Preprint / Version 1

Loss of Tousled-Like Kinase 2 Reduces Antioxidative Responses and Disrupts Brain Development

This article is a preprint and has not been certified by peer review.

Authors

    Qiannan Deng,  
    Qiannan Deng
    Fanjia Hou,  
    Fanjia Hou
    Xiaoqi Zhou,  
    Xiaoqi Zhou
    Yiwen Zhang,  
    Yiwen Zhang
    • Zhejiang University School of Medicine Children's Hospital
    Qing Zhang,  
    Qing Zhang
    Jiaqi Zhang,  
    Jiaqi Zhang
    • Chongqing Medical University
    Zihan Zhang,  
    Zihan Zhang
    Tongfei Liu,  
    Tongfei Liu
    • Chongqing Medical University
    Wen Yuan,  
    Wen Yuan
    Chaochun Zou,  
    Chaochun Zou
    • Zhejiang University School of Medicine Children's Hospital
    Chao Tong,  
    Chao Tong
    Ke Zhang
    Ke Zhang
Categories
Neuroscience & Neurology
Keywords
TLK2; rare neurodevelopmental disorder; microcephaly; oxidative stress

Abstract

Haploinsufficiency of tousled-like kinase 2 (TLK2) causes intellectual developmental disorder, autosomal dominant 57, a rare neurodevelopmental disorder with unclear pathomechanism and no effective treatment. Here, we established Drosophila models with neurodevelopmental defects resembling some disease features, such as microcephaly. Further studies suggested that oxidative stress plays a critical role in these models. Mechanistically, Tlk (the fly homolog of TLK2) loss reduces the expression of antioxidative genes via activating transcription factor 4 (ATF4) and its downstream protein, nuclear factor erythroid 2-related factor 2 (Nrf2), a transcriptional activator of antioxidative genes, in fly neural stem cells (NSCs), thereby disrupting NSC differentiation and causing microcephaly. In patient-derived brain organoids, ATF4, Nrf2, and their downstream antioxidative genes are also downregulated. Agreeing with these data, these organoids exhibited oxidative stress and microcephaly. Combined, our findings suggest that ATF4/Nrf2-mediated antioxidative stress responses play a critical role in TLK2-related disease and may be a therapeutic target.

References

Au, W.H., L. Miller-Fleming, A. Sanchez-Martinez, J.A. Lee, M.J. Twyning, H.A. Prag, L. Raik, S.P. Allen, P.J. Shaw, L. Ferraiuolo, H. Mortiboys and A.J. Whitworth (2024). "Activation of the Keap1/Nrf2 pathway suppresses mitochondrial dysfunction, oxidative stress, and motor phenotypes in C9orf72 ALS/FTD models." Life Sci Alliance 7(9).

Bruinsma, W., J. van den Berg, M. Aprelia and R.H. Medema (2016). "Tousled-like kinase 2 regulates recovery from a DNA damage-induced G2 arrest." EMBO Rep 17(5): 659-670.

Deng, Q., C. Wang, C.T. Koe, J.P. Heinen, Y.S. Tan, S. Li, C. Gonzalez, W.K. Sung and H. Wang (2022). "Parafibromin governs cell polarity and centrosome assembly in Drosophila neural stem cells." PLoS Biol 20(10): e3001834.

Doe, C.Q. (2008). "Neural stem cells: balancing self-renewal with differentiation." Development 135(9): 1575-1587.

Gao, L., F. Wang, Y. Wang, L. Hu and J. Mao (2022). "A protocol for the generation of patient-specific iPSC lines from peripheral blood mononuclear cells." STAR Protoc 3(3): 101530.

He, T., B. Xu and H. Ma (2024). "TLK2 promotes progression of hepatocellular carcinoma through Wnt/β-catenin signaling." Transl Cancer Res 13(7): 3729-3741.

Kim, J.A., Y. Tan, X. Wang, X. Cao, J. Veeraraghavan, Y. Liang, D.P. Edwards, S. Huang, X. Pan, K. Li, R. Schiff and X.S. Wang (2016). "Comprehensive functional analysis of the tousled-like kinase 2 frequently amplified in aggressive luminal breast cancers." Nat Commun 7: 12991.

