Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Coupling of NF-protocadherin signaling to axon guidance by cue-induced translation

Nature Neuroscience volume 16, pages 166–173 (2013)Cite this article

Subjects

Abstract

Cell adhesion molecules and diffusible cues both regulate axon pathfinding, yet how these two modes of signaling interact is poorly understood. The homophilic cell adhesion molecule NF-protocadherin (NFPC) is expressed in the mid-dorsal optic tract neuroepithelium and in the axons of developing retinal ganglion cells (RGC) in Xenopus laevis. Here we report that targeted disruption of NFPC function in RGC axons or the optic tract neuroepithelium results in unexpectedly localized pathfinding defects at the caudal turn in the mid-optic tract. Semaphorin 3A (Sema3A), which lies adjacent to this turn, stimulates rapid, protein synthesis–dependent increases in growth cone NFPC and its cofactor, TAF1, in vitro. In vivo, growth cones exhibit marked increases in NFPC translation reporter activity in this mid-optic tract region that are attenuated by blocking neuropilin-1 function. Our results suggest that translation-linked coupling between regionally localized diffusible cues and cell adhesion can help axons navigate discrete segments of the pathway.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Dominant negative NFPC signaling disrupts retinal axon growth and guidance in the optic tract.
Figure 2: Depletion of NFPC or TAF1 disrupts retinal axon guidance in the optic tract.
Figure 3: Live imaging of retinal axons shows that NFPC function is required at the caudal turn.
Figure 4: Homophilic NFPC interactions are required at the caudal turn.
Figure 5: NFPC is required in the neuroepithelium of the optic tract for guidance.
Figure 6: Sema3A increases NFPC levels in retinal growth cones.
Figure 7: Sema3A induces local translation of NFPC mRNA.
Figure 8: In vivo imaging shows that NFPC-translation reporter activity is upregulated at the caudal turn and depends partly on NP-1–Sema signaling.

Similar content being viewed by others

References

  1. Chien, C.B., Rosenthal, D.E., Harris, W.A. & Holt, C.E. Navigational errors made by growth cones without filopodia in the embryonic Xenopus brain. Neuron 11, 237–251 (1993).

    Article  CAS  Google Scholar 

  2. Harris, W.A. Local positional cues in the neuroepithelium guide retinal axons in embryonic Xenopus brain. Nature 339, 218–221 (1989).

    Article  CAS  Google Scholar 

  3. Atkinson-Leadbeater, K. et al. Dynamic expression of axon guidance cues required for optic tract development is controlled by fibroblast growth factor signaling. J. Neurosci. 30, 685–693 (2010).

    Article  CAS  Google Scholar 

  4. Campbell, D.S. et al. Semaphorin 3A elicits stage-dependent collapse, turning, and branching in Xenopus retinal growth cones. J. Neurosci. 21, 8538–8547 (2001).

    Article  CAS  Google Scholar 

  5. Piper, M. et al. Signaling mechanisms underlying Slit2-induced collapse of Xenopus retinal growth cones. Neuron 49, 215–228 (2006).

    Article  CAS  Google Scholar 

  6. Arikkath, J. & Reichardt, L.F. Cadherins and catenins at synapses: roles in synaptogenesis and synaptic plasticity. Trends Neurosci. 31, 487–494 (2008).

    Article  CAS  Google Scholar 

  7. Hirano, S., Suzuki, S. & Redies, C. The cadherin superfamily in neural development: diversity, function and interaction with other molecules. Front. Biosci. 8, d306–d355 (2003).

    Article  CAS  Google Scholar 

  8. Takeichi, M. The cadherin superfamily in neuronal connections and interactions. Nat. Rev. Neurosci. 8, 11–20 (2007).

    Article  CAS  Google Scholar 

  9. Bradley, R.S., Espeseth, A. & Kintner, C. NF-protocadherin, a novel member of the cadherin superfamily, is required for Xenopus ectodermal differentiation. Curr. Biol. 8, 325–334 (1998).

