Web-accessible application for identifying pathogenic transcripts with RNA-seq: Increased sensitivity in diagnosis of...

Area of Science:

• Genomics
• Molecular Biology
• Neuroscience

Background:

• Molecular diagnosis is crucial for neurodevelopmental disorders (NDDs) management and prognosis.
• Standard DNA tests often fail to identify genetic causes in NDDs.
• Transcriptome analysis, specifically RNA sequencing (RNA-seq), offers potential for improved diagnostic yield.

Purpose of the Study:

• To evaluate the diagnostic utility of RNA sequencing (RNA-seq) in individuals with NDDs.
• To assess RNA-seq's ability to detect gene expression outliers and splicing abnormalities.
• To develop and validate an accessible analysis pipeline for routine NDD diagnostics.

Main Methods:

• RNA sequencing (RNA-seq) performed on cultured skin fibroblasts from 96 individuals (67 with NDDs).
• Utilized modified OUTRIDER Z scores for detecting exonic and intronic expression outliers and mis-splicing.
• Employed cycloheximide treatment to enhance detection of aberrant splicing events.
• Developed a user-friendly web application for data analysis and candidate variant confirmation.

Main Results:

• RNA-seq increased the diagnostic yield for NDDs by 13%, identifying various pathogenic transcriptional events.
• Cycloheximide treatment and Z score analysis improved the detection and resolution of aberrant splicing.
• Identified pathogenic splicing in a neuronal-expressed gene, leading to a molecular diagnosis for one individual.
• The analysis pipeline successfully ranked pathogenic transcripts as top candidates.

Conclusions:

• RNA sequencing (RNA-seq) is a valuable complementary tool for genomic diagnosis in neurodevelopmental disorders (NDDs).
• The developed transcriptome analysis approach, with enhanced sensitivity and accessibility, facilitates wider implementation in routine diagnostics.
• Customizable RNA-seq analysis is essential for maximizing diagnostic potential in NDDs.