Unveiling the Pathogenic Role of Novel <i>CPLANE1</i> Compound Heterozygous Variants in Joubert Syndrome: Insights Into <scp>mRNA</scp> Stability and <scp>NMD</scp> Pathway

Joubert syndrome (JS) is a rare neurodevelopmental disorder associated with mutations in genes involved in ciliary function. Germline variants in CPLANE1 have been implicated in JS. In this study, we investigated a family with three adverse pregnancies characterised by fetal malformations consistent with JS. Whole-exome sequencing (WES) identified compound heterozygous variants in CPLANE1: c.8893C>T (p.Gln2965*) and c.203C>T (p.Thr68Ile). Sanger sequencing confirmed the variants in the family. Bioinformatics analysis predicted that the c.203C>T variant affects mRNA splicing and protein function. Functional studies using PBMCs demonstrated that the c.203C>T variant causes exon 3 skipping, resulting in a frameshift and premature termination codon, leading to potential nonsense-mediated mRNA degradation (NMD). The mRNA transcription and translation inhibition experiment, by treatment with actinomycin D and puromycin, indicated that the c.203C>T variant leads to accelerated mRNA degradation. Notably, the inhibition of SMG1, a key marker of the NMD pathway, partially rescued mRNA expression in mutated cells, providing further evidence of NMD activation. Based on these findings and ACMG guidelines, the c.203C>T variant was reclassified from a variant of uncertain significance (VUS) to likely pathogenic. This is the first report of novel CPLANE1 compound heterozygous variants contributing to JS in this family. Our study expands the known pathogenic variant spectrum of CPLANE1 in JS and provides new insights into the molecular mechanisms of this ciliopathy.