Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Differential cell cycle checkpoint evasion by PTEN germline mutations associated with dichotomous phenotypes of cancer versus autism spectrum disorder

Oncogene volume 42, pages 3698–3707 (2023)Cite this article

Subjects

Abstract

Individuals with a PTEN germline mutation receive the molecular diagnosis of PTEN hamartoma tumor syndrome (PHTS). PHTS displays a complex spectrum of clinical phenotypes including harmartomas, predisposition to cancers, and autism spectrum disorder (ASD). Clear-cut genotype-phenotype correlations are yet to be established due to insufficient information on the PTEN function being impacted by mutations. To fill this knowledge gap, we compared functional impacts of two selected missense PTEN mutant alleles, G132D and M134R, each respectively being associated with distinct clinical phenotype, ASD or thyroid cancer without ASD using gene-edited human induced pluripotent stem cells (hiPSCs). In homozygous hiPSCs, PTEN expression was severely reduced by M134R mutation due to shortened protein half-life. G132D suppressed PTEN expression to a lesser extent than Μ134R mutation without altering protein half-life. When challenged with γ-irradiation, G132D heterozygous cells exited radiation-induced G2 arrest earlier than wildtype and M134R heterozygous hiPSCs despite the similar DNA damage levels as the latter two. Immunoblotting analyses suggested that γ-irradiation induced apoptosis in G132D heterozygous cells to lesser degrees than in the hiPSCs of other genotypes. These data suggest that ASD-associated G132D allele promotes genome instability by premature cell cycle reentry with incomplete DNA repair.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Expression of WT, M134R, and G132D PTEN in hiPSCs.
Fig. 2: PTEN protein half-life determination.
Fig. 3: Early release from the G2 cell cycle checkpoint in γ-irradiated G132D/WT hiPSCs.
Fig. 4: Levels of DNA damage and damage-repairing activity.
Fig. 5: Western blot analyses of checkpoint kinases, PTEN signaling, and apoptosis.

Similar content being viewed by others

Data availability

The authors will share any data relating to this work upon reasonable request and execution of data sharing agreement.

References

  1. Li J, Yen C, Liaw D, Podsypanina K, Bose S, Wang SI, et al. PTEN, a putative protein tyrosine phosphatase gene mutated in human brain, breast, and prostate cancer. Science. 1997;275:1943–7.

    Article  CAS  PubMed  Google Scholar 

  2. Steck PA, Pershouse MA, Jasser SA, Yung WKA, Lin H, Ligon AH, et al. Identification of a candidate tumour suppressor gene, MMAC1, at chromosome 10q23.3 that is mutated in multiple advanced cancers. Nat Genet. 1997;15:356–62.

    Article  CAS  PubMed  Google Scholar 

  3. Nelen MR, Padberg GW, Peeters EAJ, Lin AY, Helm Bvanden, Frants RR, et al. Localization of the gene for Cowden disease to chromosome 10q22–23. Nat Genet. 1996;13:114–6.

    Article  CAS  PubMed  Google Scholar 

  4. Liaw D, Marsh DJ, Li J, Dahia PLM, Wang SI, Zheng Z, et al. Germline mutations of the PTEN gene in Cowden disease, an inherited breast and thyroid cancer syndrome. Nat Genet. 1997;16:64–67.

    Article  CAS  PubMed  Google Scholar 

  5. Yehia L, Eng C. PTEN hamartoma tumor syndrome. In: Adam MP, Everman DB, Mirzaa GM, Pagon RA, Wallace SE, Bean LJ, et al., editors. GeneReviews®. Seattle, WA: University of Washington; 1993 http://www.ncbi.nlm.nih.gov/books/NBK1488/. Accessed 22 Dec 2022.

  6. Di Cristofano A, Pesco B, Cordon-Cardo C, Pandolfi PP. Pten is essential for embryonic development and tumour suppression. Nat. Genet. 1998;19. https://doi.org/10.1038/1235.

  7. Schwerd T, Khaled AV, Schürmann M, Chen H, Händel N, Reis A, et al. A recessive form of extreme macrocephaly and mild intellectual disability complements the spectrum of PTEN hamartoma tumour syndrome. Eur J Hum Genet. 2016;24:889–94.

