Long-read genome sequencing identifies cryptic structural variants in congenital aniridia cases

Area of Science:

• Genetics
• Genomics
• Molecular Biology

Background:

• Congenital aniridia is primarily caused by PAX6 haploinsufficiency.
• 11p13 microdeletions affecting PAX6 or its regulatory regions occur in 25% of patients.
• Complex rearrangements are rarely described, leaving some cases unexplained.

Purpose of the Study:

• To investigate cryptic structural variants (SVs) in two unsolved congenital aniridia cases using nanopore-based whole-genome sequencing.
• To identify the underlying genetic cause in PAX6-negative congenital aniridia patients.

Main Methods:

• Nanopore-based whole-genome sequencing (long-read sequencing - LRS) was employed.
• Targeted polymerase chain reaction, sequencing, and FISH-based cytogenetic analysis were used for verification.
• RNA expression analysis was performed on patient-derived samples.

Main Results:

• LRS identified a de novo 4.9 Mb inversion disrupting PAX6 intron 7 in one patient.
• LRS resolved a t(6;11) balanced translocation in the second patient, with the 11p13 breakpoint affecting a PAX6 enhancer.
• Both identified SVs were confirmed as the pathogenic cause of congenital aniridia, leading to PAX6 haploinsufficiency.

Conclusions:

• Long-read sequencing is effective in detecting complex structural variants missed by short-read sequencing.
• This study highlights the utility of LRS in diagnosing rare genetic diseases with complex genomic rearrangements.
• The findings emphasize the importance of considering SVs in the genetic basis of congenital aniridia.