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:

Phase separation of TAZ compartmentalizes the transcription machinery to promote gene expression

Nature Cell Biology volume 22, pages 453–464 (2020)Cite this article

Subjects

Abstract

TAZ promotes growth, development and tumorigenesis by regulating the expression of target genes. However, the manner in which TAZ orchestrates the transcriptional responses is poorly defined. Here we demonstrate that TAZ forms nuclear condensates through liquid–liquid phase separation to compartmentalize its DNA-binding cofactor TEAD4, coactivators BRD4 and MED1, and the transcription elongation factor CDK9 for transcription. TAZ forms phase-separated droplets in vitro and liquid-like nuclear condensates in vivo, and this ability is negatively regulated by Hippo signalling through LATS-mediated phosphorylation and is mediated by the coiled-coil (CC) domain. Deletion of the TAZ CC domain or substitution with the YAP CC domain prevents the phase separation of TAZ and its ability to induce the expression of TAZ-specific target genes. Thus, we identify a mechanism of transcriptional activation by TAZ and demonstrate that pathway-specific transcription factors also engage the phase-separation mechanism for efficient and specific transcriptional activation.

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: TAZ undergoes LLPS in vitro and in vivo.
Fig. 2: TAZ nuclear condensates display liquid-like properties.
Fig. 3: The WW domain and CC domain are required for TAZ LLPS.
Fig. 4: The differential ability of TAZ and YAP to undergo phase separation lies in the CC domain.
Fig. 5: Hippo signalling negatively regulates TAZ phase separation.
Fig. 6: Hippo signalling negatively regulates TAZ phase separation through LATS2.
Fig. 7: TAZ compartmentalizes TEAD and other transcriptional factors to the nuclear puncta.
Fig. 8: Phase separation of TAZ promotes transcription.

Similar content being viewed by others

Data availability

Source data for Figs. 1–4 and 6–8 and Extended Data Figs. 1, 3, 4 and 8 are available online. The RNA-seq data are available in the Gene Expression Omnibus (GEO) with the accession number GSE142474. All other data supporting the findings of this study are available from the corresponding author on reasonable request.

References

  1. Pan, D. The Hippo signaling pathway in development and cancer. Dev. Cell 19, 491–505 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Ma, S., Meng, Z., Chen, R. & Guan, K. L. The Hippo pathway: biology and pathophysiology. Annu. Rev. Biochem. 88, 577–604 (2018).

    Article  CAS  Google Scholar 

  3. Tapon, N. & Harvey, K. F. The Hippo pathway—from top to bottom and everything in between. Semin. Cell Dev. Biol. 23, 768–769 (2012).

    Article  PubMed  Google Scholar 

  4. Badouel, C. & McNeill, H. SnapShot: the Hippo signaling pathway. Cell 145, 484 (2011).

    Article  CAS  PubMed  Google Scholar 

  5. Schroeder, M. C. & Halder, G. Regulation of the Hippo pathway by cell architecture and mechanical signals. Semin. Cell Dev. Biol. 23, 803–811 (2012).

    Article  CAS  PubMed  Google Scholar 

  6. Yu, F. X. & Guan, K. L. The Hippo pathway: regulators and regulations. Genes Dev. 27, 355–371 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Dong, J. X. et al. Elucidation of a universal size-control mechanism in Drosophila and mammals. Cell 130, 1120–1133 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Ota, M. & Sasaki, H. Mammalian Tead proteins regulate cell proliferation and contact inhibition as transcriptional mediators of Hippo signaling. Development 135, 4059–4069 (2008).

    Article  CAS  PubMed  Google Scholar 

  9. Enderle, L. & McNeill, H. Hippo gains weight: added insights and complexity to pathway control. Sci. Signal. 6, re7 (2013).

  10. Shreberk-Shaked, M. & Oren, M. New insights into YAP/TAZ nucleo-cytoplasmic shuttling: new cancer therapeutic opportunities? Mol. Oncol. 13, 1335–1341 (2019).

