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Netrin (UNC-6) mediates dendritic self-avoidance

Abstract

Dendrites from a single neuron may be highly branched but typically do not overlap. Self-avoidance behavior has been shown to depend on cell-specific membrane proteins that trigger mutual repulsion. Here we report the unexpected discovery that a diffusible cue, the axon guidance protein UNC-6 (Netrin), is required for self-avoidance of sister dendrites from the PVD nociceptive neuron in Caenorhabditis elegans. We used time-lapse imaging to show that dendrites fail to withdraw upon mutual contact in the absence of UNC-6 signaling. We propose a model in which the UNC-40 (Deleted in Colorectal Cancer; DCC) receptor captures UNC-6 at the tips of growing dendrites for interaction with UNC-5 on the apposing branch to induce mutual repulsion. UNC-40 also responds to dendritic contact through another pathway that is independent of UNC-6. Our findings offer a new model for how an evolutionarily conserved morphogenic cue and its cognate receptors can pattern a fundamental feature of dendritic architecture.

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Figure 1: UNC-6 signaling is required for contact-dependent self-avoidance.
Figure 2: UNC-6 signaling is required for contact-dependent self-avoidance.
Figure 3: UNC-6 functions as a permissive cue to prevent dendritic branch overlap.
Figure 4: UNC-40 functions in PVD to mediate self-avoidance and captures exogenous UNC-6 at the PVD cell surface.
Figure 5: UNC-5 is required in PVD and uses UNC-40-independent signaling to mediate self-avoidance.

References

  1. Jan, Y.N. & Jan, L.Y. Branching out: mechanisms of dendritic arborization. Nat. Rev. Neurosci. 11, 316–328 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Grueber, W.B. & Sagasti, A. Self-avoidance and tiling: mechanisms of dendrite and axon spacing. Cold Spring Harb. Perspect. Biol. 2, a001750 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  3. Corty, M.M., Matthews, B.J. & Grueber, W.B. Molecules and mechanisms of dendrite development in Drosophila. Development 136, 1049–1061 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Hattori, D., Millard, S., Wojtowicz, W. & Zipursky, S. Dscam-mediated cell recognition regulates neural circuit formation. Annu. Rev. Cell Dev. Biol. 24, 597–620 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Hughes, M.E. et al. Homophilic Dscam interactions control complex dendrite morphogenesis. Neuron 54, 417–427 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Matthews, B.J. et al. Dendrite self-avoidance is controlled by Dscam. Cell 129, 593–604 (2007).

    Article  CAS  PubMed  Google Scholar 

  7. Matsubara, D., Horiuchi, S.Y., Shimono, K., Usui, T. & Uemura, T. The seven-pass transmembrane cadherin Flamingo controls dendritic self-avoidance via its binding to a LIM domain protein, Espinas, in Drosophila sensory neurons. Genes Dev. 25, 1982–1996 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Long, H., Ou, Y., Rao, Y. & van Meyel, D.J. Dendrite branching and self-avoidance are controlled by Turtle, a conserved IgSF protein in Drosophila. Development 136, 3475–3484 (2009).

    Article  CAS  PubMed  Google Scholar 

  9. Zipursky, S.L. & Sanes, J.R. Chemoaffinity revisited: dscams, protocadherins, and neural circuit assembly. Cell 143, 343–353 (2010).

    Article  CAS  PubMed  Google Scholar 

  10. Wadsworth, W.G., Bhatt, H. & Hedgecock, E. Neuroglia and pioneer neurons express UNC-6 to provide global and local netrin cues for guiding migrations in C. elegans. Neuron 16, 35–46 (1996).

    Article  CAS  PubMed  Google Scholar 

  11. Serafini, T. et al. The netrins define a family of axon outgrowth-promoting proteins homologous to C. elegans UNC-6. Cell 78, 409–424 (1994).

