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Original research
Functional analysis of TLK2 variants and their proximal interactomes implicates impaired kinase activity and chromatin maintenance defects in their pathogenesis
  1. Lisa Pavinato1,2,
  2. Marina Villamor-Payà3,
  3. Maria Sanchiz-Calvo3,
  4. Cristina Andreoli4,
  5. Marina Gay3,
  6. Marta Vilaseca3,
  7. Gianluca Arauz-Garofalo3,
  8. Andrea Ciolfi5,
  9. Alessandro Bruselles6,
  10. Tommaso Pippucci7,
  11. Valentina Prota4,
  12. Diana Carli8,
  13. Elisa Giorgio1,9,
  14. Francesca Clementina Radio5,
  15. Vincenzo Antona10,
  16. Mario Giuffrè10,
  17. Kara Ranguin11,
  18. Cindy Colson11,
  19. Silvia De Rubeis12,13,14,15,
  20. Paola Dimartino16,
  21. Joseph D Buxbaum12,13,14,15,17,18,
  22. Giovanni Battista Ferrero8,
  23. Marco Tartaglia5,
  24. Simone Martinelli19,
  25. Travis H Stracker3,20,
  26. Alfredo Brusco1,21
  1. 1Department of Medical Sciences, University of Turin, Torino, Italy
  2. 2Institute of Human Genetics and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany
  3. 3The Barcelona Institute of Science and Technology, Institute for Research in Biomedicine, Barcelona, Spain
  4. 4Department of Environment and Health, Istituto Superiore di Sanità, Roma, Italy
  5. 5Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino Gesù IRCCS, Roma, Italy
  6. 6Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy
  7. 7Medical Genetics Unity, Sant'Orsola-Malpighi University Hospital, Bologna, Italy
  8. 8Department of Pediatrics and Public Health and Pediatric Sciences, University of Turin, Torino, Italy
  9. 9Department of Molecular Medicine, University of Pavia, Pavia, Italy
  10. 10Department of Sciences for Health Promotion and Mother and Child Care "G. D'Alessandro", University of Palermo, Palermo, Italy
  11. 11Department of Genetics, Reference center for Rare Diseases and Developmental Anomalies, Caen, France
  12. 12Seaver Autism Center for Research and Treatment, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  13. 13Department of Psychiatry, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  14. 14The Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  15. 15Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  16. 16Department of Medical and Surgical Sciences, University of Bologna, Bologna, Italy
  17. 17Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  18. 18Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York City, New York, USA
  19. 19Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Roma, Italy
  20. 20Radiation Oncology Branch, National Cancer Institute, Bethesda, Maryland, USA
  21. 21Unit of Medical Genetics, "Città della Salute e della Scienza" University Hospital, Torino, Italy
  1. Correspondence to Professor Alfredo Brusco, Department of Medical Sciences, University of Turin, Torino 10124, Italy; alfredo.brusco{at}


Introduction The Tousled-like kinases 1 and 2 (TLK1 and TLK2) are involved in many fundamental processes, including DNA replication, cell cycle checkpoint recovery and chromatin remodelling. Mutations in TLK2 were recently associated with ‘Mental Retardation Autosomal Dominant 57’ (MRD57, MIM# 618050), a neurodevelopmental disorder characterised by a highly variable phenotype, including mild-to-moderate intellectual disability, behavioural abnormalities, facial dysmorphisms, microcephaly, epilepsy and skeletal anomalies.

Methods We re-evaluate whole exome sequencing and array-CGH data from a large cohort of patients affected by neurodevelopmental disorders. Using spatial proteomics (BioID) and single-cell gel electrophoresis, we investigated the proximity interaction landscape of TLK2 and analysed the effects of p.(Asp551Gly) and a previously reported missense variant (c.1850C>T; p.(Ser617Leu)) on TLK2 interactions, localisation and activity.

Results We identified three new unrelated MRD57 families. Two were sporadic and caused by a missense change (c.1652A>G; p.(Asp551Gly)) or a 39 kb deletion encompassing TLK2, and one was familial with three affected siblings who inherited a nonsense change from an affected mother (c.1423G>T; p.(Glu475Ter)). The clinical phenotypes were consistent with those of previously reported cases. The tested mutations strongly impaired TLK2 kinase activity. Proximal interactions between TLK2 and other factors implicated in neurological disorders, including CHD7, CHD8, BRD4 and NACC1, were identified. Finally, we demonstrated a more relaxed chromatin state in lymphoblastoid cells harbouring the p.(Asp551Gly) variant compared with control cells, conferring susceptibility to DNA damage.

Conclusion Our study identified novel TLK2 pathogenic variants, confirming and further expanding the MRD57-related phenotype. The molecular characterisation of missense variants increases our knowledge about TLK2 function and provides new insights into its role in neurodevelopmental disorders.

  • genetic research
  • genetics
  • medical
  • loss of function mutation
  • molecular biology
  • mutation
  • missense

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  • THS and AB contributed equally.

  • Contributors MV-P, MSC, MG, MV and GA-G performed activity, localisation and BioID-MS analysis. LP wrote and edited the manuscript, interpreted exome data, collected the cases and performed variant confirmation, mRNA and splicing analysis. CA and VP performed the SCGE assay. EG interpreted exome data. DC, VAMG, KR, CC and GBF followed patients and collected their clinical information. SDR and JB performed exome sequencing. AB, TP, PD, AC, FCR and MT processed and analysed the WES data. THS and AB designed and supervised the project, analyzed data and wrote the manuscript.

  • Funding This research received funding specifically appointed to Department of Medical Sciences from the Italian Ministry for Education, University and Research (Ministero dell’Istruzione, dell’Università e della Ricerca—MIUR) under the programme 'Dipartimenti di Eccellenza 2018–2022' Project code D15D18000410001. MV-P was funded by an FPI fellowship from the Ministry of Science, Innovation and Universities (MCIU) and MSC by a Masters fellowship from the BIST and support from the IRB Barcelona. THS was funded by the MCIU (PGC2018-095616-B-I00/GINDATA and FEDER). MT was funded by Fondazione Bambino Gesù (Vite Coraggiose). The whole exome sequencing was performed as part of the Autism Sequencing Consortium and was supported by the NIMH (MH111661). Thanks to the MSPCF of IRB Barcelona, a principal unit in Proteored, PRB3, supported by PT17/0019 of the PE I+D+i 2013-2016, funded by SCIII and ERDF. This study makes use of data generated by the DECIPHER community. A full list of centres that contributed to the generation of the data is available from: and via email from: Funding for the project was provided by the Wellcome Trust.

  • Competing interests None declared.

  • Patient consent for publication Parental/Guardian consent obtained.

  • Ethics approval The study protocol was approved by the internal Ethics Committee, according to the Declaration of Helsinki and informed consent was obtained from participating families.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Data availability statement Data are available in a public, open access repository or included in the article or uploaded as supplementary information.

  • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.

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