Klimovskaia, I.M., C. Young, C.B. Stromme, P. Menard, J. Jasencakova, K. Mejlvang, K. Ask, M. Ploug, M.L. Nielsen, O.N. Jensen and A. Groth (2014). "Tousled-like kinases phosphorylate Asf1 to promote histone supply during DNA replication." Nature Communications 5(1): 3394.

Lee, S.B., S. Segura-Bayona, M. Villamor-Paya, G. Saredi, M.A.M. Todd, C.S. Attolini, T.Y. Chang, T.H. Stracker and A. Groth (2018). "Tousled-like kinases stabilize replication forks and show synthetic lethality with checkpoint and PARP inhibitors." Sci Adv 4(8): eaat4985.

Li, H.Y., C.M. Jiang, R.Y. Liu and C.C. Zou (2024). "Report of one case with de novo mutation in TLK2 and literature review." BMC Pediatr 24(1): 732.

Ohn, T., N. Kedersha, T. Hickman, S. Tisdale and P. Anderson (2008). "A functional RNAi screen links O-GlcNAc modification of ribosomal proteins to stress granule and processing body assembly." Nat. Cell Biol 10(10): 1224-1231.

Omata, T., H. Aoyama, K. Murayama, M. Takayanagi, R. Kawaguchi, R. Fujimoto and J.I. Takanashi (2024). "Efficacy of a mitochondrial drug cocktail in preventing acute encephalopathy with biphasic seizures and late reduced diffusion." J Neurol Sci 466: 123245.

Pavinato, L., M. Villamor-Paya, M. Sanchiz-Calvo, C. Andreoli, M. Gay, M. Vilaseca, G. Arauz-Garofalo, A. Ciolfi, A. Bruselles, T. Pippucci, V. Prota, D. Carli, E. Giorgio, F.C. Radio, V. Antona, M. Giuffre, K. Ranguin, C. Colson, S. De Rubeis, P. Dimartino, J.D. Buxbaum, G.B. Ferrero, M. Tartaglia, S. Martinelli, T.H. Stracker and A. Brusco (2022). "Functional analysis of TLK2 variants and their proximal interactomes implicates impaired kinase activity and chromatin maintenance defects in their pathogenesis." J Med Genet 59(2): 170-179.

Pilotto, F., D.M. Chellapandi and H. Puccio (2024). "Omaveloxolone: a groundbreaking milestone as the first FDA-approved drug for Friedreich ataxia." Trends Mol Med

Port, F. and S.L. Bullock (2016). "Augmenting CRISPR applications in Drosophila with tRNA-flanked sgRNAs." Nature Methods 13(10): 852-854.

Reijnders, M.R.F., K.A. Miller, M. Alvi, J.A.C. Goos, M.M. Lees, A. de Burca, A. Henderson, A. Kraus, B. Mikat, B.B.A. de Vries, B. Isidor, B. Kerr, C. Marcelis, C. Schluth-Bolard, C. Deshpande, C.A.L. Ruivenkamp, D. Wieczorek, D. Baralle, E.M. Blair, H. Engels, H.-J. Ludecke, J. Eason, G.W.E. Santen, J. Clayton-Smith, K. Chandler, K. Tatton-Brown, K. Payne, K. Helbig, K. Radtke, K.M. Nugent, K. Cremer, T.M. Strom, L.M. Bird, M. Sinnema, M. Bitner-Glindzicz, M.F. van Dooren, M. Alders, M. Koopmans, L. Brick, M. Kozenko, M.L. Harline, M. Klaassens, M. Steinraths, N.S. Cooper, P. Edery, P. Yap, P.A. Terhal, P.J. van der Spek, P. Lakeman, R.L. Taylor, R.O. Littlejohn, R. Pfundt, S. Mercimek-Andrews, A.P.A. Stegmann, S.G. Kant, S. McLean, S. Joss, S.M.A. Swagemakers, S. Douzgou, S.A. Wall, S. Küry, E. Calpena, N. Koelling, S.J. McGowan, S.R.F. Twigg, I.M.J. Mathijssen, C. Nellaker, H.G. Brunner and A.O.M. Wilkie (2018). "De Novo and Inherited Loss-of-Function Variants in TLK2: Clinical and Genotype-Phenotype Evaluation of a Distinct Neurodevelopmental Disorder." The American Journal of Human Genetics 102(6): 1195-1203.