    Article  CAS  PubMed  Google Scholar 

  12. Mitchell, K.J. et al. Genetic analysis of Netrin genes in Drosophila: Netrins guide CNS commissural axons and peripheral motor axons. Neuron 17, 203–215 (1996).

    Article  CAS  PubMed  Google Scholar 

  13. Hiramoto, M., Hiromi, Y., Giniger, E. & Hotta, Y. The Drosophila Netrin receptor Frazzled guides axons by controlling Netrin distribution. Nature 406, 886–889 (2000).

    Article  CAS  PubMed  Google Scholar 

  14. Keleman, K. & Dickson, B. Short- and long-range repulsion by the Drosophila Unc5 netrin receptor. Neuron 32, 605–617 (2001).

    Article  CAS  PubMed  Google Scholar 

  15. Colon-Ramos, D.A., Margeta, M. & Shen, K. Glia promote local synaptogenesis through UNC-6 (netrin) signaling in C. elegans. Science 318, 103–106 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Teichmann, H.M. & Shen, K. UNC-6 and UNC-40 promote dendritic growth through PAR-4 in Caenorhabditis elegans neurons. Nat. Neurosci. 14, 165–172 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  17. Park, J. et al. A conserved juxtacrine signal regulates synaptic partner recognition in Caenorhabditis elegans. Neural Develop. 6, 28 (2011).

    Article  CAS  Google Scholar 

  18. Chan, S.S. et al. UNC-40, a C. elegans homolog of DCC (Deleted in Colorectal Cancer), is required in motile cells responding to UNC-6 netrin cues. Cell 87, 187–195 (1996).

    Article  CAS  PubMed  Google Scholar 

  19. Leonardo, E.D. et al. Vertebrate homologues of C. elegans UNC-5 are candidate netrin receptors. Nature 386, 833–838 (1997).

    Article  CAS  PubMed  Google Scholar 

  20. Smith, C.J. et al. Time-lapse imaging and cell-specific expression profiling reveal dynamic branching and molecular determinants of a multi-dendritic nociceptor in C. elegans. Dev. Biol. 345, 18–33 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Albeg, A. et al. C. elegans multi-dendritic sensory neurons: morphology and function. Mol. Cell Neurosci. 46, 308–317 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  22. Oren-Suissa, M., Hall, D.H., Treinin, M., Shemer, G. & Podbilewicz, B. The fusogen EFF-1 controls sculpting of mechanosensory dendrites. Science 328, 1285–1288 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Hall, D.H. & Treinin, M. How does morphology relate to function in sensory arbors? Trends Neurosci. 34, 443–451 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Aguirre-Chen, C., Bülow, H.E. & Kaprielian, Z. C. elegans bicd-1, homolog of the Drosophila dynein accessory factor Bicaudal D, regulates the branching of PVD sensory neuron dendrites. Development 138, 507–518 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Ishii, N., Wadsworth, W., Stern, B., Culotti, J. & Hedgecock, E. UNC-6, a laminin-related protein, guides cell and pioneer axon migrations in C. elegans. Neuron 9, 873–881 (1992).

    Article  CAS  PubMed  Google Scholar 

  26. Schwarz, V., Pan, J., Voltmer-Irsch, S. & Hutter, H. IgCAMs redundantly control axon outgrowth in Caenorhabditis elegans. Neural Develop. 4, 13 (2009).

    Article  Google Scholar 

  27. Adler, C.E., Fetter, R. & Bargmann, C. UNC-6/Netrin induces neuronal asymmetry and defines the site of axon formation. Nat. Neurosci. 9, 511–518 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Watson, J.D. et al. Complementary RNA amplification methods enhance microarray identification of transcripts expressed in the C. elegans nervous system. BMC Genomics 9, 84 (2008).