    PubMed  PubMed Central  Google Scholar 

  11. Galli, G. G. et al. YAP drives growth by controlling transcriptional pause release from dynamic enhancers. Mol. Cell 60, 328–337 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Zanconato, F. et al. Transcriptional addiction in cancer cells is mediated by YAP/TAZ through BRD4. Nat. Med. 24, 1599–1610 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Boija, A. et al. Transcription factors activate genes through the phase-separation capacity of their activation domains. Cell 175, 1842–1855 (2018).

    Article  CAS  PubMed  Google Scholar 

  14. Chan, S. W. et al. A role for TAZ in migration, invasion, and tumorigenesis of breast cancer cells. Cancer Res. 68, 2592–2598 (2008).

    Article  CAS  PubMed  Google Scholar 

  15. Cordenonsi, M. et al. The Hippo transducer TAZ confers cancer stem cell-related traits on breast cancer cells. Cell 147, 759–772 (2011).

    Article  CAS  PubMed  Google Scholar 

  16. Plouffe, S. W. et al. The Hippo pathway effector proteins YAP and TAZ have both distinct and overlapping functions in the cell. J. Biol. Chem. 293, 11230–11240 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Xin, M. et al. Hippo pathway effector Yap promotes cardiac regeneration. Proc. Natl Acad. Sci. USA 110, 13839–13844 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  18. Nishioka, N. et al. The Hippo signaling pathway components Lats and Yap pattern Tead4 activity to distinguish mouse trophectoderm from inner cell mass. Dev. Cell 16, 398–410 (2009).

    Article  CAS  PubMed  Google Scholar 

  19. Morin-Kensicki, E. M. et al. Defects in yolk sac vasculogenesis, chorioallantoic fusion, and embryonic axis elongation in mice with targeted disruption of Yap65. Mol. Cell. Biol. 26, 77–87 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Hossain, Z. et al. Glomerulocystic kidney disease in mice with a targeted inactivation of Wwtr1. Proc. Natl Acad. Sci. USA 104, 1631–1636 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Passaniti, A., Brusgard, J. L., Qiao, Y. T., Sudol, M. & Finch-Edmondson, M. Roles of RUNX in Hippo pathway signaling. Adv. Exp. Med. Biol. 962, 435–448 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Zhao, B. et al. TEAD mediates YAP-dependent gene induction and growth control. Gene Dev. 22, 1962–1971 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Zhao, B., Tumaneng, K. & Guan, K. L. The Hippo pathway in organ size control, tissue regeneration and stem cell self-renewal. Nat. Cell Biol. 13, 877–883 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Alberti, S., Gladfelter, A. & Mittag, T. Considerations and challenges in studying liquid-liquid phase separation and biomolecular condensates. Cell 176, 419–434 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Sabari, B. R. et al. Coactivator condensation at super-enhancers links phase separation and gene control. Science 361, eaar3958 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Lu, H., Liu, R. & Zhou, Q. Balanced between order and disorder: a new phase in transcription elongation control and beyond. Transcription 10, 157–163 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Kaan, H. Y. K. et al. Crystal structure of TAZ-TEAD complex reveals a distinct interaction mode from that of YAP-TEAD complex. Sci. Rep. 7, 2035 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Piccolo, S., Dupont, S. & Cordenonsi, M. The biology of Yap/Taz: Hippo signaling and beyond. Physiol. Rev. 94, 1287–1312 (2014).

    Article  CAS  PubMed  Google Scholar 

  29. Hyman, A. A., Weber, C. A. & Juelicher, F. Liquid-liquid phase separation in biology. Annu. Rev. Cell Dev. Biol. 30, 39–58 (2014).

    Article  CAS  PubMed  Google Scholar 

  30. Boeynaems, S. et al. Protein phase separation: a new phase in cell biology. Trends Cell Biol. 28, 420–435 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Cai, D. et al. Phase separation of YAP reorganizes genome topology for long-term YAP target gene expression. Nat. Cell Biol. 21, 1578–1589 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Henis, Y. I., Rotblat, B. & Kloog, Y. FRAP beam-size analysis to measure palmitoylation-dependent membrane association dynamics and microdomain partitioning of Ras proteins. Methods 40, 183–190 (2006).