Sarcinelli, C., H. Dragic, M. Piecyk, V. Barbet, C. Duret, A. Barthelaix, C. Ferraro-Peyret, J. Fauvre, T. Renno, C. Chaveroux and S.N. Manie (2020). "ATF4-Dependent NRF2 Transcriptional Regulation Promotes Antioxidant Protection during Endoplasmic Reticulum Stress." Cancers(Basel) 12(3).

Segura-Bayona, S., P.A. Knobel, H. Gonzalez-Burón, S.A. Youssef, A. Pea-Blanco, E. Coyaud, T. López-Rovira, K. Rein, L. Palenzuela, J. Colombelli, S. Forrow, B. Raught, A. Groth, A. de Bruin and T.H. Stracker (2017). "Differential requirements for Tousled-like kinases 1 and 2 in mammalian development." Cell Death& Differentiation 24(11): 1872-1885.

Segura-Bayona, S. and T.H. Stracker (2019). "The Tousled-like kinases regulate genome and epigenome stability: implications in development and disease." Cell Mol Life Sci 76(19): 3827-3841.

Siller, K.H. and C.Q. Doe (2009). "Spindle orientation during asymmetric cell division." Nature Cell Biology 11(4): 365-374.

Suzuki, M., A. Otsuki, N. Keleku-Lukwete and M. Yamamoto (2016). "Overview of redox regulation by Keap1-Nrf2 system in toxicology and cancer." Current Opinion in Toxicology 1: 29-36.

Wang, M., J. Li, X. Yang, Q. Yan, H. Wang, X. Xu, Y. Lu, D. Li, Y. Wang, R. Sun, S. Zhang, Y. Zhang, Z. Zhang, F. Meng and Y. Li (2023). "Targeting TLK2 inhibits the progression of gastric cancer by reprogramming amino acid metabolism through the mTOR/ASNS axis." Cancer Gene Therapy 30(11): 1485-1497.

Wang, S., X.A. Chen, J. Hu, J.K. Jiang, Y. Li, K.Y. Chan-Salis, Y. Gu, G. Chen, C. Thomas, B.F. Pugh and Y. Wang (2015). "ATF4 Gene Network Mediates Cellular Response to the Anticancer PAD Inhibitor YW3-56 in Triple-Negative Breast Cancer Cells." Mol Cancer Ther 14(4): 877-888.

Yang, P., C. Mathieu, R.M. Kolaitis, P. Zhang, J. Messing, U. Yurtsever, Z. Yang, J. Wu, Y. Li, Q. Pan, J. Yu, E.W. Martin, T. Mittag, H.J. Kim and J.P. Taylor (2020). "G3BP1 Is a Tunable Switch that Triggers Phase Separation to Assemble Stress Granules." Cell 181(2): 325-345 e328.

Yoshino, H. (2019). "Edaravone for the treatment of amyotrophic lateral sclerosis." Expert Rev Neurother 19(3): 185-193.

Zhang, Y., R. Cai, R. Zhou, Y. Li and L. Liu (2016). "Tousled-like kinase mediated a new type of cell death pathway in Drosophila." Cell Death& Differentiation 23(1): 146-157.

Zhou, J., L. Xu, X. Duan, W. Liu, X. Zhao, X. Wang, W. Shang, X. Fang, H. Yang, L. Jia, J. Bai, J. Zhao, L. Wang and C. Tong (2019). "Large-scale RNAi screen identified Dhpr as a regulator of mitochondrial morphology and tissue homeostasis." Sci Adv 5(9): eaax0365.

Metrics

Favorites: 1
Views: 406
Downloads: 57

DOI:

https://doi.org/10.65215/LTSpreprints.2026.03.06.000145

Submission ID:

145

Downloads

Posted

2026-03-07

How to Cite

Deng, Q., Hou, F., Zhou, X., Zhang, Y., Zhang, Q., Zhang, J., Zhang, Z., Liu, T., Yuan, W., Zou, C., Tong, C., & Zhang, K. (2026). Loss of Tousled-Like Kinase 2 Reduces Antioxidative Responses and Disrupts Brain Development. LangTaoSha Preprint Server. https://doi.org/10.65215/LTSpreprints.2026.03.06.000145

Download Citation

Endnote/Zotero/Mendeley BibTeX

Declaration of Competing Interests

The authors declare no competing interests to disclose.

Copyright

The copyright holder for this preprint is the author/funder.

Creative Commons License

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.