    Article  PubMed  Google Scholar 

  8. Mighell TL, Thacker S, Fombonne E, Eng C, O’Roak BJ. An integrated deep-mutational-scanning approach provides clinical insights on PTEN genotype-phenotype relationships. Am J Hum Genet. 2020;106:818–29.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Yehia L, Ngeow J, Eng C. PTEN-opathies: from biological insights to evidence-based precision medicine. J Clin Investig. 2019;129:452–64.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Sarn N, Thacker S, Lee H, Eng C. Germline nuclear-predominant Pten murine model exhibits impaired social and perseverative behavior, microglial activation, and increased oxytocinergic activity. Mol Autism. 2021;12:41.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Skelton PD, Stan RV, Luikart BW. The role of PTEN in neurodevelopment. CXP. 2019;5:60–71.

    CAS  Google Scholar 

  12. Tilot AK, Bebek G, Niazi F, Altemus JB, Romigh T, Frazier TW, et al. Neural transcriptome of constitutional Pten dysfunction in mice and its relevance to human idiopathic autism spectrum disorder. Mol Psychiatry. 2016;21:118–25.

    Article  CAS  PubMed  Google Scholar 

  13. Shen WH, Balajee AS, Wang J, Wu H, Eng C, Pandolfi PP, et al. Essential role for nuclear PTEN in maintaining chromosomal integrity. Cell. 2007;128:157–70.

    Article  CAS  PubMed  Google Scholar 

  14. Chen ZH, Zhu M, Yang J, Liang H, He J, He S, et al. PTEN interacts with histone H1 and controls chromatin condensation. Cell Rep. 2014;8:2003–14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Groszer M, Erickson R, Scripture-Adams DD, Dougherty JD, Le Belle J, Zack JA, et al. PTEN negatively regulates neural stem cell self-renewal by modulating G0-G1 cell cycle entry. Proc Natl Acad Sci USA. 2006;103:111–6.

    Article  CAS  PubMed  Google Scholar 

  16. McEllin B, Camacho CV, Mukherjee B, Hahm B, Tomimatsu N, Bachoo RM, et al. PTEN loss compromises homologous recombination repair in astrocytes: implications for GBM therapy with temozolomide or PARP inhibitors. Cancer Res. 2010;70:5457–64.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ming M, Feng L, Shea CR, Soltani K, Zhao B, Han W, et al. PTEN positively regulates UVB-induced DNA damage repair. Cancer Res. 2011;71:5287–95.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Puc J, Keniry M, Li HS, Pandita TK, Choudhury AD, Memeo L, et al. Lack of PTEN sequesters CHK1 and initiates genetic instability. Cancer Cell. 2005;7:193–204.

    Article  CAS  PubMed  Google Scholar 

  19. Mighell TL, Evans-Dutson S, O’Roak BJ. A saturation mutagenesis approach to understanding PTEN lipid phosphatase activity and genotype-phenotype relationships. Am J Hum Genet. 2018;102:943–55.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. He X, Ni Y, Wang Y, Romigh T, Eng C. Naturally occurring germline and tumor-associated mutations within the ATP-binding motifs of PTEN lead to oxidative damage of DNA associated with decreased nuclear p53. Hum Mol Genet. 2011;20:80–89.

    Article  PubMed  Google Scholar 

  21. He X, Arrotta N, Radhakrishnan D, Wang Y, Romigh T, Eng C. Cowden syndrome-related mutations in PTEN associate with enhanced proteasome activity. Cancer Res. 2013;73:3029–40.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Bax M, McKenna J, Do-Ha D, Stevens CH, Higginbottom S, Balez R, et al. The ubiquitin proteasome system is a key regulator of pluripotent stem cell survival and motor neuron differentiation. Cells. 2019;8:E581.