    Article  CAS  PubMed  Google Scholar 

  33. Eisenberg, S., Giehl, K., Henis, Y. I. & Ehrlich, M. Differential interference of chlorpromazine with the membrane interactions of oncogenic K-Ras and its effects on cell growth. J. Biol. Chem. 283, 27279–27288 (2008).

    Article  CAS  PubMed  Google Scholar 

  34. Shvartsman, D. E. et al. Src kinase activity and SH2 domain regulate the dynamics of Src association with lipid and protein targets. J. Cell Biol. 178, 675–686 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Wolfenson, H. et al. A role for the juxtamembrane cytoplasm in the molecular dynamics of focal adhesions. PLoS ONE 4, e4304 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Molliex, A. et al. Phase separation by low complexity domains promotes stress granule assembly and drives pathological fibrillization. Cell 163, 123–133 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Nott, T. J. et al. Phase transition of a disordered nuage protein generates environmentally responsive membraneless organelles. Mol. Cell 57, 936–947 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Fan, R., Kim, N. G. & Gumbiner, B. M. Regulation of Hippo pathway by mitogenic growth factors via phosphoinositide 3-kinase and phosphoinositide-dependent kinase-1. Proc. Natl Acad. Sci. USA 110, 2569–2574 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  39. Kim, N. G. & Gumbiner, B. M. Adhesion to fibronectin regulates Hippo signaling via the FAK-Src-PI3K pathway. J. Cell Biol. 210, 503–515 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Totaro, A., Panciera, T. & Piccolo, S. YAP/TAZ upstream signals and downstream responses. Nat. Cell Biol. 20, 888–899 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Yu, F. X. et al. Regulation of the Hippo-YAP pathway by G-protein-coupled receptor signaling. Cell 150, 780–791 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Panciera, T., Azzolin, L., Cordenonsi, M. & Piccolo, S. Mechanobiology of YAP and TAZ in physiology and disease. Nat. Rev. Mol. Cell Biol. 18, 758–770 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Hergovich, A. The roles of NDR protein kinases in Hippo signalling. Genes 7, 21 (2016).

    Article  CAS  PubMed Central  Google Scholar 

  44. Chan, S. W. et al. TEADs mediate nuclear retention of TAZ to promote oncogenic transformation. J. Biol. Chem. 284, 14347–14358 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Noland, C. L. et al. Palmitoylation of TEAD transcription factors is required for their stability and function in Hippo pathway signaling. Structure 24, 179–186 (2016).

    Article  CAS  PubMed  Google Scholar 

  46. Zhang, H. et al. TEAD transcription factors mediate the function of TAZ in cell growth and epithelial-mesenchymal transition. J. Biol. Chem. 284, 13355–13362 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Rausch, V. et al. The Hippo pathway regulates caveolae expression and mediates flow response via caveolae. Curr. Biol. 29, 242–255 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Misra, J. R. & Irvine, K. D. The Hippo signaling network and its biological functions. Annu. Rev. Genet. 52, 65–87 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Zhang, L., Yue, T. & Jiang, J. Hippo signaling pathway and organ size control. Fly 3, 68–73 (2009).

    Article  PubMed  Google Scholar 

  50. Gill, M. K. et al. A feed forward loop enforces YAP/TAZ signaling during tumorigenesis. Nat. Commun. 9, 3510 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Richardson, H. E. & Portela, M. Tissue growth and tumorigenesis in Drosophila: cell polarity and the Hippo pathway. Curr Opin. Cell Biol. 48, 1–9 (2017).

    Article  CAS  PubMed  Google Scholar 

  52. Zeng, M. et al. Phase transition in postsynaptic densities underlies formation of synaptic complexes and synaptic plasticity. Cell 166, 1163–1175 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Fang, X. et al. Arabidopsis FLL2 promotes liquid-liquid phase separation of polyadenylation complexes. Nature 569, 265–269 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Varelas, X. et al. TAZ controls Smad nucleocytoplasmic shuttling and regulates human embryonic stem-cell self-renewal. Nat. Cell Biol. 10, 837–848 (2008).