    Article  PubMed  PubMed Central  Google Scholar 

  29. Xu, Z., Li, H. & Wadsworth, W. The roles of multiple UNC-40 (DCC) receptor-mediated signals in determining neuronal asymmetry induced by the UNC-6 (netrin) ligand. Genetics 183, 941–949 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Yang, L., Garbe, D. & Bashaw, G. A frazzled/DCC-dependent transcriptional switch regulates midline axon guidance. Science 324, 944–947 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Tsalik, E.L. & Hobert, O. Functional mapping of neurons that control locomotory behavior in Caenorhabditis elegans. J. Neurobiol. 56, 178–197 (2003).

    Article  PubMed  Google Scholar 

  32. Eastman, C., Horvitz, H.R. & Jin, Y. Coordinated transcriptional regulation of the unc-25 glutamic acid decarboxylase and the unc-47 GABA vesicular transporter by the Caenorhabditis elegans UNC-30 homeodomain protein. J. Neurosci. 19, 6225–6234 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Leung-Hagesteijn, C. et al. UNC-5, a transmembrane protein with immunoglobulin and thrombospondin type 1 domains, guides cell and pioneer axon migrations in C. elegans. Cell 71, 289–299 (1992).

    Article  CAS  PubMed  Google Scholar 

  34. Killeen, M. et al. UNC-5 function requires phosphorylation of cytoplasmic tyrosine 482, but its UNC-40-independent functions also require a region between the ZU-5 and death domains. Dev. Biol. 251, 348–366 (2002).

    Article  CAS  PubMed  Google Scholar 

  35. Asakura, T., Ogura, K. & Goshima, Y. UNC-6 expression by the vulval precursor cells of Caenorhabditis elegans is required for the complex axon guidance of the HSN neurons. Dev. Biol. 304, 800–810 (2007).

    Article  CAS  PubMed  Google Scholar 

  36. Bashaw, G.J. & Goodman, C.S. Chimeric axon guidance receptors: the cytoplasmic domains of slit and netrin receptors specify attraction versus repulsion. Cell 97, 917–926 (1999).

    Article  CAS  PubMed  Google Scholar 

  37. Gitai, Z., Yu, T., Lundquist, E., Tessier-Lavigne, M. & Bargmann, C. The netrin receptor UNC-40/DCC stimulates axon attraction and outgrowth through enabled and, in parallel, Rac and UNC-115/AbLIM. Neuron 37, 53–65 (2003).

    Article  CAS  PubMed  Google Scholar 

  38. Hiramoto, M. & Hiromi, Y. ROBO directs axon crossing of segmental boundaries by suppressing responsiveness to relocalized Netrin. Nat. Neurosci. 9, 58–66 (2006).

    Article  CAS  PubMed  Google Scholar 

  39. Kuzina, I., Song, J.K. & Giniger, E. How Notch establishes longitudinal axon connections between successive segments of the Drosophila CNS. Development 138, 1839–1849 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Alexander, M. et al. An UNC-40 pathway directs postsynaptic membrane extension in Caenorhabditis elegans. Development 136, 911–922 (2009).

    Article  CAS  PubMed  Google Scholar 

  41. MacNeil, L., Hardy, W., Pawson, T., Wrana, J. & Culotti, J. UNC-129 regulates the balance between UNC-40 dependent and independent UNC-5 signaling pathways. Nat. Neurosci. 12, 150–155 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Merz, D.C., Zheng, H., Killeen, M.T., Krizus, A. & Culotti, J.G. Multiple signaling mechanisms of the UNC-6/netrin receptors UNC-5 and UNC-40/DCC in vivo. Genetics 158, 1071–1080 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Chang, C. et al. MIG-10/lamellipodin and AGE-1/PI3K promote axon guidance and outgrowth in response to slit and netrin. Curr. Biol. 16, 854–862 (2006).

