses MaxEntScan-calculated splice site strength to compare skipping-isoform-dominated groups (SIDG) and inclusion-isoform-dominated groups (IIDG) around middle exons.
• Splicing factor positioning: Identifies positional preference and enrichment of splicing factors at intronic splice sites adjacent to middle exons.
• Epigenetic influence analysis: Assesses how epigenetic histone modifications can create variable barriers at exon-intron boundaries that affect skipping events.
• Experimental validation: Produces predictions that have been validated experimentally by RT-PCR.

Scientific Applications:

• ENCODE cell-line analysis: Applied to publicly available RNA-seq datasets from GM12878 and K562 cells (ENCODE) for large-scale alternative splicing analysis.
• Validation of computational predictions: Supporting RT-PCR validation of predicted exon-skipping events to confirm biological relevance.
• Regulatory mechanism investigation: Used to explore how splicing activities, TFs, and epigenetic histone modifications influence exon-skipping outcomes.
• Splice-site strength comparisons: Enabled discovery that SIDG generally exhibit weaker MaxEntScan-calculated splice site strength around middle exons compared to IIDG.
• Splicing factor mapping: Revealed enrichment patterns of splicing factors at intronic splice sites adjacent to middle exons.
• Epigenetic impact studies: Facilitated analysis showing that different epigenetic modifications affect the establishment of exon-intron boundaries and skipping events.

Methodology:

Graph-based mapping and analysis of raw RNA-seq reads to detect exon-skipping sites and determine dominant isoforms without gene annotations; splice-site strength evaluation using MaxEntScan; and integration with sequencing-based genomic data for splicing activities, transcription factors (TFs), and epigenetic histone modifications.