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Myosin 1b promotes the formation of post-Golgi carriers by regulating actin assembly and membrane remodelling at the trans-Golgi network

Abstract

The function of organelles is intimately associated with rapid changes in membrane shape. By exerting force on membranes, the cytoskeleton and its associated motors have an important role in membrane remodelling. Actin and myosin 1 have been implicated in the invagination of the plasma membrane during endocytosis. However, whether myosin 1 and actin contribute to the membrane deformation that gives rise to the formation of post-Golgi carriers is unknown. Here we report that myosin 1b regulates the actin-dependent post-Golgi traffic of cargo, generates force that controls the assembly of F-actin foci and, together with the actin cytoskeleton, promotes the formation of tubules at the TGN. Our results provide evidence that actin and myosin 1 regulate organelle shape and uncover an important function for myosin 1b in the initiation of post-Golgi carrier formation by regulating actin assembly and remodelling TGN membranes.

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Figure 1: Distribution of Myo1b, MPR, F-actin and the Arp2/3 complex in the perinuclear region.
Figure 2: Myo1b controls the steady-state distribution of MPR.
Figure 3: Myo1b knockdown impairs the TGN exit of cargo.
Figure 4: Myo1b controls the formation of tubular carriers at the TGN.
Figure 5: Myo1b depletion reduces the number of F-actin foci and of Arp2/3 complex structures in the vicinity of the TGN.
Figure 6: Functional Myo1b is required for organizing F-actin–Arp2/3 foci.
Figure 7: Depletion of the Arp2/3 complex induces MPR accumulation in TGN and inhibits post-Golgi carrier formation.
Figure 8: Model for the role of Myo1b in the formation of tubular carriers at the TGN.

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References

  1. Zimmerberg, J. & Kozlov, M. M. How proteins produce cellular membrane curvature. Nat. Rev. Mol. Cell Biol. 7, 9–19 (2006).

    Article  CAS  PubMed  Google Scholar 

  2. Anitei, M. et al. Protein complexes containing CYFIP/Sra/PIR121 coordinate Arf1 and Rac1 signalling during clathrin-AP-1-coated carrier biogenesis at the TGN. Nat. Cell Biol. 12, 330–340 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Carreno, S., Engqvist-Goldstein, A. E., Zhang, C. X., McDonald, K. L. & Drubin, D. G. Actin dynamics coupled to clathrin-coated vesicle formation at the trans-Golgi network. J. Cell Biol. 165, 781–788 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Poupon, V. et al. Clathrin light chains function in mannose phosphate receptor trafficking via regulation of actin assembly. Proc. Natl Acad. Sci. USA 105, 168–173 (2008).

    Article  CAS  PubMed  Google Scholar 

  5. Salvarezza, S. B. et al. LIM kinase 1 and cofilin regulate actin filament population required for dynamin-dependent apical carrier fission from the trans-Golgi network. Mol. Biol. Cell 20, 438–451 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Musch, A., Cohen, D. & Rodriguez-Boulan, E. Myosin II is involved in the production of constitutive transport vesicles from the TGN. J. Cell Biol. 138, 291–306 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Sahlender, D. A. et al. Optineurin links myosin VI to the Golgi complex and is involved in Golgi organization and exocytosis. J. Cell Biol. 169, 285–295 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Warner, C. L. et al. Loss of myosin VI reduces secretion and the size of the Golgi in fibroblasts from Snell’s waltzer mice. Embo J. 22, 569–579 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Miserey-Lenkei, S. et al. Rab and actomyosin-dependent fission of transport vesicles at the Golgi complex. Nat. Cell Biol. 12, 645–654 (2010).

    Article  CAS  PubMed  Google Scholar 

  10. Cordonnier, M. N., Dauzonne, D., Louvard, D. & Coudrier, E. Actin filaments and myosin I alpha cooperate with microtubules for the movement of lysosomes. Mol. Biol. Cell 12, 4013–4029 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Raposo, G. et al. Association of myosin I alpha with endosomes and lysosomes in mammalian cells. Mol. Biol. Cell 10, 1477–1494 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Salas-Cortes, L. et al. Myosin Ib modulates the morphology and the protein transport within multi-vesicular sorting endosomes. J. Cell Sci. 118, 4823–4832 (2005).

    Article  CAS  PubMed  Google Scholar 

  13. Tang, N. & Ostap, E. M. Motor domain-dependent localization of myo1b (myr-1). Curr. Biol. 11, 1131–1135 (2001).

    Article  CAS  PubMed  Google Scholar 

  14. Ruppert, C. et al. Localization of the rat myosin I molecules myr 1 and myr 2 and in vivo targeting of their tail domains. J. Cell Sci. 108, 3775–3786 (1995).

    CAS  PubMed  Google Scholar 

  15. Ghosh, P., Dahms, N. M. & Kornfeld, S. Mannose 6-phosphate receptors: new twists in the tale. Nat. Rev. Mol. Cell Biol. 4, 202–212 (2003).