    Article  CAS  PubMed  Google Scholar 

  44. Quinn, C.C., Pfeil, D. & Wadsworth, W. CED-10/Rac1 mediates axon guidance by regulating the asymmetric distribution of MIG-10/lamellipodin. Curr. Biol. 18, 808–813 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Fleming, T. et al. The role of C. elegans Ena/VASP homolog UNC-34 in neuronal polarity and motility. Dev. Biol. 344, 94–106 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Grueber, W.B. & Sagasti, A. Self-avoidance and tiling: mechanisms of dendrite and axon spacing. Cold Spring Harb. Perspect. Biol. 2, a001750 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  47. Emoto, K., Parrish, J., Jan, L. & Jan, Y.-N. The tumour suppressor Hippo acts with the NDR kinases in dendritic tiling and maintenance. Nature 443, 210–213 (2006).

    Article  CAS  PubMed  Google Scholar 

  48. Brenner, S. The genetics of Caenorhabditis elegans. Genetics 77, 71–94 (1974).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Poon, V.Y., Klassen, M. & Shen, K. UNC-6/netrin and its receptor UNC-5 locally exclude presynaptic components from dendrites. Nature 455, 669–673 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Sulston, J.E. & Horvitz, H. Post-embryonic cell lineages of the nematode, Caenorhabditis elegans. Dev. Biol. 56, 110–156 (1977).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank C. Bargmann (Rockefeller University) for unc-86UNC-40GFP, hsp16.2UNC-6HA and the CX6488 strain; W. Wadsworth (Rutgers University) for the pIM97 unc-6 expression construct and unc-6(rh46); K. Shen (Stanford University) for constructs used to make pCJS01, F49H12.4gatewaymcherry; P. Roy (University of Toronto) for the unc-40(e271) sequence; Y. Goshima (Yokohama City University) for ghIs9; J. Culotti (Mount Sinai Hospital, Toronto) for the unc-5 rescue construct and for the modified UNC-5 protein strains used for structure–function analysis and members of the D.M.M., R.B. and D.A.C.-R. laboratories for technical advice and for comments on the manuscript. Some of the strains used in this work were provided by the C. elegans Genetics Center, which is supported by the US National Institutes of Health (NIH) National Center for Research Resources. This work was supported by NIH R01 NS26115 (D.M.M.), NIH R21 NS06882 (D.M.M.), NIH F31 NS071801 (C.J.S.), NIH R00 NS057931 (D.A.C.-R.), the Klingenstein Foundation and an Alfred P. Sloan Foundation fellowship (D.A.C.-R.), and NIH MBRS SC3 GM089595 (M.K.V.).

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Authors and Affiliations

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Contributions

C.J.S. and D.M.M. designed the experiments. C.J.S. performed experiments with advice from D.M.M. J.D.W. helped with the phenotypic analysis of UNC-6 signaling mutants. D.A.C.-R. and M.K.V. provided reagents to test the cell-specific requirement of UNC-40 and UNC-6 and provided advice. C.J.S. and D.M.M. wrote the paper with input from coauthors.

Corresponding author

Correspondence to David M Miller III.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–9, Supplementary Table 1 (PDF 870 kb)

Supplementary Movie 1

Wild-type self-avoidance. Time-lapse confocal movie of PVD::GFP in wild-type background. 3° dendrites contact but quickly retract (arrows). Note the intervening distance between 3O dendrites at the end of the movie is comparable to distance visualized in mature PVD neurons. Arrows indicate locations of contact-dependent self-avoidance. (MOV 8295 kb)

Supplementary Movie 2

Self-avoidance defect in unc-40(e271). Time-lapse confocal movie of PVD::GFP in unc-40(e271). 3° dendrites grow toward each other but upon contact fail to retract. Arrow indicates location of failed self-avoidance. (MOV 1892 kb)

Supplementary Movie 3

Self-avoidance defect in unc-5(e152). Imaging of unc-5(e152) shows PVD dendrites fail to retract after contact. Arrow indicates location of failed self-avoidance. (MOV 33275 kb)

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Smith, C., Watson, J., VanHoven, M. et al. Netrin (UNC-6) mediates dendritic self-avoidance. Nat Neurosci 15, 731–737 (2012). https://doi.org/10.1038/nn.3065

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