    Article  CAS  PubMed  Google Scholar 

  55. Zhang, J. M. et al. YAP-dependent induction of amphiregulin identifies a non-cell-autonomous component of the Hippo pathway. Nat. Cell Biol. 11, 1444–1450 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Kwon, I. et al. Phosphorylation-regulated binding of rna polymerase ii to fibrous polymers of low-complexity domains. Cell 155, 1049–1060 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Kato, M. et al. Cell-free formation of RNA granules: low complexity sequence domains form dynamic fibers within hydrogels. Cell 149, 753–767 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Hagenbeek, T. J. et al. The Hippo pathway effector TAZ induces TEAD-dependent liver inflammation and tumors. Sci. Signal. 11, eaaj1757 (2018).

    Article  CAS  PubMed  Google Scholar 

  59. Lu, H. et al. Phase-separation mechanism for C-terminal hyperphosphorylation of RNA polymerase II. Nature 558, 318–323 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Plouffe, S. W. et al. Characterization of Hippo pathway components by gene inactivation. Mol. Cell 64, 993–1008 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Zhu, Q. et al. SnoN antagonizes the Hippo kinase complex to promote TAZ signaling during breast carcinogenesis. Dev. Cell 37, 399–412 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Eisenberg, S. et al. Raft protein clustering alters N-Ras membrane interactions and activation pattern. Mol. Cell. Biol 31, 3938–3952 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Efron, B. & Tibshirani, R. in An Introduction to Bootstrap (eds Cox, D. R. et al.) 124–130 (Chapman & Hall, 1993).

  64. Rashidian, J. et al. Ski regulates Hippo and TAZ signaling to suppress breast cancer progression. Sci. Signal. 8, ra14 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).

    Article  CAS  PubMed  Google Scholar 

  66. Anders, S., Pyl, P. T. & Huber, W. HTSeq–a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166–169 (2015).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank K.-L. Guan and A. Mauviel for providing cDNAs of components of the Hippo pathway and H. Sasaki for the 8xGT-IIC-δ51LucII construct; J. He for technical assistance and Q. Zhu for helpful suggestions, discussions and help with experimental procedures; and D. Schichnes and S. Ruzin at the CNR biological imaging facility at the University of California, Berkeley for assistance with microscopy. This study was supported by DOD/US Army Medical Research And Materiel Command W81XWH-15-1-0068 (to K.L. and Q.Z.), a Tel Aviv University-University of California Berkeley collaborative research grant (to Y.I.H. and K.L.), and NIH R01AI41757 (to Q.Z.). Y.I.H. is an incumbent of the Zalman Weinberg Chair in Cell Biology. Y.L. is supported by the Berkeley Scholars program, and T.W. was supported by the China Scholarship Council.

Author information

Author notes

  1. These authors contributed equally: Yi Lu, Tiantian Wu.

Authors and Affiliations

  1. Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, USA

    Yi Lu, Tiantian Wu, Huasong Lu, Qiang Zhou & Kunxin Luo

  2. State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, China

    Tiantian Wu

  3. Department of Neurobiology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel

    Orit Gutman & Yoav I. Henis

Authors
  1. Yi Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tiantian Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Orit Gutman
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Huasong Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qiang Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yoav I. Henis
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Kunxin Luo
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Y.L., T.W. and K.L. designed the research. Y.L. performed in vivo experiments. T.W. performed in vitro experiments. Y.I.H. designed and O.G. performed FRAP experiments. Y.L., T.W., H.L., Y.I.H., Q.Z. and K.L. analysed data and wrote the paper. K.L. conceived and directed the project. All of the authors discussed the results and commented on the manuscript.

Corresponding author

Correspondence to Kunxin Luo.

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.

Extended data

Extended Data Fig. 1 Regulation of TAZ droplet formation in vitro and nuclear puncta formation in vivo.

a, GFP-TAZ purified from E. coil were analysed by SDS-PAGE and visualized by Coomassie blue staining. b. 50 μM GFP-TAZ were heated-inactivated (5 min at 95 °C and immediately put on ice for 5 min) or treated with 100 μg/ml Proteinase K for 30 min at 40 °C, and then subjected to droplet formation assay in vitro in the presence of 500 mM NaCl at room temperature. c, Ectopically expressed GFP-TAZ was expressed at a lower level than endogenous TAZ in MCF-10A cells as shown by western blotting. GAPDH was used as a loading control. d, Flag-TAZ formed nuclear puncta when transfected into the MCF-10A cells, as detected by immunofluorescence staining with anti-Flag. Scale bar, 10 μm. Experiments in a–d were repeated independently three times with similar results. Unprocessed blots are provided in Unprocessed Blots Extended Data Fig. 1.

