Article

Epithelial rotation promotes the global alignment of contractile actin bundles during Drosophila egg chamber elongation

  • Nature Communications 5, Article number: 5511 (2014)
  • doi:10.1038/ncomms6511
  • Download Citation
Received:
Accepted:
Published online:

Abstract

Tissues use numerous mechanisms to change shape during development. The Drosophila egg chamber is an organ-like structure that elongates to form an elliptical egg. During elongation the follicular epithelial cells undergo a collective migration that causes the egg chamber to rotate within its surrounding basement membrane. Rotation coincides with the formation of a ‘molecular corset’, in which actin bundles in the epithelium and fibrils in the basement membrane are all aligned perpendicular to the elongation axis. Here we show that rotation plays a critical role in building the actin-based component of the corset. Rotation begins shortly after egg chamber formation and requires lamellipodial protrusions at each follicle cell’s leading edge. During early stages, rotation is necessary for tissue-level actin bundle alignment, but it becomes dispensable after the basement membrane is polarized. This work highlights how collective cell migration can be used to build a polarized tissue organization for organ morphogenesis.

Additional access options:

Already a subscriber?  Log in  now or  Register  for online access.

References

  1. 1.

    & Mass transit: epithelial morphogenesis in the Drosophila egg chamber. Dev. Dyn 232, 559–574 (2005).

  2. 2.

    & Expanding the morphogenetic repertoire: perspectives from the Drosophila egg. Dev. Cell 22, 12–23 (2012).

  3. 3.

    Drosophila egg chamber elongation: insights into how tissues and organs are shaped. Fly (Austin) 6, 213–227 (2012).

  4. 4.

    The Drosophila egg chamber-a new spin on how tissues elongate. Integr. Comp. Biol. 54, 667–676 (2014).

  5. 5.

    The microfilament pattern in the somatic follicle cells of mid-vitellogenic ovarian follicles of Drosophila. Eur. J. Cell Biol. 53, 349–356 (1990).

  6. 6.

    , & Laminin and basement membrane-associated microfilaments in wild-type and mutant Drosophila ovarian follicles. J. Cell Sci. 100, (Pt 4): 781–788 (1991).

  7. 7.

    , , & The receptor tyrosine phosphatase Dlar and integrins organize actin filaments in the Drosophila follicular epithelium. Curr. Biol. 11, 1317–1327 (2001).

  8. 8.

    , , & The serine/threonine kinase dPak is required for polarized assembly of F-actin bundles and apical-basal polarity in the Drosophila follicular epithelium. Dev. Biol. 305, 470–482 (2007).

  9. 9.

    & The receptor-like tyrosine phosphatase lar is required for epithelial planar polarity and for axis determination within drosophila ovarian follicles. Development 128, 3209–3220 (2001).

  10. 10.

    & Global tissue revolutions in a morphogenetic movement controlling elongation. Science 331, 1071–1074 (2011).

  11. 11.

    , , , & A screen for round egg mutants in Drosophila identifies tricornered, furry, and misshapen as regulators of egg chamber elongation. G3 (Bethesda) 2, 371–378 (2012).

  12. 12.

    , , & The cadherin Fat2 is required for planar cell polarity in the Drosophila ovary. Development 136, 4123–4132 (2009).

  13. 13.

    , , & Tissue elongation requires oscillating contractions of a basal actomyosin network. Nat. Cell Biol. 12, 1133–1142 (2010).

  14. 14.

    Organization and in vitro activity of microfilament bundles associated with the basement membrane of Drosophila follicles. Acta Histochem. Suppl. 41, 201–210 (1991).

  15. 15.

    , & Misshapen decreases integrin levels to promote epithelial motility and planar polarity in Drosophila. J. Cell Biol. 200, 721–729 (2013).

  16. 16.

    , , & Mechanical integration of actin and adhesion dynamics in cell migration. Annu. Rev. Cell Dev. Biol. 26, 315–333 (2010).

  17. 17.

    Life at the leading edge. Cell 145, 1012–1022 (2011).

  18. 18.

    & Filopodia: molecular architecture and cellular functions. Nat. Rev. Mol. Cell Biol. 9, 446–454 (2008).

  19. 19.

    & The integrin adhesion complex changes its composition and function during morphogenesis of an epithelium. J. Cell Sci. 122, 4363–4374 (2009).

  20. 20.

    & The role of the actomyosin cytoskeleton in coordination of tissue growth during Drosophila oogenesis. Curr. Biol. 17, 1349–1355 (2007).

  21. 21.

    , , , & The large Maf factor Traffic Jam controls gonad morphogenesis in Drosophila. Nat. Cell Biol. 5, 994–1000 (2003).

  22. 22.

    et al. A Rab10-dependent mechanism for polarized basement membrane secretion during organ morphogenesis. Dev. Cell 24, 159–168 (2013).

  23. 23.

    & Microtubule polarity predicts direction of egg chamber rotation in Drosophila. Curr. Biol. 23, 1472–1477 (2013).

  24. 24.

    , & Dynamic and structural signatures of lamellar actomyosin force generation. Mol. Biol. Cell 22, 1330–1339 (2011).

