Article | Published:

STRIPAK components determine mode of cancer cell migration and metastasis

Nature Cell Biology volume 17, pages 6880 (2015) | Download Citation

Abstract

The contractile actomyosin cytoskeleton and its connection to the plasma membrane are critical for control of cell shape and migration. We identify three STRIPAK complex components, FAM40A, FAM40B and STRN3, as regulators of the actomyosin cortex. We show that FAM40A negatively regulates the MST3 and MST4 kinases, which promote the co-localization of the contractile actomyosin machinery with the Ezrin/Radixin/Moesin family proteins by phosphorylating the inhibitors of PPP1CB, PPP1R14A–D. Using computational modelling, in vitro cell migration assays and in vivo breast cancer metastasis assays we demonstrate that co-localization of contractile activity and actin–plasma membrane linkage reduces cell speed on planar surfaces, but favours migration in confined environments similar to those observed in vivo. We further show that FAM40B mutations found in human tumours uncouple it from PP2A and enable it to drive a contractile phenotype, which may underlie its role in human cancer.

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Acknowledgements

C.D.M., S.H., M.T., A.B., G.F., P.A.B., B.T. and E.S. are financially supported by Cancer Research UK. C.D.M. was further supported by a FEBS long-term fellowship. We thank laboratory members for help and advice throughout this work. We thank N. O’Reilly and S. Kjaer for help with peptide synthesis and protein purification, and members of the BRU for help with metastasis assays.

Author information

Affiliations

  1. Tumour Cell Biology Laboratory, Cancer Research UK London Research Institute, 44 Lincoln’s Inn Fields London WC2A 3LY, UK

    • Chris D. Madsen
    • , Steven Hooper
    •  & Erik Sahai
  2. Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5 2200 Copenhagen N, Denmark

    • Chris D. Madsen
    •  & Janine T. Erler
  3. Biomolecular Modelling Laboratory, Cancer Research UK London Research Institute, 44 Lincoln’s Inn Fields London WC2A 3LY, UK

    • Melda Tozluoglu
    •  & Paul A. Bates
  4. Lymphocyte Interaction Laboratory, Cancer Research UK London Research Institute, 44 Lincoln’s Inn Fields London WC2A 3LY, UK

    • Andreas Bruckbauer
  5. Epithelial Biology Laboratory, Cancer Research UK London Research Institute, 44 Lincoln’s Inn Fields London, WC2A 3LY, UK

    • Georgina Fletcher
    •  & Barry Thompson

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Contributions

C.D.M. and E.S. carried out all experiments except those noted otherwise. S.H. performed all kinase assays and immunoprecipitation assays. G.F. and B.T. performed the fly screen. M.T. and P.A.B. made the mathematical model. A.B. helped with structured illumination microscopy. C.D.M. and E.S. conceived the study and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Erik Sahai.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    Supplementary Information

Excel files

  1. 1.

    Supplementary Table 1: Information of the siRNA screen's.

    Sheet 1. Fly screen. List of genes that were depleted in the fly screen. Sheet 2. Fly genes and human homologous. Genes with similarities are also included. Sheet 3. siRNA sequences and catalogue number of all siRNA used in the screen. All siRNA's were purchased by Dharmacon. Sheet 4. Fly and human gene names, fly and A431 phenotypes.

  2. 2.

    Supplementary Table 2: Peptide kinase screen.

    Peptide sequences and specific phospho site are shown. Raw data from each experiment is shown.

  3. 3.

    Supplementary Table 3: qPCR primers and siRNA oligo's.

    qPCR primer sequences and siRNAs used in the study including catalog numbers.

  4. 4.

    Supplementary Table 4: Expression vectors.

    All expression vectors used in this study are shown.

  5. 5.

    Supplementary Table 5: Antibodies.

    All antibodies used in this study including provider, catalog numbers and dilutions are shown.

Videos

  1. 1.

    3D morphologies of siRNA depleted A431 cells.

    3D reconstruction of confocal stacks taken of siRNA transfected A431 cells stained for F-actin (red) and pS19-MLC (green). The cells were plated on top of collagen-1/matrigels. The movie includes siCtr, siFAM40A, siFAM40B and STRN3 depleted A431 cells sequentially.

  2. 2.

    Spatiotemporal regulation of MST3-GFP.

    Confocal time lapse movie of siRNA transfected A431-MST3-GFP cells. The cells have been serum starved for 24 h and then stimulated with FBS. Imaging is then initiated immediately and frames are taken every 20 s. When cells were treated with ROCK inhibitor (Y27632) the drug was added during serum starvation. The movie includes siCtr, ROCK inhibitor (Y27632) treated, and siCCM3 depleted A431-MST3-GFP cells sequentially.

  3. 3.

    Time-lapse movie of siRNA depleted MDA-MD231 cells on hard surfaces.

    Phase contrast time lapse movie of siRNA transfected MDA-MB-231 cells plated on a 2D planar surface. Images were taken every 5 min. The movie includes siCtr, siFAM40A, siFAM40B and siMST3&4 depleted MDA-MB-231 cells sequentially.

  4. 4.

    Time-lapse movie of siRNA depleted MDA-MD231 cells on soft surfaces.

    Phase contrast time lapse movie of siRNA transfected MDA-MB-231 cells plated on top of collagen-I/matrigels. Images were taken every 5 min. The movie includes siCtr, siFAM40A, siFAM40B and siMST3&4 depleted MDA-MB-231 cells sequentially.

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DOI

https://doi.org/10.1038/ncb3083

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