    Article  CAS  Google Scholar 

  10. Heggem, M.A. & Bradley, R.S. The cytoplasmic domain of Xenopus NF-protocadherin interacts with TAF1/set. Dev. Cell 4, 419–429 (2003).

    Article  CAS  Google Scholar 

  11. Rashid, D., Newell, K., Shama, L. & Bradley, R. A requirement for NF-protocadherin and TAF1/Set in cell adhesion and neural tube formation. Dev. Biol. 291, 170–181 (2006).

    Article  CAS  Google Scholar 

  12. Piper, M., Dwivedy, A., Leung, L., Bradley, R.S. & Holt, C.E. NF-protocadherin and TAF1 regulate retinal axon initiation and elongation in vivo. J. Neurosci. 28, 100–105 (2008).

    Article  CAS  Google Scholar 

  13. Campbell, D.S. & Holt, C.E. Chemotropic responses of retinal growth cones mediated by rapid local protein synthesis and degradation. Neuron 32, 1013–1026 (2001).

    Article  CAS  Google Scholar 

  14. Leung, K.-M. et al. Asymmetrical β-actin mRNA translation in growth cones mediates attractive turning to netrin-1. Nat. Neurosci. 9, 1247–1256 (2006).

    Article  CAS  Google Scholar 

  15. Lin, A.C. & Holt, C.E. Local translation and directional steering in axons. EMBO J. 26, 3729–3736 (2007).

    Article  CAS  Google Scholar 

  16. Wu, K.Y. et al. Local translation of RhoA regulates growth cone collapse. Nature 436, 1020–1024 (2005).

    Article  CAS  Google Scholar 

  17. Yao, J., Sasaki, Y., Wen, Z., Bassell, G.J. & Zheng, J.Q. An essential role for β-actin mRNA localization and translation in Ca2+-dependent growth cone guidance. Nat. Neurosci. 9, 1265–1273 (2006).

    Article  CAS  Google Scholar 

  18. Falk, J. et al. Electroporation of cDNA/morpholinos to targeted areas of embryonic CNS in Xenopus. BMC Dev. Biol. 7, 107 (2007).

    Article  Google Scholar 

  19. Holt, C.E. A single-cell analysis of early retinal ganglion cell differentiation in Xenopus: from soma to axon tip. J. Neurosci. 9, 3123–3145 (1989).

    Article  CAS  Google Scholar 

  20. Piper, M., Salih, S., Weinl, C., Holt, C.E. & Harris, W.A. Endocytosis-dependent desensitization and protein synthesis-dependent resensitization in retinal growth cone adaptation. Nat. Neurosci. 8, 179–186 (2005).

    Article  CAS  Google Scholar 

  21. Lin, A.C. et al. Cytoplasmic polyadenylation and cytoplasmic polyadenylation element-dependent mRNA regulation are involved in Xenopus retinal axon development. Neural Dev. 4, 8 (2009).

    Article  Google Scholar 

  22. Leung, K.-M. & Holt, C.E. Live visualization of protein synthesis in axonal growth cones by microinjection of photoconvertible Kaede into Xenopus embryos. Nat. Protoc. 3, 1318–1327 (2008).

    Article  CAS  Google Scholar 

  23. Uemura, M., Nakao, S., Suzuki, S.T., Takeichi, M. & Hirano, S. OL-protocadherin is essential for growth of striatal axons and thalamocortical projections. Nat. Neurosci. 10, 1151–1159 (2007).

    Article  CAS  Google Scholar 

  24. Garrett, A.M., Schreiner, D., Lobas, M.A. & Weiner, J.A. γ-Protocadherins control cortical dendrite arborization by regulating the activity of a FAK/PKC/MARCKS signaling pathway. Neuron 74, 269–276 (2012).

    Article  CAS  Google Scholar 

  25. Lefebvre, J.L., Kostadinov, D., Chen, W.V., Maniatis, T. & Sanes, J.R. Protocadherins mediate dendritic self-avoidance in the mammalian nervous system. Nature 488, 517–521 (2012).