Source data

Extended Data Fig. 2 YAP does not form droplets in vitro and in vivo in the absence of crowding agents.

a, GFP-YAP purified from E. coil were analyzed by SDS-PAGE and visualized by Coomassie blue staining. b, GFP-YAP at varying concentrations was subjected to the droplet formation assay at room temperature and in the presence of 500 mM NaCl. c, 50 μM GFP-YAP was subjected to the droplet formation assay at room temperature in the presence of indicated salt concentrations. d, 50 μM GFP-YAP was subjected to droplet formation in the presence of 150 mM NaCl at 4 °C or 37 °C. e, Two YAP isoforms, GFP-YAP1–1β or GFP-YAP1–2α, did not form droplets (50 μM protein, 500 mM NaCl and room temperature). aa, amino acids. f, 50 μM GFP-YAP formed droplets in the presence of 10% PEG-8000, Ficoll or Dextran but not 10% glycerol or sucrose. Droplet formation assay was performed in the presence of 500 mM NaCl at room temperature. g, 50 μM GFP-YAP did not form droplets in the presence of BSA at varying concentrations. h, GFP-YAP did not form nuclear puncta in both HeLa cells and 293T cells. Scale bars, 10 μm. Experiments in a–h were repeated independently three times with similar results.

Extended Data Fig. 3 The CC and WW domains are required for TAZ to form nuclear puncta.

a, Domain structure of TAZ and TAZ truncations. The numbers above indicate the position of amino acid residues. b, Bacterially purified GFP-TAZ, ∆TB, ∆WW, ∆CC, and ∆WW+∆CC proteins were analyzed by SDS-PAGE and detected by Coomasssie blue staining. c, Localization of GFP-TAZ and various mutants in HeLa cells. d, Localization of GFP-TAZ and various TAZ/YAP chimera in HeLa cells. Scale bars, 10 μm. e, A GST pull-down assay was performed by incubating immobilized GST fusion proteins with lysates of cells expressing HA-tagged WT or mutant TAZ, and the associated TAZ proteins were detected by western blotting with anti-HA (upper). GST fusion proteins were assessed by western blotting with anti-GST, and HA-TAZ proteins in the cell lysates were measured by western blotting (lower). Experiments in b–e were repeated independently three times with similar results. Unprocessed blots are provided in Unprocessed Blots Extended Data Fig. 3.

Source data

Extended Data Fig. 4 TAZ CC domain enhances YAP phase separation in the presence of PEG.

a, Domain structure of YAP chimera. b, Substitution of the YAP CC and WW domains with that of TAZ is not sufficient to enable YAP to undergo LLPS in MCF10A cells in the absence of PEG. c, Coomasssie blue staining of various recombinant proteins purified from E. coil. d, 25 μM bacterially purified GFP-YAP chimera proteins were subjected to droplet formation assay in the presence of 10% PEG-8000. Quantification of the droplets is on the right. Scale bar, 10 μm. Data shown as the mean ± s.e.m. Statistical significance was evaluated using One-way ANOVA with Krusk-Wallis test. Droplets in n = 3 fields in each group were quantified. e, The TAZ CC and WW domains enhanced LLPS by GFP-YAP in transfected MCF10A cells in the presence of PEG as shown by confocal microscopy. Scale bar, 10 μm. Quantification of the percentage of cells that displayed nuclear puncta is shown on the right. Data shown as the mean ± s.e.m.. P value was determined by unpaired two-tailed Student’s t-test. 80 transfected cells in each group were quantified. n = 3 biologically independent samples. Experiments in b, c, e were repeated independently three times with similar results. Experiments in d were repeated twice with similar results. Statistical source data for d, e, are provided in Statistical Source Date Extended Data Fig. 4.

