Study of complex structural variations of X-linked deafness-2 based on single-molecule sequencing

Area of Science:

• Genetics
• Otolaryngology
• Molecular Biology

Background:

• X-linked deafness-2 (DFNX2) is a form of inner ear malformation, specifically cochlear incomplete partition type III (IP-III).
• The POU3F4 gene is identified as the causative gene for DFNX2.
• DFNX2 is characterized by specific anatomical abnormalities and potential complications during otologic surgery.

Purpose of the Study:

• To investigate the genetic causes of DFNX2 in 12 unrelated patients.
• To compare the diagnostic efficiency of targeted next-generation sequencing (NGS) and long-read sequencing for DFNX2.
• To identify novel genetic variations, including structural variations, associated with DFNX2.

Main Methods:

• Targeted next-generation sequencing (NGS) was employed to analyze the POU3F4 gene in 12 patients.
• Long-read sequencing was utilized for patients with negative NGS results to detect structural variations.
• Analysis of deletion (DEL) and inversion (INV) mechanisms, including non-homologous end joining (NHEJ) and non-allelic homologous recombination (NAHR).

Main Results:

• NGS identified six POU3F4 variants in the cohort.
• Long-read sequencing detected significant structural variations (870-kb deletion and 8-Mb inversion) upstream of POU3F4 in two patients initially negative by NGS.
• Common POU3F4 mutations include point mutations, insertions/deletions (INDELs), and exon mutations.

Conclusions:

• NGS is effective for detecting common POU3F4 mutations in DFNX2.
• Single-molecule long-read sequencing is crucial for identifying pathogenic structural variations in IP-III patients with negative NGS results.
• Combining NGS and long-read sequencing provides a comprehensive approach to diagnosing DFNX2.