Matching variants for functional characterization of genetic variants

Area of Science:

• Genetics and Genomics
• Cell Biology
• Developmental Biology

Background:

• Genetic diagnosis advances rapidly, yet interpreting genetic variants, especially missense mutations, remains challenging.
• Computational predictions and high-throughput experiments have limitations in variant interpretation.
• Utilizing model organisms and existing mutant resources can aid in understanding gene function and variant effects.

Purpose of the Study:

• To functionally characterize missense and stop-codon variants in the intraflagellar transport protein 140 (IFT-140) gene.
• To use Caenorhabditis elegans as a model to assess the phenotypic impact of IFT-140 variants, including those matching human disease variants.
• To provide experimental support for the interpretation of human genetic variants associated with ciliopathies.

Main Methods:

• Generated and analyzed ten variants (eight missense, two stop codons) in the C. elegans IFT-140 gene.
• Created specific human-matching variants (MatchVars) using CRISPR/Cas9 gene editing in C. elegans.
• Assessed ciliary phenotypes, including cilia length, intraflagellar transport (IFT) accumulation, and protein localization.

Main Results:

• Two variants, IFT-140(P702A) and IFT-140(W937stop), phenocopied the null mutant phenotype, exhibiting short cilia, IFT accumulations, and protein mislocalization.
• Other analyzed variants did not show significant ciliary defects in C. elegans.
• The study successfully generated and functionally validated C. elegans MatchVars for human IFT-140 variants.

Conclusions:

• The C. elegans model is effective for the functional assessment of IFT-140 variants.
• The identified phenotypic variants (P702A and W937stop) provide valuable insights into IFT-140 function and human disease mechanisms.
• This approach using model organism mutants aids in the interpretation of genetic variants of unknown significance in human diseases.