PDBsum entry 1w8a

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protein links
Signaling protein PDB id
Protein chain
189 a.a. *
Waters ×5
* Residue conservation analysis
PDB id:
Name: Signaling protein
Title: Third lrr domain of drosophila slit
Structure: Slit protein. Chain: a. Fragment: third lrr domain, residues 542-733. Engineered: yes
Source: Drosophila melanogaster. Organism_taxid: 7227. Expressed in: human. Expression_system_taxid: 9606. Expression_system_cell_line: 293-ebna.
2.8Å     R-factor:   0.199     R-free:   0.268
Authors: J.A.Howitt,N.J.Clout,E.Hohenester
Key ref:
J.A.Howitt et al. (2004). Binding site for Robo receptors revealed by dissection of the leucine-rich repeat region of Slit. EMBO J, 23, 4406-4412. PubMed id: 15496984 DOI: 10.1038/sj.emboj.7600446
17-Sep-04     Release date:   25-Oct-04    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P24014  (SLIT_DROME) -  Protein slit
1504 a.a.
189 a.a.
Key:    PfamA domain  Secondary structure  CATH domain


DOI no: 10.1038/sj.emboj.7600446 EMBO J 23:4406-4412 (2004)
PubMed id: 15496984  
Binding site for Robo receptors revealed by dissection of the leucine-rich repeat region of Slit.
J.A.Howitt, N.J.Clout, E.Hohenester.
Recognition of the large secreted protein Slit by receptors of the Robo family provides fundamental signals in axon guidance and other developmental processes. In Drosophila, Slit-Robo signalling regulates midline crossing and the lateral position of longitudinal axon tracts. We report the functional dissection of Drosophila Slit, using structure analysis, site-directed mutagenesis and in vitro assays. The N-terminal region of Slit consists of a tandem array of four independently folded leucine-rich repeat (LRR) domains, connected by disulphide-tethered linkers. All three Drosophila Robos were found to compete for a single highly conserved site on the concave face of the second LRR domain of Slit. We also found that this domain is sufficient for biological activity in a chemotaxis assay. Other Slit activities may require Slit dimerisation mediated by the fourth LRR domain. Our results show that a small portion of Slit is able to induce Robo signalling and indicate that the distinct functions of Drosophila Robos are encoded in their divergent cytosolic domains.
  Selected figure(s)  
Figure 2.
Figure 2 Structure of Slit LRR domains. (A) Cartoon drawing of the crystal structure of Slit D3. The N- and C-terminal caps are in violet and red, respectively, and LRRs 1 -5 are in cyan. Disulphide bridges are in yellow. The N- and C-terminus are labelled. (B) Sequence conservation of the concave faces of D1 -4. The domains are shown schematically in the orientation and colour scheme used in (A). The LRR consensus sequence is indicated on the left, running vertically from top to bottom. Within each domain, the LRRs are numbered horizontally and invariant residues at solvent-exposed positions X[1] -X[5] are shown. Four residues in D2 that were mutated in this study are highlighted.
Figure 4.
Figure 4 Conserved residues on the concave face of Slit D2 are important in Robo binding. (A) Sequence conservation mapped onto the concave face of a Slit D2 homology model (see text and Materials and methods). The colour scheme ranges from red (identity) to blue (no conservation). The orientation is similar to Figure 2A. Residues mutated in this study are labelled. (B -D) Binding of FLAG-tagged Robo D1 -5 (B), Robo2 D1 -5 (C) and Robo3 D1 -5 (D) to immobilised Slit D2 -3 proteins. Each experiment was carried out at least three times with similar results.
  The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: EMBO J (2004, 23, 4406-4412) copyright 2004.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21479274 E.Leonardi, S.Andreazza, S.Vanin, G.Busolin, C.Nobile, and S.C.Tosatto (2011).
A computational model of the LGI1 protein suggests a common binding site for ADAM proteins.
  PLoS One, 6, e18142.  
20697954 M.W.Poon, W.H.Tsang, S.O.Chan, H.M.Li, H.K.Ng, and M.M.Waye (2011).
Dyslexia-associated kiaa0319-like protein interacts with axon guidance receptor nogo receptor 1.
  Cell Mol Neurobiol, 31, 27-35.  
