Interaction between the TBC1D24 TLDc domain and the KIBRA C2 domain is disrupted by two epilepsy-associated TBC1D24 missense variants

Mutations of human TBC1D24 are associated with deafness,
epilepsy, or DOORS syndrome (deafness, onychodystrophy, osteodystrophy,
cognitive disability, and seizures). The causal relationships
between TBC1D24 variants and the different clinical
phenotypes are not understood. Our hypothesis is that phenotypic
heterogeneity of missense mutations of TBC1D24 results,
in part, from perturbed binding of different protein partners. To discover
novel protein partners of TBC1D24, we conducted yeast two-hybrid (Y2H)
screen using mouse full-length TBC1D24 as bait. Kidney and brain protein
(KIBRA), a scaffold protein encoded by Wwc1, was identified as a
partner of TBC1D24. KIBRA functions in the Hippo signaling pathway and is
important for human cognition and memory. The TBC1D24 TLDc domain binds to
KIBRA full-length and to its C2 domain, confirmed by Y2H assays. No
interaction was detected with Y2H assays between the KIBRA C2 domain and
TLDc domains of NCOA7, MEAK7, and OXR1. Two epilepsy-associated recessive
variants (Gly511Arg and Ala515Val) in the TLDc domain of human TBC1D24
disrupt the interaction with the human KIBRA C2 domain. In the case of
mouse constructs, Y2H assays show only mouse TBC1D24 Ala517Val
corresponding to Ala515Val of human TBC1D24 abolished the interaction
between mouse TBC1D24 and mouse KIBRA. This study reveals a pathogenic
mechanism of TBC1D24-associated epilepsy, linking the TBC1D24 and KIBRA
pathways. The interaction of TBC1D24-KIBRA is physiologically meaningful
and necessary to reduce the risk of epilepsy.