Abstract
Seamless tubes form intracellularly without cell–cell or autocellular junctions. Such tubes have been described across phyla, but remain mysterious despite their simple architecture. In Drosophila, seamless tubes are found within tracheal terminal cells, which have dozens of branched protrusions extending hundreds of micrometres. We find that mutations in multiple components of the dynein motor complex block seamless tube growth, raising the possibility that the lumenal membrane forms through minus-end-directed transport of apical membrane components along microtubules. Growth of seamless tubes is polarized along the proximodistal axis by Rab35 and its apical membrane-localized GAP, Whacked. Strikingly, loss of whacked (or constitutive activation of Rab35) leads to tube overgrowth at terminal cell branch tips, whereas overexpression of Whacked (or dominant-negative Rab35) causes formation of ectopic tubes surrounding the terminal cell nucleus. Thus, vesicle trafficking has key roles in making and shaping seamless tubes.
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References
Samakovlis, C. et al. Development of the Drosophila tracheal system occurs by a series of morphologically distinct but genetically coupled branching events. Development 122, 1395–1407 (1996).
Ribeiro, C., Neumann, M. & Affolter, M. Genetic control of cell intercalation during tracheal morphogenesis in Drosophila. Curr. Biol. 14, 2197–2207 (2004).
Buechner, M. Tubes and the single C. elegans excretory cell. Trends Cell Biol. 12, 479–484 (2002).
Bar, T., Guldner, F. H. & Wolff, J. R. ‘Seamless’ endothelial cells of blood capillaries. Cell Tissue Res. 235, 99–106 (1984).
Lubarsky, B. & Krasnow, M. A. Tube morphogenesis: making and shaping biological tubes. Cell 112, 19–28 (2003).
Uv, A., Cantera, R. & Samakovlis, C. Drosophila tracheal morphogenesis: intricate cellular solutions to basic plumbing problems. Trends Cell Biol. 13, 301–309 (2003).
Gervais, L. & Casanova, J. In vivo coupling of cell elongation and lumen formation in a single cell. Curr. Biol. 20, 359–366 (2010).
Brodu, V., Baffet, A. D., Le Droguen, P. M., Casanova, J. & Guichet, A. A developmentally regulated two-step process generates a noncentrosomal microtubule network in Drosophila tracheal cells. Dev. Cell 18, 790–801 (2010).
Sutherland, D., Samakovlis, C. & Krasnow, M. A. branchless encodes a Drosophila FGF homolog that controls tracheal cell migration and the pattern of branching. Cell 87, 1091–1101 (1996).
Ribeiro, C., Ebner, A. & Affolter, M. In vivo imaging reveals different cellular functions for FGF and Dpp signaling in tracheal branching morphogenesis. Dev. Cell 2, 677–683 (2002).
Jarecki, J., Johnson, E. & Krasnow, M. A. Oxygen regulation of airway branching in Drosophila is mediated by branchless FGF. Cell 99, 211–220 (1999).
Berry, K. L., Bulow, H. E., Hall, D. H. & Hobert, O. A. C. elegans CLIC-like protein required for intracellular tube formation and maintenance. Science 302, 2134–2137 (2003).
Ulmasov, B., Bruno, J., Gordon, N., Hartnett, M. E. & Edwards, J. C. Chloride intracellular channel protein-4 functions in angiogenesis by supporting acidification of vacuoles along the intracellular tubulogenic pathway. Am. J. Pathol. 174, 1084–1096 (2009).
Grawe, F., Wodarz, A., Lee, B., Knust, E. & Skaer, H. The Drosophila genes crumbs and stardust are involved in the biogenesis of adherens junctions. Development 122, 951–959 (1996).
Wodarz, A., Hinz, U., Engelbert, M. & Knust, E. Expression of crumbs confers apical character on plasma membrane domains of ectodermal epithelia of Drosophila. Cell 82, 67–76 (1995).
Waterman-Storer, C. M. et al. The interaction between cytoplasmic dynein and dynactin is required for fast axonal transport. Proc. Natl Acad. Sci. USA 94, 12180–12185 (1997).
Ghabrial, A. S., Levi, B. P. & Krasnow, M. A. A systematic screen for tube morphogenesis and branching genes in the Drosophila tracheal system. PLoS Genet. 7, e1002087 (2011).
Levi, B. P., Ghabrial, A. S. & Krasnow, M. A. Drosophila talin and integrin genes are required for maintenance of tracheal terminal branches and luminal organization. Development 133, 2383–2393 (2006).
Albert, S. & Gallwitz, D. Two new members of a family of Ypt/Rab GTPase activating proteins. Promiscuity of substrate recognition. J. Biol. Chem. 274, 33186–33189 (1999).
Albert, S., Will, E. & Gallwitz, D. Identification of the catalytic domains and their functionally critical arginine residues of two yeast GTPase-activating proteins specific for Ypt/Rab transport GTPases. EMBO J. 18, 5216–5225 (1999).
Strom, M., Vollmer, P., Tan, T. J. & Gallwitz, D. A yeast GTPase-activating protein that interacts specifically with a member of the Ypt/Rab family. Nature 361, 736–739 (1993).
Zhang, J. et al. Thirty-one flavors of Drosophila rab proteins. Genetics 176, 1307–1322 (2007).
Hsu, C. et al. Regulation of exosome secretion by Rab35 and its GTPase-activating proteins TBC1D10A-C. J. Cell Biol. 189, 223–232 (2010).