    Article  CAS  Google Scholar 

  26. Yasuda, S. et al. Activity-induced protocadherin arcadlin regulates dendritic spine number by triggering N-cadherin endocytosis via TAO2β and p38 MAP kinases. Neuron 56, 456–471 (2007).

    Article  CAS  Google Scholar 

  27. Garrett, A.M. & Weiner, J.A. Control of CNS synapse development by γ-protocadherin-mediated astrocyte-neuron contact. J. Neurosci. 29, 11723–11731 (2009).

    Article  CAS  Google Scholar 

  28. Williams, E.O. et al. Delta protocadherin 10 is regulated by activity in the mouse main olfactory system. Front. Neural Circuits 5, 9 (2011).

    Article  CAS  Google Scholar 

  29. Lefebvre, J.L., Zhang, Y., Meister, M., Wang, X. & Sanes, J.R. γ-Protocadherins regulate neuronal survival but are dispensable for circuit formation in retina. Development 135, 4141–4151 (2008).

    Article  CAS  Google Scholar 

  30. Castellani, V., De Angelis, E., Kenwrick, S. & Rougon, G. Cis and trans interactions of L1 with neuropilin-1 control axonal responses to semaphorin 3A. EMBO J. 21, 6348–6357 (2002).

    Article  CAS  Google Scholar 

  31. Castellani, V., Falk, J. & Rougon, G. Semaphorin3A-induced receptor endocytosis during axon guidance responses is mediated by L1 CAM. Mol. Cell. Neurosci. 26, 89–100 (2004).

    Article  CAS  Google Scholar 

  32. Bechara, A. et al. FAK-MAPK-dependent adhesion disassembly downstream of L1 contributes to semaphorin3A-induced collapse. EMBO J. 27, 1549–1562 (2008).

    Article  CAS  Google Scholar 

  33. Falk, J. et al. Dual functional activity of semaphorin 3B is required for positioning the anterior commissure. Neuron 48, 63–75 (2005).

    Article  Google Scholar 

  34. Wolman, M.A., Regnery, A.M., Becker, T., Becker, C.G. & Halloran, M.C. Semaphorin3D regulates axon–axon interactions by modulating levels of L1 cell adhesion molecule. J. Neurosci. 27, 9653–9663 (2007).

    Article  CAS  Google Scholar 

  35. Nawabi, H. et al. A midline switch of receptor processing regulates commissural axon guidance in vertebrates. Genes Dev. 24, 396–410 (2010).

    Article  CAS  Google Scholar 

  36. Kuwajima, T. et al. Optic chiasm presentation of Semaphorin6D in the context of Plexin-A1 and Nr-CAM promotes retinal axon midline crossing. Neuron 74, 676–690 (2012).

    Article  CAS  Google Scholar 

  37. Lemmon, V., Burden, S.M., Payne, H.R., Elmslie, G.J. & Hlavin, M.L. Neurite growth on different substrates: permissive versus instructive influences and the role of adhesive strength. J. Neurosci. 12, 818–826 (1992).

    Article  CAS  Google Scholar 

  38. Goodman, C.S. Mechanisms and molecules that control growth cone guidance. Annu. Rev. Neurosci. 19, 341–377 (1996).

    Article  CAS  Google Scholar 

  39. Barnes, S.H., Price, S.R., Wentzel, C. & Guthrie, S.C. Cadherin-7 and cadherin-6B differentially regulate the growth, branching and guidance of cranial motor axons. Development 137, 805–814 (2010).

    Article  CAS  Google Scholar 

  40. Shima, Y. et al. Opposing roles in neurite growth control by two seven-pass transmembrane cadherins. Nat. Neurosci. 10, 963–969 (2007).

    Article  CAS  Google Scholar 

  41. Nakao, S., Platek, A., Hirano, S. & Takeichi, M. Contact-dependent promotion of cell migration by the OL-protocadherin-Nap1 interaction. J. Cell Biol. 182, 395–410 (2008).

