![]() Surprisingly, different pathogenic mutations of human TBC1D24 (TBC1 domain family member 24, OMIM 613577 Fig. Studies of these disorders have contributed to an appreciation of the broad range of in vivo functions of TBC-containing proteins, although their pathophysiology is not fully understood. Pathogenic variants of several genes encoding TBC domains contribute to a range of disorders in humans, including epilepsy, intellectual disability, pontocerebellar hypoplasia, dysmorphic features and deafness ( Table 1) ( 3–7). The human genome encodes 42 proteins that contain TBC domains (Tre-2–Bub2–Cdc16) ( 1), an ancient, conserved sequence consisting of approximately 200 amino acids ( 2). This is the first characterization of a mouse model of human TBC1D24-associated EIEE that can now be used to screen for antiepileptogenic drugs ameliorating TBCID24 seizure disorders. Immunogold electron microscopy using a TBC1D24-specific antibody revealed that TBC1D24 is associated with clathrin-coated vesicles and synapses of hippocampal neurons, suggesting a crucial role of TBC1D24 in vesicle trafficking important for neuronal signal transmission. Wild-type Tbc1d24 mRNA is abundantly expressed in the hippocampus using RNAscope in situ hybridization. A mouse deficient for alternative splice factor SRRM3 impairs incorporation of the Tbc1d24 micro-exon. S324Tfs *3 homozygotes show an abrupt onset of seizures at P15 that correlates with a developmental switch to utilization of the micro-exon. During embryonic and early postnatal development, S324Tfs *3 homozygotes produce predominantly the shorter wild-type TBC1D24 protein isoform that omits the micro-exon. The S324Tfs *3 variant is located in an alternatively spliced micro-exon encoding six perfectly conserved amino acids incorporated postnatally into TBC1D24 protein due to a micro-exon utilization switch. Homozygous S324Tfs *3 mice have normal auditory and vestibular functions but show an abrupt onset of spontaneous seizures at postnatal day 15 recapitulating human EIEE. Using CRISPR-Cas9 genome editing, we engineered a mouse with a premature translation stop codon equivalent to human S324Tfs *3, a recessive mutation of TBC1D24 associated with early infantile epileptic encephalopathy (EIEE). The mechanisms underlying TBC1D24-associated disorders and the functions of TBC1D24 are not well understood. Epilepsy, deafness, onychodystrophy, osteodystrophy and intellectual disability are associated with a spectrum of mutations of human TBC1D24. ![]()
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