Pan, F. et al. Feedback inhibition of calcineurin and Ras by a dual inhibitory protein Carabin. Nature 445, 433–436 (2007).
Itoh, T. & Fukuda, M. Identification of EPI64 as a GTPase-activating protein specific for Rab27A. J. Biol. Chem. 281, 31823–31831 (2006).
Ishibashi, K., Kanno, E., Itoh, T. & Fukuda, M. Identification and characterization of a novel Tre-2/Bub2/Cdc16 (TBC) protein that possesses Rab3A-GAP activity. Genes Cells 14, 41–52 (2009).
Chevallier, J. et al. Rab35 regulates neurite outgrowth and cell shape. FEBS Lett. 583, 1096–1101 (2009).
Chua, C. E., Lim, Y. S. & Tang, B. L. Rab35—a vesicular traffic-regulating small GTPase with actin modulating roles. FEBS Lett. 584, 1–6 (2010).
Echard, A. Membrane traffic and polarization of lipid domains during cytokinesis. Biochem. Soc. Trans. 36, 395–399 (2008).
Gao, Y. et al. Recycling of the Ca2+-activated K+ channel, KCa2.3, is dependent upon RME-1, Rab35/EPI64C, and an N-terminal domain. J. Biol. Chem. 285, 17938–17953 (2010).
Kouranti, I., Sachse, M., Arouche, N., Goud, B. & Echard, A. Rab35 regulates an endocytic recycling pathway essential for the terminal steps of cytokinesis. Curr. Biol. 16, 1719–1725 (2006).
Patino-Lopez, G. et al. Rab35 and its GAP EPI64C in T cells regulate receptor recycling and immunological synapse formation. J. Biol. Chem. 283, 18323–18330 (2008).
Sato, M. et al. Regulation of endocytic recycling by C. elegans Rab35 and its regulator RME-4, a coated-pit protein. EMBO J. 27, 1183–1196 (2008).
Shim, J. et al. Rab35 mediates transport of Cdc42 and Rac1 to the plasma membrane during phagocytosis. Mol. Cell Biol. 30, 1421–1433 (2010).
Uytterhoeven, V., Kuenen, S., Kasprowicz, J., Miskiewicz, K. & Verstreken, P. Loss of skywalker reveals synaptic endosomes as sorting stations for synaptic vesicle proteins. Cell 145, 117–132 (2011).
Zhang, J., Fonovic, M., Suyama, K., Bogyo, M. & Scott, M. P. Rab35 controls actin bundling by recruiting fascin as an effector protein. Science 325, 1250–1254 (2009).
Lee, S. & Kolodziej, P. A. The plakin Short Stop and the RhoA GTPase are required for E-cadherin-dependent apical surface remodeling during tracheal tube fusion. Development 129, 1509–1520 (2002).
Lee, T., Hacohen, N., Krasnow, M. & Montell, D. J. Regulated Breathless receptor tyrosine kinase activity required to pattern cell migration and branching in the Drosophila tracheal system. Genes Dev. 10, 2912–2921 (1996).
Pfeffer, S. Filling the Rab GAP. Nat. Cell Biol. 7, 856–857 (2005).
Fuchs, E. et al. Specific Rab GTPase-activating proteins define the Shiga toxin and epidermal growth factor uptake pathways. J. Cell Biol. 177, 1133–1143 (2007).
Huang, J., Zhou, W., Dong, W., Watson, A. M. & Hong, Y. Directed, efficient, and versatile modifications of the Drosophila genome by genomic engineering. Proc. Natl Acad. Sci. USA 106, 8284–8289 (2009).
Rubin, G. M. & Spradling, A. C. Genetic transformation of Drosophila with transposable element vectors. Science 218, 348–353 (1982).
Bellen, H. J. et al. The BDGP gene disruption project: single transposon insertions associated with 40% of Drosophila genes. Genetics 167, 761–781 (2004).
Acknowledgements
The authors would like to acknowledge: D. Willis, a former Stanford undergraduate student who helped with the rough mapping of wkd; B. Levi, who helped with the third chromosome screen; and M. Krasnow, in whose laboratory the screen and early phases of these studies were carried out. We also thank J. Zhang and the laboratories of M. Scott and H. Bellen (Baylor College of Medicine, USA) for making RabCA and RabDN stocks available to us before publication, the Engels laboratory for sharing deficiency strains, and M. Metzstein (The University of Utah, USA) for sharing 4x-SRF-GAL4 flies. We thank S. DiNardo, C. Burd, E. Bi and members of the Ghabrial and DiNardo laboratories for fruitful discussions. We thank A. S. Burguete, B. Levi and N. Speck for comments on the manuscript. J.S-R. was supported by NIH training grant 5-T32-HD007516-12 and, subsequently, by an NIH postdoctoral fellowship (NRSA—GM090438-01). A.S.G. gratefully acknowledges support from the University of Pennsylvania and the NIH (1R01GM089782-01A1). This work was supported in part by Basil O’Connor Starter Scholar Research Award Grant No. 5-FY09-43 from the March of Dimes Foundation.
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J.S-R. and A.S.G. conceived of and carried out all experiments described here. A.S.G. wrote the manuscript with input from J.S-R. Figures were assembled by J.S-R. and A.S.G.
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Schottenfeld-Roames, J., Ghabrial, A. Whacked and Rab35 polarize dynein-motor-complex-dependent seamless tube growth. Nat Cell Biol 14, 386–393 (2012). https://doi.org/10.1038/ncb2454
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DOI: https://doi.org/10.1038/ncb2454
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