    Article  CAS  Google Scholar 

  42. Brittis, P.A., Lu, Q. & Flanagan, J.G. Axonal protein synthesis provides a mechanism for localized regulation at an intermediate target. Cell 110, 223–235 (2002).

    Article  CAS  Google Scholar 

  43. Kuwako, K. et al. Neural RNA-binding protein Musashi1 controls midline crossing of precerebellar neurons through posttranscriptional regulation of Robo3/Rig-1 expression. Neuron 67, 407–421 (2010).

    Article  CAS  Google Scholar 

  44. Zivraj, K.H. et al. Subcellular profiling reveals distinct and developmentally regulated repertoire of growth cone mRNAs. J. Neurosci. 30, 15464–15478 (2010).

    Article  CAS  Google Scholar 

  45. Seo, S.B. et al. Regulation of histone acetylation and transcription by INHAT, a human cellular complex containing the Set oncoprotein. Cell 104, 119–130 (2001).

    Article  CAS  Google Scholar 

  46. Nieuwkoop, F.J. & Faber, J. Normal Table of Xenopus laevis (Daudin) (North-Holland, 1967).

  47. Scotto-Lavino, E., Du, G. & Frohman, M.A. 3′ end cDNA amplification using classic RACE. Nat. Protoc. 1, 2742–2745 (2006).

    Article  CAS  Google Scholar 

  48. Leung, L. & Holt, C.E. Imaging axon pathfinding in Xenopus in vivo. Cold Spring Harb. Protoc. 2012, 984–991 (2012).

    PubMed  Google Scholar 

  49. Wizenmann, A. et al. Extracellular engrailed participates in the topographic guidance of retinal axons in vivo. Neuron 64, 355–366 (2009).

    Article  CAS  Google Scholar 

  50. Luo, Y., Raible, D. & Raper, J.A. Collapsin: a protein in brain that induces the collapse and paralysis of neuronal growth cones. Cell 75, 217–227 (1993).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank R. Bradley (Montana State University) for kind gifts of NFPC antibody and sharing constructs, S. McFarlane for advice on the Slit1 in situ hybridization, and F. van Horck for NFPC in situ hybridization data. We thank A. Pungaliya, A. McNabb, T. Dyl, G. Stooke-Vaughan, C. Purmann, K. Holmes, H. Lynn, G. Lupo, D. Maurus and N. Coutts for technical assistance. We thank members of the Harris and Holt laboratories for assistance and comments on the manuscript. This work was funded by Wellcome Trust Programme grant no. 085314/Z/08/Z (C.E.H., W.A.H.), a UK Medical Research Council Doctoral Training Grant (L.C.L.), a Wellcome Trust Studentship (V.U.), the National Science and Engineering Research Council of Canada and the Alberta Heritage Foundation for Medical Research (M-L.B.), a UK Medical Research Council studentship (T.G.B.) and an EMBO fellowship (A.C.L.).

Author information

Author notes

  1. Louis C Leung, Marie-Laure Baudet & Timothy G Bayley

    Present address: Present addresses: Department of Psychiatry & Behavioral Sciences, Stanford University, Stanford, California, USA (L.C.L.), Centre for Integrative Biology (CIBIO), University of Trento, Mattarello, Italy (M.-L.B.) and Department of Zoology, University of Cambridge, Cambridge, UK (T.G.B.).,

  2. Louis C Leung and Vasja Urbančič: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Physiology, Development and Neuroscience, University of Cambridge, Cambridge, UK

    Louis C Leung, Vasja Urbančič, Marie-Laure Baudet, Asha Dwivedy, Timothy G Bayley, Aih Cheun Lee, William A Harris & Christine E Holt

Authors
  1. Louis C Leung
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Vasja Urbančič
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Marie-Laure Baudet
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Asha Dwivedy
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Timothy G Bayley
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Aih Cheun Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. William A Harris
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Christine E Holt
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