    Article  CAS  PubMed  Google Scholar 

  16. Riedl, J. et al. Lifeact: a versatile marker to visualize F-actin. Nat. Methods 5, 605–607 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Zhong, Q. et al. Endosomal localization and function of sorting nexin 1. Proc. Natl Acad. Sci. USA 99, 6767–6772 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Cao, H. et al. Actin and Arf1-dependent recruitment of a cortactin–dynamin complex to the Golgi regulates post-Golgi transport. Nat. Cell Biol. 7, 483–492 (2005).

    Article  CAS  PubMed  Google Scholar 

  19. Kreitzer, G., Marmorstein, A., Okamoto, P., Vallee, R. & Rodriguez-Boulan, E. Kinesin and dynamin are required for post-Golgi transport of a plasma-membrane protein. Nat. Cell Biol. 2, 125–127 (2000).

    Article  CAS  PubMed  Google Scholar 

  20. Ludwig, T. et al. Differential sorting of lysosomal enzymes in mannose 6-phosphate receptor-deficient fibroblasts. EMBO J. 13, 3430–3437 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Lazaro-Dieguez, F. et al. Variable actin dynamics requirement for the exit of different cargo from the trans-Golgi network. FEBS Lett. 581, 3875–3881 (2007).

    Article  CAS  PubMed  Google Scholar 

  22. von Blume, J. et al. Actin remodeling by ADF/cofilin is required for cargo sorting at the trans-Golgi network. J. Cell Biol. 187, 1055–1069 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Waguri, S. et al. Visualization of TGN to endosome trafficking through fluorescently labelled MPR and AP-1 in living cells. Mol. Biol. Cell 14, 142–155 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Loubery, S. & Coudrier, E. Myosins in the secretory pathway: tethers or transporters? Cell Mol. Life Sci. 65, 2790–2800 (2008).

    Article  CAS  PubMed  Google Scholar 

  25. Sokac, A. M., Schietroma, C., Gundersen, C. B. & Bement, W. M. Myosin-1c couples assembling actin to membranes to drive compensatory endocytosis. Dev. Cell 11, 629–640 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Friedman, A. L., Geeves, M. A., Manstein, D. J. & Spudich, J. A. Kinetic characterization of myosin head fragments with long-lived myosin.ATP states. Biochemistry 37, 9679–9687 (1998).

    Article  CAS  PubMed  Google Scholar 

  27. Shimada, T., Sasaki, N., Ohkura, R. & Sutoh, K. Alanine scanning mutagenesis of the switch I region in the ATPase site of Dictyostelium discoideum myosin II. Biochemistry 36, 14037–14043 (1997).

    Article  CAS  PubMed  Google Scholar 

  28. Klemm, R. W et al. Segregation of sphingolipids and sterols duringformation of secretory vesicles at the trans-Golgi network. J. Cell Biol. 185, 601–612 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Polishchuk, R., Di Pentima, A. & Lippincott-Schwartz, J. Delivery of raft-associated, GPI-anchored proteins to the apical surface of polarized MDCK cells by a transcytotic pathway. Nat. Cell Biol. 6, 297–307 (2004).

    Article  CAS  PubMed  Google Scholar 

  30. Roux, A. et al. A minimal system allowing tubulation with molecular motors pulling on giant liposomes. Proc. Natl Acad. Sci. USA 99, 5394–5399 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Laakso, J. M., Lewis, J. H., Shuman, H. & Ostap, E. M. Myosin I can act as a molecular force sensor. Science 321, 133–136 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Laakso, J. M., Lewis, J. H., Shuman, H. & Ostap, E. M. Control of myosin-I force sensing by alternative splicing. Proc. Natl Acad. Sci. USA 107, 698–702 (2010).

    Article  CAS  PubMed  Google Scholar 

  33. Leduc, C. et al. Cooperative extraction of membrane nanotubes by molecular motors. Proc. Natl Acad. Sci. USA 101, 17096–17101 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Sun, Y., Martin, A. C. & Drubin, D. G. Endocytic internalization in budding yeast requires coordinated actin nucleation and myosin motor activity. Dev. Cell 11, 33–46 (2006).

    Article  CAS  PubMed  Google Scholar 

  35. Naccache, S. N., Hasson, T. & Horowitz, A. Binding of internalized receptors to the PDZ domain of GIPC/synectin recruits myosin VI to endocytic vesicles. Proc. Natl Acad. Sci. USA 103, 12735–12740 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Noguchi, T., Lenartowska, M. & Miller, K. G. Myosin VI stabilizes an actin network during Drosophila spermatid individualization. Mol. Biol. Cell 17, 2559–2571 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Noguchi, T., Lenartowska, M., Rogat, A. D., Frank, D. J. & Miller, K. G. Proper cellular reorganization during Drosophila spermatid individualization depends on actin structures composed of two domains, bundles and meshwork, that are differentially regulated and have different functions. Mol. Biol. Cell 19, 2363–2372 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Dippold, H. C. et al. GOLPH3 bridges phosphatidylinositol-4-phosphate and actomyosin to stretch and shape the Golgi to promote budding. Cell 139, 337–351 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Veigel, C. et al. The motor protein myosin-I produces its working stroke in two steps. Nature 398, 530–533 (1999).