  25. 25.

    & Characterization of a dorsal-eye Gal4 line in Drosophila. Genesis 48, 3–7 (2010).

  26. 26.

    et al. Characterization of two classes of small molecule inhibitors of Arp2/3 complex. Nature 460, 1031–1034 (2009).

  27. 27.

    , , , & Symmetry breaking in cultured mammalian cells. In Vitro Cell Dev. Biol. Anim. 36, 563–565 (2000).

  28. 28.

    et al. Guidance of collective cell migration by substrate geometry. Integr. Biol. (Camb) 5, 1026–1035 (2013).

  29. 29.

    , , , & ROCK-mediated contractility, tight junctions and channels contribute to the conversion of a preapical patch into apical surface during isochoric lumen initiation. J. Cell Sci. 121, 3649–3663 (2008).

  30. 30.

    , , & Symmetry-breaking in mammalian cell cohort migration during tissue pattern formation: role of random-walk persistence. Cell Motil. Cytoskeleton 61, 201–213 (2005).

  31. 31.

    et al. A mathematical method for the 3D analysis of rotating deformable systems applied on lumen-forming MDCK cell aggregates. Cytoskeleton (Hoboken) 67, 224–240 (2010).

  32. 32.

    Fellow travellers: emergent properties of collective cell migration. EMBO Rep. 13, 984–991 (2012).

  33. 33.

    , , , & Coherent angular motion in the establishment of multicellular architecture of glandular tissues. Proc. Natl Acad. Sci. USA 109, 1973–1978 (2012).

  34. 34.

    , , , & Rotational motion during three-dimensional morphogenesis of mammary epithelial acini relates to laminin matrix assembly. Proc. Natl Acad. Sci. USA 110, 163–168 (2013).

  35. 35.

    & Principles of planar polarity in animal development. Development 138, 1877–1892 (2011).

  36. 36.

    et al. The carnegie protein trap library: a versatile tool for Drosophila developmental studies. Genetics 175, 1505–1531 (2007).

  37. 37.

    , & Planar polarized actomyosin contractile flows control epithelial junction remodelling. Nature 468, 1110–1114 (2010).

  38. 38.

    , & Pulsed contractions of an actin-myosin network drive apical constriction. Nature 457, 495–499 (2009).

  39. 39.

    , , , & A protocol for culturing Drosophila melanogaster stage 9 egg chambers for live imaging. Nat. Protoc. 2, 2467–2473 (2007).

  40. 40.

    & Variable-angle epifluorescence microscopy: a new way to look at protein dynamics in the plant cell cortex. Plant J. 53, 186–196 (2008).

  41. 41.

    , , & Single-molecule analysis of cell surface dynamics in Caenorhabditis elegans embryos. Nat. Methods 11, 677–682 (2014).

  42. 42.

    , & Highly inclined thin illumination enables clear single-molecule imaging in cells. Nat. Methods 5, 159–161 (2008).

  43. 43.

    , , , & Abi activates WASP to promote sensory organ development. Nat. Cell Biol. 7, 977–984 (2005).

  44. 44.

    , , & Developmental expression of Drosophila Wiskott-Aldrich syndrome family proteins. Dev. Dyn. 241, 608–626 (2012).

Download references

Acknowledgements

We thank C. Klämbt, T. Lecuit, B. McCartney, D. Montell, T. Schu¨pbach, E. Wieschaus and J. Zallen for generously providing reagents, and F. Robin and E. Munro for assistance with near-total internal reflection fluorescence microscopy. We are also grateful to E. Ferguson and members of the Horne-Badovinac lab for insightful comments on the manuscript and to N. Ellis for illustrations. This work was supported by NIH T32 GM007183 to M.C., a NSF Graduate Research Fellowship to G.R.R-SJ., an ACS Postdoctoral Fellowship (PF-12-135-01-CSM) and an award from the AHA to L.L., an NSERC Alexander Graham Bell Canada Graduate Scholarship to M.J.F., a Canadian Institutes of Health Research operating grant (MOP-272122) to G.T., a Burroughs Wellcome Career Award, American Asthma Early Excellence Award and funding from the Packard Foundation to M.L.G., and grants from the Edward Mallinckrodt, Jr Foundation and NIH (R01 GM094276) to S.H.-B.

Author information

Author notes

    • Maureen Cetera
    •  & Guillermina R. Ramirez-San Juan

    These authors contributed equally to this work

Affiliations

  1. Department of Molecular Genetics and Cell Biology, The University of Chicago, 920 East 58th Street, Chicago, Illinois 60637, USA

    • Maureen Cetera
    • , Guillermina R. Ramirez-San Juan
    • , Lindsay Lewellyn
    •  & Sally Horne-Badovinac
  2. Committee on Development, Regeneration and Stem Cell Biology, The University of Chicago, 920 East 58th Street, Chicago, Illinois 60637, USA

    • Maureen Cetera
    •  & Sally Horne-Badovinac
  3. Institute for Biophysical Dynamics, James Franck Institute and Department of Physics, The University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA

