Multiple Regulation of Roundabout (Robo) Phosphorylation in a Heterologous Cell System

  • Park, Hwan-Tae (Department of Physiology and Institute of Medical Science, College of Medicine, Dong-A University)
  • Published : 2004.04.21

Abstract

Roundabout (Robo) is the transmembrane receptor for slit, the neuronal guidance molecule. In this study, the tyrosine phosphorylation of Robo was observed in Robo-transfected human embryonic kidney cells and developing rat brains, and found to be increased by the treatment with protein kinase A activator, forskolin. In contrast, protein kinase C activation by phorbol-12-myristate-13-acetate decreased the phosphorylation of Robo. Intracellular calcium was required for the tyrosine phosphorylation. Furthermore, the transfection of an Eph receptor tyrosine kinase dramatically enhanced the tyrosine phosphorylation. These findings indicate that the tyrosine phosphorylation of Robo is regulated by multiple mechanisms, and that Eph receptor kinases may play a role in the regulation of tyrosine phosphorylation of Robo in the rat brain.

Keywords

References

  1. Bagri A, Marin O, Plump AS, Mak J, Pleasure SJ, Rubenstein JL, Tessier-Lavigne M. Slit proteins prevent midline crossing and determine the dorsoventral position of major axonal pathways in the mammalian forebrain. Neuron 33: 233-248, 2002 https://doi.org/10.1016/S0896-6273(02)00561-5
  2. Bashaw GJ, Kidd T, Murray D, Pawson T, Goodman CS. Repulsive axon guidance: Abelson and Enabled play opposing roles downstream of the roundabout receptor. Cell 101: 703-715, 2000 https://doi.org/10.1016/S0092-8674(00)80883-1
  3. Brose K, Bland KS, Wang KH, Arnott D, Henzel W, Goodman CS, Tessier-Lavigne M, Kidd T. Slit proteins bind Robo receptors and have an evolutionarily conserved role in repulsive axon guidance. Cell 96: 795-806, 1999 https://doi.org/10.1016/S0092-8674(00)80590-5
  4. Castellani V, Chedotal A, Schachner M, Faivre-Sarrailh C, Rougon G. Analysis of the L1-deficient mouse phenotype reveals crosstalk between Sema3A and L1 signaling pathways in axonal guidance. Neuron 27: 237-249, 2000 https://doi.org/10.1016/S0896-6273(00)00033-7
  5. Davy A, Gale NW, Murray EW, Klinghoffer RA, Soriano P, Feuerstein C, Robbins SM. Compartmentalized signaling by GPI-anchored ephrin-A5 requires the Fyn tyrosine kinase to regulate cellular adhesion. Genes Dev 13: 3125-3135, 1999 https://doi.org/10.1101/gad.13.23.3125
  6. Erskine L, Williams SE, Brose K, Kidd T, Rachel RA, Goodman CS, Tessier-Lavigne M, Mason CA. Retinal ganglion cell axon guidance in the mouse optic chiasm: expression and function of robos and slits. J Neurosci 20: 4975-4982, 2000
  7. Gao Z, Chen T, Weber MJ, Linden J. A2B adenosine and P2Y2 receptors stimulate mitogen-activated protein kinase in human embryonic kidney-293 cells. cross-talk between cyclic AMP and protein kinase c pathways. J Biol Chem 274: 5972-5980, 1999 https://doi.org/10.1074/jbc.274.9.5972
  8. Godenschwege TA, Simpson JH, Shan X, Bashaw GJ, Goodman CS, Murphey RK. Ectopic expression in the giant fiber system of Drosophila reveals distinct roles for roundabout (Robo), Robo2, and Robo3 in dendritic guidance and synaptic connectivity. J Neurosci 22: 3117-3129, 2002
  9. Hao JC, Yu TW, Fujisawa K, Culotti JG, Gengyo-Ando K, Mitani S, Moulder G, Barstead R, Tessier-Lavigne M, Bargmann CI. C. elegans slit acts in midline, dorsal-ventral, and anteriorposterior guidance via the SAX-3/Robo receptor. Neuron 32: 25-38, 2001 https://doi.org/10.1016/S0896-6273(01)00448-2
  10. Hong K, Nishiyama M, Henley J, Tessier-Lavigne M, Poo M. Calcium signalling in the guidance of nerve growth by netrin-1. Nature 403: 93-98, 2000 https://doi.org/10.1038/47507
  11. Jeon ES, Bae KW, Park HT. Fyn tyrosine kinase-mediated tyrosine phosphorylation of Roundabout (Robo), the slit receptor. Mol Brain Res (submitted)
  12. Kidd T, Bland KS, Goodman CS. Slit is the midline repellent for the robo receptor in Drosophila. Cell 96: 785-794, 1999 https://doi.org/10.1016/S0092-8674(00)80589-9
  13. Kidd T, Brose K, Mitchell KJ, Fetter RD, Tessier-Lavigne M, Goodman CS, Tear G. Roundabout controls axon crossing of the CNS midline and defines a novel subfamily of evolutionarily conserved guidance receptors. Cell 92: 205-215, 1998 https://doi.org/10.1016/S0092-8674(00)80915-0