    Article  Google Scholar 

  23. Hartwell LH, Weinert TA. Checkpoints: controls that ensure the order of cell cycle events. Science. 1989;246:629–34.

    Article  CAS  PubMed  Google Scholar 

  24. Kastan MB, Bartek J. Cell-cycle checkpoints and cancer. Nature. 2004;432:316–23.

    Article  CAS  PubMed  Google Scholar 

  25. Hitomi M, Yang K, Stacey AW, Stacey DW. Phosphorylation of cyclin D1 regulated by ATM or ATR controls cell cycle progression. Mol Cell Biol. 2008;28:5478–93.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Firsanov DV, Solovjeva LV, Svetlova MP. H2AX phosphorylation at the sites of DNA double-strand breaks in cultivated mammalian cells and tissues. Clin Epigenet. 2011;2:283–97.

    Article  CAS  Google Scholar 

  27. Cruz C, Castroviejo-Bermejo M, Gutiérrez-Enríquez S, Llop-Guevara A, Ibrahim YH, Gris-Oliver A, et al. RAD51 foci as a functional biomarker of homologous recombination repair and PARP inhibitor resistance in germline BRCA-mutated breast cancer. Ann Oncol. 2018;29:1203–10.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Mariotti LG, Pirovano G, Savage KI, Ghita M, Ottolenghi A, Prise KM, et al. Use of the γ-H2AX assay to investigate DNA repair dynamics following multiple radiation exposures. PLoS ONE. 2013;8:e79541.

    Article  PubMed  PubMed Central  Google Scholar 

  29. Yang J, Yu Y, Hamrick HE, Duerksen-Hughes PJ. ATM, ATR and DNA-PK: initiators of the cellular genotoxic stress responses. Carcinogenesis. 2003;24:1571–80.

    Article  CAS  PubMed  Google Scholar 

  30. So S, Davis AJ, Chen DJ. Autophosphorylation at serine 1981 stabilizes ATM at DNA damage sites. J Cell Biol. 2009;187:977–90.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Hirose Y, Katayama M, Mirzoeva OK, Berger MS, Pieper RO. Akt activation suppresses Chk2-mediated, methylating agent–induced G2 arrest and protects from temozolomide-induced mitotic catastrophe and cellular senescence. Cancer Res. 2005;65:4861–9.

    Article  CAS  PubMed  Google Scholar 

  32. Keane NA, Glavey SV, Krawczyk J, O’Dwyer M. AKT as a therapeutic target in multiple myeloma. Expert Opin Ther Targets. 2014;18:897–915.

    Article  CAS  PubMed  Google Scholar 

  33. Jackson MP, Hewitt EW. Cellular proteostasis: degradation of misfolded proteins by lysosomes. Essays Biochem. 2016;60:173–80.

    Article  PubMed  PubMed Central  Google Scholar 

  34. Smith IN, Thacker S, Jaini R, Eng C. Dynamics and structural stability effects of germline PTEN mutations associated with cancer versus autism phenotypes. J Biomol Struct Dyn. 2019;37:1766–82.

    Article  CAS  PubMed  Google Scholar 

  35. Kang SC, Jaini R, Hitomi M, Lee H, Sarn N, Thacker S, et al. Decreased nuclear Pten in neural stem cells contributes to deficits in neuronal maturation. Mol Psychiatry. 2020;11:43.

    CAS  Google Scholar 

  36. Tilot AK, Gaugler MK, Yu Q, Romigh T, Yu W, Miller RH, et al. Germline disruption of Pten localization causes enhanced sex-dependent social motivation and increased glial production. Hum Mol Genet. 2014;23:3212–27.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Papa A, Wan L, Bonora M, Salmena L, Song MS, Hobbs RM, et al. Cancer-associated PTEN mutants act in a dominant-negative manner to suppress PTEN protein function. Cell. 2014;157:595–610.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Yehia L, Seyfi M, Niestroj L-M, Padmanabhan R, Ni Y, Frazier TW, et al. Copy number variation and clinical outcomes in patients with germline PTEN mutations. JAMA Netw Open. 2020;3:e1920415.

    Article  PubMed  PubMed Central  Google Scholar 

  39. Markkanen E, Meyer U, Dianov GL. DNA damage and repair in schizophrenia and autism: implications for cancer comorbidity and beyond. Int J Mol Sci. 2016;17:856.