    Article  CAS  PubMed  Google Scholar 

  40. Loubery, S. et al. Different microtubule motors move early and late endocytic compartments. Traffic 9, 492–509 (2008).

    Article  CAS  PubMed  Google Scholar 

  41. Chen, H. J., Remmler, J., Delaney, J. C., Messner, D. J. & Lobel, P. Mutational analysis of the cation-independent mannose 6-phosphate/insulin-like growth factor II receptor. A consensus casein kinase II site followed by 2 leucines near the carboxyl terminus is important for intracellular targeting of lysosomal enzymes. J. Biol. Chem. 268, 22338–22346 (1993).

    CAS  PubMed  Google Scholar 

  42. Grosshans, B. L. et al. TEDS site phosphorylation of the yeast myosins I is required for ligand-induced but not for constitutive endocytosis of the G protein-coupled receptor Ste2p. J. Biol. Chem. 281, 11104–11114 (2006).

    Article  CAS  PubMed  Google Scholar 

  43. Lin, S. X., Mallet, W. G., Huang, A. Y. & Maxfield, F. R. Endocytosed cation-independent mannose 6-phosphate receptor traffics via the endocytic recycling compartment en route to the trans-Golgi network and a subpopulation of late endosomes. Mol. Biol. Cell 15, 721–733 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Sibarita, J. B. Deconvolution microscopy. Adv. Biochem. Eng. Biotechnol. 95, 201–243 (2005).

    PubMed  Google Scholar 

  45. Racine, V. et al. Visualization and quantification of vesicle trafficking on athree-dimensional cytoskeleton network in living cells. J. Microsc. 225, 214–228 (2007).

    Article  PubMed  Google Scholar 

  46. Mardones, G. A. et al. The trans-Golgi network accessory protein p56 promotes long-range movement of GGA/clathrin-containing transport carriers and lysosomal enzyme sorting. Mol. Biol. Cell 18, 3486–3501 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Piccirillo, R. et al. An unconventional dileucine-based motif and a novel cytosolic motif are required for the lysosomal and melanosomal targeting of OA1. J. Cell Sci. 119, 2003–2014 (2006).

    Article  CAS  PubMed  Google Scholar 

  48. Riederer, M. A., Soldati, T., Shapiro, A. D., Lin, J. & Pfeffer, S. R. Lysosome biogenesis requires Rab9 function and receptor recycling from endosomes to the trans-Golgi network. J. Cell Biol. 125, 573–582 (1994).

    Article  CAS  PubMed  Google Scholar 

  49. Mallard, F. et al. Direct pathway from early/recycling endosomes to the Golgi apparatus revealed through the study of shiga toxin B-fragment transport. J. Cell Biol. 143, 973–990 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Derivery, E., Lombard, B., Loew, D. & Gautreau, A. The wave complex is intrinsically inactive. Cell Motil. Cytoskeleton. 66, 777–790 (2009).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank L. Salas-Cortes for designing the Myo1b siRNA, M. Prospéri for Myo1b antibodies and the Cherry–Myo1b, FlagHA–Myo1b-5MR and FlagHA–Myo1b-5ME constructs and J. Lee-Tin-Wah and P. Martin for helping set up the actin sliding assay. We thank V. Fraisier, J-B. Sibarita, F. Waharte and L. Sengmanivong for their expertise in microscopy and the Nikon imaging centre@Institut Curie-CNRS. We thank E. Derivery for setting up the Myo1b purification with the FlipIn system. We thank J. Kean for critical reading of the manuscript.

This work has been supported by the Institut Curie, the CNRS and the Agence Nationale pour la Recherche (grant ANR 09-BLAN-0027). C.G.A. has been the recipient of an EMBO long-term fellowship (ALTF 607-2006) and a Marie Curie action intra-European fellowship for career development (FP7-PEOPLE-2007-2-1-IEF N. 2200088).

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C.G.A. and E.C. conceived the project and wrote the manuscript; C.G.A. generated and analysed most of the data; A.Y. carried out Myo1b and Myo1b mutant purification as well as their characterization in vitro; D.T. and G.R. generated and analysed the electron microscopy data; D.L. revised the manuscript.

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Correspondence to Evelyne Coudrier.

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Almeida, C., Yamada, A., Tenza, D. et al. Myosin 1b promotes the formation of post-Golgi carriers by regulating actin assembly and membrane remodelling at the trans-Golgi network. Nat Cell Biol 13, 779–789 (2011). https://doi.org/10.1038/ncb2262

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