  14. Kramer SG, Kidd T, Simpson JH, Goodman CS. Switching repulsion to attraction: changing responses to slit during transition in mesoderm migration. Science 292: 737-740, 2001 https://doi.org/10.1126/science.1058766
  15. Kullander K, Klein R. Mechanisms and functions of Eph and ephrin signalling. Nat Rev Mol Cell Biol 3: 475-486, 2002 https://doi.org/10.1038/nrm856
  16. Li HS, Chen JH, Wu W, Fagaly T, Zhou L, Yuan W, Dupuis S, Jiang ZH, Nash W, Gick C, Ornitz DM, Wu JY, Rao Y. Vertebrate slit, a secreted ligand for the transmembrane protein roundabout, is a repellent for olfactory bulb axons. Cell 96: 807- 818, 1999 https://doi.org/10.1016/S0092-8674(00)80591-7
  17. Ming GL, Song HJ, Berninger B, Holt CE, Tessier-Lavigne M, Poo MM. cAMP-dependent growth cone guidance by netrin-1. Neuron 19: 1225-1235, 1997 https://doi.org/10.1016/S0896-6273(00)80414-6
  18. Nguyen-Ba-Charvet KT, Chedotal A. Role of Slit proteins in the vertebrate brain. J Physiol Paris 96: 91-98, 2002 https://doi.org/10.1016/S0928-4257(01)00084-5
  19. Nishiyama M, Hoshino A, Tsai L, Henley JR, Goshima Y, Tessier-Lavigne M, Poo MM, Hong K. Cyclic AMP/GMP-dependent modulation of Ca2$^+$ channels sets the polarity of nerve growth-cone turning. Nature 424: 990-995, 2003 https://doi.org/10.1038/424990a
  20. Park HT, Wu J, Rao Y. Molecular control of neuronal migration. Bioessays 24: 821-827, 2002 https://doi.org/10.1002/bies.10141
  21. Park KW, Morrison CM, Sorensen LK, Jones CA, Rao Y, Chien CB, Wu JY, Urness LD, Li DY, Robo4 is a vascular-specific receptor that inhibits endothelial migration. Dev Biol 261: 251- 267, 2003 https://doi.org/10.1016/S0012-1606(03)00258-6
  22. Serafini T, Colamarino SA, Leonardo ED, Wang H, Beddington R, Skarnes WC, Tessier-Lavigne M. Netrin-1 is required for commissural axon guidance in the developing vertebrate nervous system. Cell 87: 1001-1014, 1996 https://doi.org/10.1016/S0092-8674(00)81795-X
  23. Song H, Ming G, He Z, Lehmann M, McKerracher L, Tessier- Lavigne M, Poo M. Conversion of neuronal growth cone responses from repulsion to attraction by cyclic nucleotides. Science 281: 1515-1518, 1998 https://doi.org/10.1126/science.281.5382.1515
  24. Stein E, Tessier-Lavigne M. Hierarchical organization of guidance receptors: silencing of netrin attraction by slit through a Robo/ DCC receptor complex. Science 291: 1928-1938, 2001 https://doi.org/10.1126/science.1058445
  25. Steinle JJ, Meininger CJ, Forough R, Wu G, Wu MH, Granger HJ. Eph B4 receptor signaling mediates endothelial cell migration and proliferation via the phosphatidylinositol 3-kinase pathway. J Biol Chem 277: 43830-43835, 2002 https://doi.org/10.1074/jbc.M207221200
  26. Tessier-Lavigne M, Goodman CS. The molecular biology of axon guidance. Science 274: 1123-1133, 1996 https://doi.org/10.1126/science.274.5290.1123
  27. Wang B, Xiao Y, Ding BB, Zhang N, Yuan X, Gui L, Qian KX, Duan S, Chen Z, Rao Y, Geng JG. Induction of tumor angiogenesis by Slit-Robo signaling and inhibition of cancer growth by blocking Robo activity. Cancer Cell 4: 19-29, 2003 https://doi.org/10.1016/S1535-6108(03)00164-8
  28. Wang KH, Brose K, Arnott D, Kidd T, Goodman CS, Henzel W, Tessier-Lavigne M. Biochemical purification of a mammalian slit protein as a positive regulator of sensory axon elongation and branching. Cell 96: 771-784, 1999 https://doi.org/10.1016/S0092-8674(00)80588-7
  29. Whitford KL, Marillat V, Stein E, Goodman CS, Tessier-Lavigne M, Chedotal A, Ghosh A. Regulation of cortical dendrite development by Slit-Robo interactions. Neuron 33: 47-61, 2002 https://doi.org/10.1016/S0896-6273(01)00566-9
  30. Wong K, Park HT, Wu JY, Rao Y. Slit proteins: molecular guidance cues for cells ranging from neurons to leukocytes. Curr Opin Genet Dev 12: 583-591, 2002 https://doi.org/10.1016/S0959-437X(02)00343-X
  31. Wu JY, Feng L, Park HT, Havlioglu N, Wen L, Tang H, Bacon KB, Jiang Zh, Zhang Xc, Rao Y. The neuronal repellent Slit inhibits leukocyte chemotaxis induced by chemotactic factors. Nature 410: 948-952, 2001 https://doi.org/10.1038/35073616
  32. Xiang Y, Li Y, Zhang Z, Cui K, Wang S, Yuan XB, Wu CP, Poo MM, Duan S. Nerve growth cone guidance mediated by G protein-coupled receptors. Nat Neurosci 5: 843-848, 2002 https://doi.org/10.1038/nn899
  33. Yu HH, Zisch AH, Dodelet VC, Pasquale EB. Multiple signaling interactions of Abl and Arg kinases with the EphB2 receptor. Oncogene 20: 3995-4006, 2001 https://doi.org/10.1038/sj.onc.1204524