L.C.L., C.E.H. and W.A.H. conceived the project, designed the experiments and wrote the manuscript. L.C.L. produced the constructs and peptides, and generated and analyzed all the data in Figures 1,2,3,4,5,6,7, and Supplementary Figures 1–7 unless otherwise stated. V.U. and T.G.B. performed and analyzed the in vivo Kaede translation reporter experiments in Figure 8a,b. V.U. conducted the combined antibody in vivo Kaede reporter experiments in Figure 8c,d and A.D. assisted with in vivo antibody experiments in Supplementary Figure 4e. M.-L.B. performed in situ hybridization in Supplementary Figure 3. A.C.L. performed the western blots in Supplementary Figure 1a,b and the cycloheximide experiments in Supplementary Figure 5n.

Corresponding author

Correspondence to Christine E Holt.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8 and Supplementary Table 1 (PDF 21990 kb)

Supplementary Video 1

Timelapse movie of a GAP-GFP axon. An example of a timelapse movie of GAP-GFP axons navigating through the Xenopus optic tract (lateral view), from which the kymograph in Figure 3a was derived. Each frame represents 3 min elapsed and the movie is played at 5 frames per second. This movie was acquired at 20× magnification. Scale bar, 10 μm. (MOV 517 kb)

Supplementary Video 2

Timelapse movie of a NFΔEeGFP axon. An example of a timelapse movie of an NFΔEeGFP axon navigating through the Xenopus optic tract, from which the kymograph in Figure 3b was derived. Each frame represents 3 min elapsed and the movie is played at 5 frames per second. The growth cone has stalled at the caudal turn, and once it has made the turn growth in the dorsal tract is appreciably slower. This movie was acquired at 20× magnification. Scale bar, 10 μm. (MOV 574 kb)

Supplementary Video 3

Timelapse movie of Kaede-NFPC-3′UTR translation activity in the ventral optic tract. An example of a retinal growth cone in the ventral optic tract (arrowheads), expressing Kaede-NFPC-3′UTR, in which the green signal is not recovered over the course of 30 min, indicative of the absence of NFPC-3′UTR–driven translation in the ventral optic tract. Each frame represents 5 min and the movie is played at 1.5 frames per second. The timelapse was acquired at 60× magnification. Scale bar, 5 μm. (MOV 329 kb)

Supplementary Video 4

Timelapse movie of Kaede-NFPC-3′UTR translation activity in the mid-optic tract. Photoconversion and post-conversion imaging of Kaede in retinal growth cones expressing Kaede-NFPC-′UTR in the mid-optic tract. Arrowheads highlight a single growth cone in which the green signal recovers over the course of 30 min after photoconversion, indicative of the translation of Kaede-NFPC-3′UTR. Magnification and playback speed are as in Supplementary Movie 3. (MOV 330 kb)

Supplementary Video 5

Timelapse movie of Kaede-NFPC-3′UTR translation in brains treated with a control antibody. An example of a retinal growth cone in mid-optic tract with Kaede-green signal recovery after photoconversion, following incubation with a control (IgG2B) antibody. Magnification and playback speed are as in Supplementary Video 3. (MOV 459 kb)

Supplementary Video 6

Timelapse movie of Kaede-NFPC-3′UTR translation in brains treated with a neuropilin-1 function blocking antibody. An example of a retinal growth cone in mid-optic tract with a reduced level of Kaede-green signal recovery after photoconversion following incubation with a neuropilin-1 function-blocking antibody, indicative of decreased translation of Kaede-NFPC-3′UTR upon anti–NP-1 treatment. Magnification and playback speed are as in Supplementary Video 3. (MOV 361 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Leung, L., Urbančič, V., Baudet, ML. et al. Coupling of NF-protocadherin signaling to axon guidance by cue-induced translation. Nat Neurosci 16, 166–173 (2013). https://doi.org/10.1038/nn.3290

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn.3290

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing