Minigene assay for verifying the splicing effects of the rare variants in the SLC12A2 gene at c.2930-1G>a.

Background: Hearing loss (HL) is a prevalent sensory impairment with a genetic basis. The SLC12A2 gene, encoding NKCC1, is vital for inner ear ion balance. The c.2930-1G > A variant is a novel mutation potentially linked to sensorineural hearing loss.

Objectives: To investigate the splicing and protein expression effects of the c.2930-1G>A variant in SLC12A2 and its role in hearing loss.

Material and methods: Minigene assays and plasmid transfection in HEK-293T and Hela cells were used to study splicing. Protein expression and modification were also assessed.

Results: The c.2930-1G>A variant caused partial exon skipping, altering mRNA splicing in both cell types. This suggests a potential involvement in sensorineural hearing loss. Protein analysis showed the E21del mutation increased expression without altering modification patterns, whereas the 2930_2977del mutation reduced both, possibly impacting stability or modification sites.

Conclusions and significance: The c.2930-1G>A variant likely contribute to hearing loss by disrupting splicing and protein expression. Currently a Variant of Uncertain Significance (VUS), its pathogenicity is supported by these findings. Further research is needed to confirm its role, emphasizing the need for integrated genetic and clinical data in auditory disorders management.