    Article  PubMed  PubMed Central  Google Scholar 

  40. Qing X, Zhang G, Wang Z-Q. DNA damage response in neurodevelopment and neuromaintenance. FEBS J. 2023;290:3300–10.

    Article  CAS  PubMed  Google Scholar 

  41. Wang M, Wei P-C, Lim CK, Gallina IS, Marshall S, Marchetto MC, et al. Increased Neural progenitor proliferation in a hiPSC model of autism induces replication stress-associated genome instability. Cell Stem Cell. 2020;26:221–233.e6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Nelen MR, Kremer H, Konings IB, Schoute F, Essen AJ, van, Koch R, et al. Novel PTEN mutations in patients with Cowden disease: absence of clear genotype–phenotype correlations. Eur J Hum Genet. 1999;7:267–73.

    Article  CAS  PubMed  Google Scholar 

  43. Yehia L, Ni Y, Sadler T, Frazier TW, Eng C. Distinct metabolic profiles associated with autism spectrum disorder versus cancer in individuals with germline PTEN mutations. npj Genom Med. 2022;7:1–9.

    Article  Google Scholar 

  44. Chen Y-H, Pruett-Miller SM. Improving single-cell cloning workflow for gene editing in human pluripotent stem cells. Stem Cell Res. 2018;31:186–92.

    Article  CAS  PubMed  Google Scholar 

  45. Hitomi M, Stacey DW. The checkpoint kinase ATM protects against stress-induced elevation of cyclin D1 and potential cell death in neurons. Cytometry A. 2010;77:524–33.

    Article  PubMed  Google Scholar 

  46. Hitomi M, Stacey DW. Cyclin D1 production in cycling cells depends on ras in a cell-cycle-specific manner. Curr Biol. 1999;9:1075–84.

    Article  CAS  PubMed  Google Scholar 

  47. Chumakova AP, Hitomi M, Sulman EP, Lathia JD. High-throughput automated single-cell imaging analysis reveals dynamics of glioblastoma stem cell population during state transition. Cytometry A. 2019;95:290–301.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We would like to thank Washington University Genome Engineering and hiPSC Core (GEiC) facility for preparing gene-edited human induced pluripotent stem cells used in this study. This study was supported, in part, by the Lisa Dean Moseley Foundation Grant (to CE and MH) and the Ambrose Monell Foundation (PTEN Switch grant to CE). CE is the Sondra J. and Stephen R. Hardis Endowed Chair of Cancer Genomic Medicine at the Cleveland Clinic, and an ACS Clinical Research Professor.

Author information

Authors and Affiliations

  1. Genomic Medicine Institute, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, 44195, USA

    Masahiro Hitomi, Juan Venegas, Shin Chung Kang & Charis Eng

  2. Department of Molecular Medicine, The Cleveland Clinic Lerner College of Medicine of Case Western Reserve University, Cleveland, OH, 44195, USA

    Masahiro Hitomi & Charis Eng

  3. Center for Personalized Genetic Healthcare, Medical Specialties Institute, Cleveland Clinic, Cleveland, OH, 44195, USA

    Charis Eng

  4. Department of Genetics and Genome Science, Case Western Reserve University School of Medicine, Cleveland, OH, 44116, USA

    Charis Eng

  5. Case Comprehensive Cancer Center, Case Western Reserve University School of Medicine, Cleveland, OH, 44116, USA

    Charis Eng

Authors
  1. Masahiro Hitomi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Juan Venegas
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shin Chung Kang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Charis Eng
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

CE was responsible for conception and design of the entire study, interpretation of data, and drafting and critical revision of the manuscript. MH was responsible for study design, experimental execution, data acquisition, data analysis, data interpretation, and drafting the manuscript. JV was responsible for designing the project and critical review and revision of the manuscript. SCK was responsible for data acquisition and analysis and reviewing the manuscript. All authors approved of the final manuscript.

Corresponding author

Correspondence to Charis Eng.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Hitomi, M., Venegas, J., Kang, S.C. et al. Differential cell cycle checkpoint evasion by PTEN germline mutations associated with dichotomous phenotypes of cancer versus autism spectrum disorder. Oncogene 42, 3698–3707 (2023). https://doi.org/10.1038/s41388-023-02867-4

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41388-023-02867-4

Search

Advanced search

Quick links