Source data

Extended Data Fig. 5 Hippo signaling negatively regulates TAZ phase separation in HeLa cells.

TAZ localization was examined by immunofluorescence staining with anti-TAZ (green) in HeLa cells that have been subjected to the following treatments: a, Serum-starved HeLa cells were treated with 1 μM LPA or 50 ng/ml EGF for 1 h. b, Serum-starved HeLa cells were seeded on fibronectin-coated coverslips for 10 min or 2 h in serum-free medium. c, HeLa cells were grown on fibronectin-coated polyacrylamide hydrogels of 1 kPa and 40 kPa stiffness. d, HeLa cells were treated with 1 μg/ml Latrunculin B for 1 h. Alexa Fluor 555-conjugated phalloidin (Red) staining was performed to detect F-actin in b-d. Scale bar, 10 µm. Experiments in a–d were repeated independently three times with similar results.

Extended Data Fig. 6 LATS2 regulates TAZ LLPS and recruitment of TEAD4 and BRD4.

a, MCF-10A cells transfected with GFP-TAZ-S89A and Flag-LATS2 were subjected to immunofluorescence staining with anti-Flag (Red). Scale bar, 10 µm. b, MCF-10A cells stably expressing siLATS1/2 were transfected with GFP-TAZ and Flag-TEAD4. TEAD localization at high cell density was detected by immunofluorescence staining with anti-Flag (Red). Scale bar, 10 μm. c, MCF-10A cells stably expressing siLATS1/2 were transfected with GFP-TAZ. Endogenous BRD4 localization was examined by immunofluorescence staining with anti-BRD4 (Red). Scale bar, 10 μm. All experiments were repeated independently three times with similar results.

Extended Data Fig. 7 TAZ nuclear condensates do not co-localize with the PML bodies, Cajal bodies or nucleoli.

The PML nuclear bodies, Cajal Bodies and nucleoli in MCF-10A cells expressing GFP-TAZ (green) were detected by immunofluorescence staining with antibodies targeting PML, Coilin and Fibrillarin, respectively (red). Scale bar, 10 μm. Experiments were repeated independently three times with similar results.

Extended Data Fig. 8 TAZ mutants lacking the CC domain still bind to LAST2 and TEAD4.

a, HA-tagged WT and mutant TAZ were co-transfected into 293T cells with Flag-LATS2. TAZ proteins associated with LATS2 were isolated by immunoprecipitation with anti-Flag and detected by western blotting with anti-HA antibodies (upper panels). The abundance of these proteins in the cell lysates was assessed by western blotting (lower panels). GAPDH was used as a loading control. b, Interaction of various TAZ mutants with Flag-TEAD4 was analyzed by co-IP assay as described in a. c, Interaction of various TAZ/YAP chimera with LATS2 was analyzed by co-IP as described in a. All experiments were repeated independently three times with similar results. Unprocessed blots are provided in Unprocessed Blots Extended Data Fig. 8.

Source data

Supplementary information

Source data

Source Data Fig. 1

Statistical source data.

Source Data Fig. 2

Statistical source data.

Source Data Fig. 3

Unprocessed blots.

Source Data Fig. 3

Statistical source data.

Source Data Fig. 4

Unprocessed blots.

Source Data Fig. 4

Statistical source data.

Source Data Fig. 6

Unprocessed blots.

Source Data Fig. 6

Statistical source data.

Source Data Fig. 7

Unprocessed blots.

Source Data Fig. 8

Unprocessed blots.

Source Data Fig. 8

Statistical source data.

Source Data Extended Data Fig. 1

Unprocessed blots.

Source Data Extended Data Fig. 3

Unprocessed blots.

Source Data Extended Data Fig. 4

Statistical source data.

Source Data Extended Data Fig. 8

Unprocessed blots.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Lu, Y., Wu, T., Gutman, O. et al. Phase separation of TAZ compartmentalizes the transcription machinery to promote gene expression. Nat Cell Biol 22, 453–464 (2020). https://doi.org/10.1038/s41556-020-0485-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41556-020-0485-0

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing