Multilevel Genetic and Functional Assessment of an ARR3 Frameshift Variant in Early-Onset High Myopia

<h4>Background</h4>Early-onset high myopia (eoHM) is a highly heritable ocular disorder, with pathogenic variants in the X-linked ARR3 gene (which encodes cone arrestin) being associated with a female-limited form of eoHM. However, the clinical manifestations of early truncating variants in ARR3 and their potential impact on the classic sex-limited pattern remain insufficiently understood.<h4>Methods</h4>This study examined individuals with eoHM from a multigenerational Chinese family, utilizing whole-exome sequencing, Sanger validation, and segregation analysis to identify candidate variants. In silico predictions and protein structure modeling were conducted to assess the variant's pathogenicity. Furthermore, in cultured cells, quantitative real-time PCR, Western blotting, and fluorescence microscopy were used to evaluate ARR3 transcript expression and ARR3-related immunodetected signal after transfection with wild-type or mutant ARR3 constructs, thereby providing a preliminary functional assessment of the variant.<h4>Results</h4>We discovered a novel frameshift variant, c.721delT (p.Y241Ifs∗3), which cosegregated with eoHM in the family and was absent in public population databases. Affected individuals, including hemizygous males, exhibited early-onset, high-degree myopia and fundus changes consistent with pathologic myopia. Functional assays in ARPE-19 cells showed that the mutant construct was associated with markedly reduced ARR3 mRNA expression and decreased ARR3 immunoreactivity compared with the wild-type construct. Because no direct cellular injury assay was performed, these in vitro findings were interpreted as preliminary expression-level evidence supporting a likely loss-of-function mechanism.<h4>Conclusion</h4>The novel ARR3 frameshift variant c.721delT (p.Y241Ifs∗3) is likely pathogenic for eoHM and may alter the previously understood female-limited inheritance pattern. These findings expand the known mutation spectrum of ARR3 and enhance understanding of the role of cone arrestin dysfunction in the development of eoHM.