    • Guillermina R. Ramirez-San Juan
    • , Patrick W. Oakes
    •  & Margaret L. Gardel
  4. Department of Biological Sciences, Butler University, 4600 Sunset Boulevard, Indianapolis, Indiana 46208, USA

    • Lindsay Lewellyn
  5. Life Sciences Centre, Department of Cellular and Physiological Sciences, University of British Columbia, 2350 Health Sciences Mall, Vancouver, British Columbia, Canada V6T 1Z3

    • Michael J. Fairchild
    •  & Guy Tanentzapf

Authors

  1. Search for Maureen Cetera in:

  2. Search for Guillermina R. Ramirez-San Juan in:

  3. Search for Patrick W. Oakes in:

  4. Search for Lindsay Lewellyn in:

  5. Search for Michael J. Fairchild in:

  6. Search for Guy Tanentzapf in:

  7. Search for Margaret L. Gardel in:

  8. Search for Sally Horne-Badovinac in:

Contributions

M.C. and G.R.R.-S.J. designed and performed the experiments, and analysed the data. S.H.-B. supervised the project with critical input from M.L.G. P.W.O. made important intellectual contributions and designed the quantitative method used to measure actin bundle alignment. L.L. made important intellectual contributions. M.J.F. and G.T. provided key reagents. M.C., G.R.R.-S.J. and S.H.-B. wrote the manuscript. All authors helped to edit the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Sally Horne-Badovinac.

Supplementary information

PDF files

  1. 1.

    Supplementary Figures and Tables

    Supplementary Figures 1-5 and Supplementary Tables 1-3

Videos

  1. 1.

    Supplementary Movie 1

    Membrane protrusions extend from the leading edge of each follicle cell.Membranes of a stage seven egg chamber expressing Nrg and Indy GFP protein traps. follicle cellswere imaged at the basal surface. Laser-scanning confocal images. 25.5 mins/ 30 sec interval. 190x play back rate. Scale bar,10 μm.

  2. 2.

    Supplementary Movie 2

    Abi depletion in all follicle cells blocks rotation. Nrg and Indy GFP protein traps mark follicle cell membranes in a stage eight egg chamber. Control egg chambers rotate normally (left) while rotation is blocked when Abi RNAi is expressed in all follicle cells (right). Laser-scanning confocal images. 17.5 mins/ 30 sec interval. 210x play back rate. Scale bar,10 μm.

  3. 3.

    Supplementary Movie 3

    A small clone expressing Abi RNAi is carried along within the migrating epithelium. The follicle cell membranes of a stage seven egg chamber are marked with the Nrg GFP protein trap. Follicle cells expressing Abi RNAi are marked with RFP. The clone is carried along by wild-type neighbors. Laser-scanning confocal images. 45 mins/1 min interval. 450x play back rate. Scale bar, 10μm

  4. 4.

    Supplementary Movie 4

    Egg chambers rotate prior to stage five. Stage one, three, and four egg chambers expressing a nuclear marker, His2Av RFP, rotate. A row of cells in each egg chamber is indicated by colored dots through time. Spinning disk confocal images. 180 mins/ 1 min interval. 600x play back rate. Scale bar,10 μm.

  5. 5.

    Supplementary Movie 5

    fat2 mutant egg chambers do not rotate at early stages. A fat2n103-2 ovariole containing stage one, two, three and four egg chambers that do not rotate during a twenty minute time lapse. FM4-64 dye was used to label cell membranes. Spinning disk confocal images. 20 mins/ 1 min interval. 600x play back rate. Scale bar,10 μm

  6. 6.

    Supplementary Movie 6

    Global actin bundle alignment of a stage four egg chamber increases as follicle cells migrate. TIRF microscopy of an egg chamber expressing the actin binding domain of Utrophin fused to GFP to visualize F-actin live. Basal actin bundles are initially not well aligned. As the follicle cells migrate, the global alignment of the actin bundles fluctuates. At the end of the time series, the basal actin bundles are globally aligned. 60 mins/ 1 min interval. 515x play back rate. Scale bar,10 μm.

  7. 7.

    Supplementary Movie 7

    Follicle cell migration is blocked by the Arp2/3 inhibitor. Membranes of a stage seven egg chamber expressing the Indy GFP protein trap. The inhibitor (CK-666, right) or control molecule (CK-689, left) was added at the 20-minute time point (blank frame). The control egg chamber continues to rotate while the inhibitor-treated egg chamber stops rotating within an hour. Spinning disk confocal images. 80 mins/ 1 min interval. 370x play back rate. Scale bar,15 μm.

  8. 8.

    Supplementary Movie 8

    Acutely blocking follicle cell migration in a stage seven egg chamber with the Arp2/3 inhibitor does not alter global actin bundle alignment. An Arp2/3 inhibitor-treated egg chamber expressing the actin binding domain of Utrophin fused to GFP. The egg chamber is not rotating but the basal actin bundles maintain their global alignment. Images were acquired on a laser-scanning confocal. 21 mins/ 30 sec interval. 250x play back rate. Scale bar,10 μm.

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.