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Identification of a novel alternative splicing isoform of the Hippo kinase STK3/MST2 with impaired kinase and cell growth suppressing activities

Oncogene (2024)Cite this article

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Abstract

Mammalian Ste-20-like Kinases 1 and 2 (MST1/2) are core serine-threonine kinases of the Hippo pathway regulating several cellular processes, including cell cycle arrest and cell death. Here, we discovered a novel alternative splicing variant of the MST2 encoding gene, STK3, in malignant cells and tumor datasets. This variant, named STK3∆7 or MST2∆7 (for mRNA or protein, respectively), resulted from the skipping of exon 7. MST2∆7 exhibited increased ubiquitylation and interaction with the E3 ubiquitin-protein ligase CHIP compared to the full-length protein (MST2FL). Exon 7 in STK3 encodes a segment within the kinase domain, and its exclusion compromised MST2 interaction with and phosphorylation of MOB, a major MST1/2 substrate. Nevertheless, MST2∆7 was capable of interacting with MST1 and MST2FL. Unlike MST2FL, overexpression of MST2∆7 did not lead to increased cell death and growth arrest. Strikingly, we observed the exclusion of STK3 exon 7 in 3.2–15% of tumor samples from patients of several types of cancer, while STK3∆7 was seldomly found in healthy tissues. Our study identified a novel STK3 splicing variant with loss of function and the potential to disturb tissue homeostasis by impacting on MST2 activities in the regulation of cell death and quiescence.

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Fig. 1: Identification of a new variant of STK3.
Fig. 2: MST2Δ7 is more ubiquitylated than MST2FL and interacts with CHIP-E3 ligase.
Fig. 3: MST2Δ7 overexpression resulted in decreased MOB1 phosphorylation.
Fig. 4: Cells overexpressing MST2Δ7 lose their ability to trigger cell death and quiescence.
Fig. 5: STK3Δ7 is expressed in tumor samples and tumor cell lines and displays potential clinical implications.

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Data availability

All data related to this study are available in the main figures and supplementary material.

Code availability

Code used for image quantification can be found at https://github.com/brunicardoso/e-signal-lab. Code used for bioinformatics analysis can be made available upon request.

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Acknowledgements

The authors are grateful to Antonio Manucci, Beatriz Moraes, Eduardo Moraes Rego Reis, and Nicolas Hoch for scientific discussions, advice on experiments and insightful inputs. The authors also wish to thank Celia Ludio for technical assistance.

Funding

This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP #2014/10492–0 and #2019/26767–2). RT was initially funded by Instituto Serrapilheira and subsequently by a CAPES postdoctoral fellowship (88882.315500/2019–01). APZPF was a recipient of FAPESP postdoctoral fellowship (#2014/25832–1). AMR was supported by a PhD scholarship from CAPES (88882.332986/2019–01).

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Authors and Affiliations

  1. Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, Brazil

    Ana Maria Rodrigues, Ana Paula Zen Petisco Fiore, Rebeka Tomasin, André Azevedo Reis Teixeira, Ricardo Jose Giordano, Deborah Schechtman & Alexandre Bruni-Cardoso

  2. Department of Biology, New York University, New York, NY, USA

    Ana Paula Zen Petisco Fiore

  3. Centro de Oncologia Molecular, Hospital Sírio-Libanês, São Paulo, Brazil

    Gabriela D. A. Guardia & Pedro A. F. Galante

  4. Department of Biochemistry and Molecular Pharmacology, Howard Hughes Medical Institute, New York University Grossman School of Medicine, New York, NY, USA

    Michele Pagano

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Contributions

AMR, APZPF, and AB-C conceptualized the work; AMR. APZPF, DS, RT, RJG, AB-C, and MP designed the experiments; PAFG and GADG designed and performed bioinformatics analysis. AMR, APZPF, RT, and AAT performed experiments; AMR, AB-C, and APZPF were responsible for data curation; AB-C and AMR wrote the manuscript; AB-C supervised the work; AB-C, MP, and DS acquired funding to support this work.

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Correspondence to Alexandre Bruni-Cardoso.

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The authors declare no competing interests.

Ethics approval and consent to participate

Our research involved human material and human data used in accordance with the Declaration of Helsinki. The RNA samples utilized in the analysis presented in Fig. 5F were extracted from patient samples stored in the Biobank of the São Paulo Cancer Institute Octavio Frias de Oliveira (ICESP). Informed consent was obtained from all subjects. To use these samples, we obtained approval from the National Ethics Committee in Brazil (Comissão Nacional de Ética em Pesquisa; CAAE: 06373719.0.3001.0065).

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Rodrigues, A.M., Paula Zen Petisco Fiore, A., Guardia, G.D.A. et al. Identification of a novel alternative splicing isoform of the Hippo kinase STK3/MST2 with impaired kinase and cell growth suppressing activities. Oncogene (2024). https://doi.org/10.1038/s41388-024-03104-2

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