Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy
Mutations in the GAP activity toward RAGs 1 (GATOR1) complex genes (DEPDC5, NPRL2 and NPRL3) have been associated with focal epilepsy and focal cortical dysplasia (FCD). GATOR1 functions as an inhibitor of the mTORC1 signalling pathway, indicating that the downstream effects of mTORC1 deregulation u...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2020-02-01
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| Series: | Neurobiology of Disease |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0969996119303158 |
| _version_ | 1818573260069011456 |
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| author | Ruby E. Dawson Alvaro F. Nieto Guil Louise J. Robertson Sandra G. Piltz James N. Hughes Paul Q. Thomas |
| author_facet | Ruby E. Dawson Alvaro F. Nieto Guil Louise J. Robertson Sandra G. Piltz James N. Hughes Paul Q. Thomas |
| author_sort | Ruby E. Dawson |
| collection | DOAJ |
| description | Mutations in the GAP activity toward RAGs 1 (GATOR1) complex genes (DEPDC5, NPRL2 and NPRL3) have been associated with focal epilepsy and focal cortical dysplasia (FCD). GATOR1 functions as an inhibitor of the mTORC1 signalling pathway, indicating that the downstream effects of mTORC1 deregulation underpin the disease. However, the vast majority of putative disease-causing variants have not been functionally assessed for mTORC1 repression activity. Here, we develop a novel in vitro functional assay that enables rapid assessment of GATOR1-gene variants. Surprisingly, of the 17 variants tested, we show that only six showed significantly impaired mTORC1 inhibition. To further investigate variant function in vivo, we generated a conditional Depdc5 mouse which modelled a ‘second-hit’ mechanism of disease. Generation of Depdc5 null ‘clones’ in the embryonic brain resulted in mTORC1 hyperactivity and modelled epilepsy and FCD symptoms including large dysmorphic neurons, defective migration and lower seizure thresholds. Using this model, we validated DEPDC5 variant F164del to be loss-of-function. We also show that Q542P is not functionally compromised in vivo, consistent with our in vitro findings. Overall, our data show that mTORC1 deregulation is the central pathological mechanism for GATOR1 variants and also indicates that a significant proportion of putative disease variants are pathologically inert, highlighting the importance of GATOR1 variant functional assessment. |
| first_indexed | 2024-12-15T00:08:48Z |
| format | Article |
| id | doaj.art-e1fc166522f74d62b0762995995d9340 |
| institution | Directory Open Access Journal |
| issn | 1095-953X |
| language | English |
| last_indexed | 2024-12-15T00:08:48Z |
| publishDate | 2020-02-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Neurobiology of Disease |
| spelling | doaj.art-e1fc166522f74d62b0762995995d93402022-12-21T22:42:39ZengElsevierNeurobiology of Disease1095-953X2020-02-01134Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsyRuby E. Dawson0Alvaro F. Nieto Guil1Louise J. Robertson2Sandra G. Piltz3James N. Hughes4Paul Q. Thomas5School of Biological Sciences, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, AustraliaSchool of Medicine, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, AustraliaSchool of Medicine, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, AustraliaSchool of Medicine, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, AustraliaSchool of Biological Sciences, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, AustraliaSchool of Medicine, University of Adelaide, Adelaide, SA 5005, Australia; Robinson Research Institute, University of Adelaide, Adelaide, SA 5005, Australia; Precision Medicine Theme, South Australia Health and Medical Research Institute, Adelaide, SA 5000, Australia; Corresponding author at: School of Medicine, University of Adelaide, Adelaide, SA 5005, AustraliaMutations in the GAP activity toward RAGs 1 (GATOR1) complex genes (DEPDC5, NPRL2 and NPRL3) have been associated with focal epilepsy and focal cortical dysplasia (FCD). GATOR1 functions as an inhibitor of the mTORC1 signalling pathway, indicating that the downstream effects of mTORC1 deregulation underpin the disease. However, the vast majority of putative disease-causing variants have not been functionally assessed for mTORC1 repression activity. Here, we develop a novel in vitro functional assay that enables rapid assessment of GATOR1-gene variants. Surprisingly, of the 17 variants tested, we show that only six showed significantly impaired mTORC1 inhibition. To further investigate variant function in vivo, we generated a conditional Depdc5 mouse which modelled a ‘second-hit’ mechanism of disease. Generation of Depdc5 null ‘clones’ in the embryonic brain resulted in mTORC1 hyperactivity and modelled epilepsy and FCD symptoms including large dysmorphic neurons, defective migration and lower seizure thresholds. Using this model, we validated DEPDC5 variant F164del to be loss-of-function. We also show that Q542P is not functionally compromised in vivo, consistent with our in vitro findings. Overall, our data show that mTORC1 deregulation is the central pathological mechanism for GATOR1 variants and also indicates that a significant proportion of putative disease variants are pathologically inert, highlighting the importance of GATOR1 variant functional assessment.http://www.sciencedirect.com/science/article/pii/S0969996119303158NeurodevelopmentEpilepsyFocal cortical dysplasiaMolecular geneticsFunctional testingDevelopmental genetics |
| spellingShingle | Ruby E. Dawson Alvaro F. Nieto Guil Louise J. Robertson Sandra G. Piltz James N. Hughes Paul Q. Thomas Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy Neurobiology of Disease Neurodevelopment Epilepsy Focal cortical dysplasia Molecular genetics Functional testing Developmental genetics |
| title | Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy |
| title_full | Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy |
| title_fullStr | Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy |
| title_full_unstemmed | Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy |
| title_short | Functional screening of GATOR1 complex variants reveals a role for mTORC1 deregulation in FCD and focal epilepsy |
| title_sort | functional screening of gator1 complex variants reveals a role for mtorc1 deregulation in fcd and focal epilepsy |
| topic | Neurodevelopment Epilepsy Focal cortical dysplasia Molecular genetics Functional testing Developmental genetics |
| url | http://www.sciencedirect.com/science/article/pii/S0969996119303158 |
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