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Published May 16, 2019 | Published + Supplemental Material
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WASH phosphorylation balances endosomal versus cortical actin network integrities during epithelial morphogenesis


Filamentous actin (F-actin) networks facilitate key processes like cell shape control, division, polarization and motility. The dynamic coordination of F-actin networks and its impact on cellular activities are poorly understood. We report an antagonistic relationship between endosomal F-actin assembly and cortical actin bundle integrity during Drosophila airway maturation. Double mutants lacking receptor tyrosine phosphatases (PTP) Ptp10D and Ptp4E, clear luminal proteins and disassemble apical actin bundles prematurely. These defects are counterbalanced by reduction of endosomal trafficking and by mutations affecting the tyrosine kinase Btk29A, and the actin nucleation factor WASH. Btk29A forms protein complexes with Ptp10D and WASH, and Btk29A phosphorylates WASH. This phosphorylation activates endosomal WASH function in flies and mice. In contrast, a phospho-mimetic WASH variant induces endosomal actin accumulation, premature luminal endocytosis and cortical F-actin disassembly. We conclude that PTPs and Btk29A regulate WASH activity to balance the endosomal and cortical F-actin networks during epithelial tube maturation.

Additional Information

© 2019 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Received 29 October 2018; Accepted 24 April 2019; Published 16 May 2019. Data availability: The source data underlying Figs. 1d, e, f, h, 2b, c, 3c, e, g, h, 4b–e, h, 5b, 6c–e, 7c, e, g, 8b, c, e, f and Supplementary Figs. 1b, 2b, 3c, g, i, l, n, 4b, 5b, 7b, d, f, 8b, f, 9d, e, 10b, d, e are provided as Source Data file. All reagents and further experimental data are available from the corresponding author upon reasonable request. We would like to thank Maura Strigini, Marcos Gonzalez-Gaitan, Kazuo Emoto, Markus Affolter, Susan Parkhurst, the Bloomington Drosophila Stock Center, Kyoto Drosophila Stock Center, the Drosophila Genomics Resource Center (DGRC; IN) and the Developmental Studies Hybridoma Bank (DSHB; IA) for fly strains, clones and antibodies. We thank fly community that isolated, characterized or distributed mutants or antibodies. Special thanks to Flybase for the Drosophila genomic resources. We thank the Stockholm University Imaging Facility (IFSU). We thank members of the M. Mannervik, C. Samakovlis, Q. Dai and S. Åström laboratories for comments and support during the project, especially K. Senti, B. Arefin and Y. Zhang for constructs and strains and M. Björk for fly service. This work was funded by the Swedish Research Council and the Swedish Cancer Society to C.S. C.S. was supported by the German Research Foundation (DFG), grant KFO309 (project number 284237345). D.D.B. was supported by National Institutes of Health grant 1R01 DK107733. Author Contributions: V.T. conceived the project, designed and executed the genetic analysis, analysed data and wrote the paper. D.L. planned and executed the in vivo biochemical analysis, generated transgenic flies, analysed data and contributed to writing. G.T. planned and executed experiments to map WASH phosphorylation and the biochemical analysis in Drosophila S2 cells, generated transgenic flies, analysed data and wrote the paper. A.F. planned, analysed and executed the WASH mutant analysis in mouse fibroblasts, provided reagents and wrote the paper. K.Z. provided fly strains, analysed data and commented on the manuscript. R.M. proposed experiments, analysed data and wrote the paper. D.D.B. planned and analysed the WASH mutant analysis in mouse fibroblasts, provided reagents and wrote the paper. C.S. conceived the project, proposed experiments, analysed data and wrote the paper. V.T created the graphical illustration in Fig. 9b and G.T. the graphics in Fig. 3b and Supplementary Fig. 6a. The authors declare no competing interests.

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