Out-of-frame translation rescues a loss-of-function variant in a novel TBCE phenotype

Pathogenic variants in the TBCE gene, encoding tubulin-specific chaperone E crucial for tubulin folding, are linked to three severe neurodevelopmental disorders: Hypoparathyroidism-retardation-dysmorphism (HRD) syndrome, Kenny-Caffey syndrome type 1, and progressive encephalopathy with amyotrophy and optic atrophy. We identified patients with a novel, milder TBCE-associated phenotype and aimed to characterize it at the clinical and molecular levels. We conducted splicing analysis using deep NGS sequencing of RT-PCR products and detected TBCE through Western blotting. Translation efficiency was measured using a luciferase reporter assay. Overexpression experiments were performed in Hela cells with tubulin staining. Immunofluorescence analysis was used for Golgi complex assessment, while microtubule dynamics were studied post-nocodazole treatment. Electron microscopy facilitated ultrastructural studies. We report seven patients with a novel, milder TBCE phenotype, presenting with amyotrophy, testicular failure, and mild intellectual disability, with or without short stature. All patients were homozygous or compound-heterozygous for the NM_003193.5:c.100+1G>A variant, which causes a splicing alteration and early frameshift. However, we found that the mild phenotype arises due to translation from an alternative open reading frame, producing a partially functional protein. Dermal fibroblasts showed reduced Golgi compactness but normal microtubule dynamics. Electron microscopy revealed varying levels of acto-myosin degradation. The c.100+1G>A variant was found to be 10 times more frequent in Slavic samples than in gnomAD, suggesting underdiagnosis of this phenotype. This study uncovers complex molecular mechanisms contributing to the milder phenotype in patients with the c.100+1G>A variant, providing insights into a new TBCE-related disorder.