Interaction between the TBC1D24 TLDc domain and the KIBRA C2 domain is disrupted by two epilepsy-associated TBC1D24 missense variants
Data files
Sep 30, 2025 version files 2.63 MB
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Figure_1-2-9.pdf
989.17 KB
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Figure_10A.pdf
420.90 KB
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Figure_10B.pdf
670.18 KB
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Figure_4-S3.pdf
540.17 KB
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README.md
5.54 KB
Abstract
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.
This folder contains the data files for the article:
Tona, R., Inagaki, S., Ishibashi, Y., Faridi, R., Yousaf, R., Roux, I., Wilson, E., Fenollar-Ferrer, C., Chien, W.W., Belyantseva, I.A. and Friedman, T.B. (2024). Interaction between the TBC1D24 TLDc domain and the KIBRA C2 domain is disrupted by two epilepsy-associated TBC1D24 missense variants. Journal of Biological Chemistry, 300, 107725.
DOI: 10.1016/j.jbc.2024.107725
This dataset contains previously unpublished data collected by the Laboratory of Molecular Genetics, National Institute on Deafness and Communication Disorders, National Institutes of Health, and associated with published data in Journal of Biological Chemistry (DOI: 10.1016/j.jbc.2024.107725). If you use any of the data in this dataset, please cite the associated publication
Description of the data
Yeast two-hybrid (Y2H) assays presented here show our published results are reproducible and verified by a separate source, Hybrigenics Service in France Link.
The experimental protocol for the Y2H assays, yeast strains and sequence information of bait and prey constructs were described in “yeast two hybrid screening” in Experimental procedures and Supplementary Table S6 of the original paper (DOI: 10.1016/j.jbc.2024.107725).
Data
The data folder contains four files. Original data are provided by Hybrigenics Service.
Figure_1-2-9.pdf: Y2H assays showing that mouse TBC1D24 TLDc domain interacts with mouse KIBRA C2 domain, and human TBC1D24 TLDc domain interacts with human KIBRA C2 domain as well.
Figure_4-S3.pdf: Y2H assays using the TLDc domains of the five mouse TLDc domain-containing proteins and mouse KIBRA C2 domain as bait and prey, respectively, to indicate only the mouse TBC1D24 TLDc domain interacts with the mouse KIBRA C2 domain.
Figure_10A.pdf: Y2H assay showing human TBC1D24 Arg515Val abolished the interaction between human TBC1D24 and human KIBRA, and human TBC1D24 Gly511Arg reduces the interaction between human TBC1D24 and human KIBRA.
Figure_10B.pdf: Y2H assay showing only mouse TBC1D24 A517V abolished the interaction between mouse TBC1D24 and mouse KIBRA.
Files:
File 1: Figure_1-2-9.pdf
- Content: one pdf file
- Description: Y2H assays showing that mouse TBC1D24 TLDc domain interacts with mouse KIBRA C2 domain, and human TBC1D24 TLDc domain interacts with the human KIBRA C2 domain as well.
- Page 1: Title
- Page 2: Abbreviations
- Page 3: Summary interaction matrix and results
- Page 4-5: Results of growth assays using mouse TBC1D24 and mouse KIBRA as bait and prey, respectively. Data is associated with Figures 1B and 2B of the original paper Link.
- Page 5-6: Results of growth assays using human TBC1D24 and human KIBRA as bait and prey, respectively. Data is associated with Figure 9 of the original paper Link.
File 2: Figure_4-S3.pdf
- Content: one pdf file
- Description: Y2H assays using the TLDc domains of the five mouse TLDc domain-containing proteins and mouse KIBRA C2 domain as bait and prey, respectively, to indicate only the mouse TBC1D24 TLDc domain interacts with the mouse KIBRA C2 domain.
- Page 1: Title
- Page 2: Abbreviations
- Page 3: Summary interaction matrix and results
- Page 4-6: Results of growth assays using TLDc domains of five TLDc domain-containing proteins and mouse KIBRA as bait and prey, respectively. Data is associated with Figures 4, S3D, and S3E of the original paper Link.
File 3: Figure_10A.pdf
- Content: one pdf file
- Description: Y2H assays showing that human TBC1D24 Ala515Val abolishes the interaction with human KIBRA, while the human TBC1D24 Gly511Arg disrupts the interaction only with a higher concentration of an inhibitor, 3-amino-1,2,4-triazole.
- Page 1: Title
- Page 2: Abbreviations
- Page 3: Summary interaction matrix and results
- Page 4: Results of growth assays using human TBC1D24 WT, Gly511Arg, and Ala515Val and mouse KIBRA as bait and prey, respectively.
- Page 5-6: Results of semiquantitative Y2H analysis using human TBC1D24 wild type (WT), G511R, and A515V and mouse KIBRA as bait and prey, respectively. Part of data was presented in Figure 10A of the original paper Link.
File 4: Figure_10B.pdf
- Content: one pdf file
- Description: Y2H assays using mouse TBC1D24 WT, Gly513Arg and Ala517Val and mouse KIBRA, as bait and prey, respectively. Mouse TBC1D24 Gly513 and Ala517 correspond to Gly511 and Ala515 of human TBC1D24, respectively. Mouse TBC1D24 Ala517Val abolishes the interaction with mouse KIBRA in all conditions tested. However, mouse TBC1D24 Gly513Arg allows for an interaction with mouse KIBRA unlike human TBC1D24 Gly511Arg presented in Figure 10A of the original paper Link.
- Page 1: Title
- Page 2: Abbreviations
- Page 3: Summary interaction matrix and results
- Page 4: Results of growth assays using human TBC1D24 WT, G511R, and A515V and mouse KIBRA as bait and prey, respectively.
- Page 5-6: Results of semiquantitative Y2H analysis using mouse TBC1D24 WT, Gly513Arg, and Ala515Val and mouse KIBRA as bait and prey, respectively. Part of data was presented in Figure 10B of the original paper Link.