20144763 B.Spitzweck, M.Brankatschk, and B.J.Dickson (2010).
Distinct protein domains and expression patterns confer divergent axon guidance functions for Drosophila Robo receptors.
  Cell, 140, 409-420.  
20498081 R.Marlow, M.Binnewies, L.K.Sorensen, S.D.Monica, P.Strickland, E.C.Forsberg, D.Y.Li, and L.Hinck (2010).
Vascular Robo4 restricts proangiogenic VEGF signaling in breast.
  Proc Natl Acad Sci U S A, 107, 10520-10525.  
20206526 T.A.Evans, and G.J.Bashaw (2010).
Functional diversity of Robo receptor immunoglobulin domains promotes distinct axon guidance decisions.
  Curr Biol, 20, 567-572.  
19934013 C.E.Shiau, and M.Bronner-Fraser (2009).
N-cadherin acts in concert with Slit1-Robo2 signaling in regulating aggregation of placode-derived cranial sensory neurons.
  Development, 136, 4155-4164.  
19498462 E.Seiradake, A.C.von Philipsborn, M.Henry, M.Fritz, H.Lortat-Jacob, M.Jamin, W.Hemrika, M.Bastmeyer, S.Cusack, and A.A.McCarthy (2009).
Structure and functional relevance of the Slit2 homodimerization domain.
  EMBO Rep, 10, 736-741.
PDB code: 2wfh
19452560 K.L.Hindle, J.Bella, and S.C.Lovell (2009).
Quantitative analysis and prediction of curvature in leucine-rich repeat proteins.
  Proteins, 77, 342-358.  
18270976 A.Anitha, K.Nakamura, K.Yamada, S.Suda, I.Thanseem, M.Tsujii, Y.Iwayama, E.Hattori, T.Toyota, T.Miyachi, Y.Iwata, K.Suzuki, H.Matsuzaki, M.Kawai, Y.Sekine, K.Tsuchiya, G.Sugihara, Y.Ouchi, T.Sugiyama, K.Koizumi, H.Higashida, N.Takei, T.Yoshikawa, and N.Mori (2008).
Genetic analyses of roundabout (ROBO) axon guidance receptors in autism.
  Am J Med Genet B Neuropsychiatr Genet, 147, 1019-1027.  
18611862 A.Prasad, V.Paruchuri, A.Preet, F.Latif, and R.K.Ganju (2008).
Slit-2 induces a tumor-suppressive effect by regulating beta-catenin in breast cancer cells.
  J Biol Chem, 283, 26624-26633.  
18345009 C.A.Jones, N.R.London, H.Chen, K.W.Park, D.Sauvaget, R.A.Stockton, J.D.Wythe, W.Suh, F.Larrieu-Lahargue, Y.S.Mukouyama, P.Lindblom, P.Seth, A.Frias, N.Nishiya, M.H.Ginsberg, H.Gerhardt, K.Zhang, and D.Y.Li (2008).
Robo4 stabilizes the vascular network by inhibiting pathologic angiogenesis and endothelial hyperpermeability.
  Nat Med, 14, 448-453.  
18227064 G.K.Seabold, P.Y.Wang, K.Chang, C.Y.Wang, Y.X.Wang, R.S.Petralia, and R.J.Wenthold (2008).
The SALM family of adhesion-like molecules forms heteromeric and homomeric complexes.
  J Biol Chem, 283, 8395-8405.  
19077264 H.Park, J.Huxley-Jones, R.P.Boot-Handford, P.N.Bishop, T.K.Attwood, and J.Bella (2008).
LRRCE: a leucine-rich repeat cysteine capping motif unique to the chordate lineage.
  BMC Genomics, 9, 599.  
18264786 J.A.Legg, J.M.Herbert, P.Clissold, and R.Bicknell (2008).
Slits and Roundabouts in cancer, tumour angiogenesis and endothelial cell migration.
  Angiogenesis, 11, 13-21.  
18359766 N.Fukuhara, J.A.Howitt, S.A.Hussain, and E.Hohenester (2008).
Structural and functional analysis of slit and heparin binding to immunoglobulin-like domains 1 and 2 of Drosophila Robo.
  J Biol Chem, 283, 16226-16234.
PDB codes: 2vr9 2vra
18980679 S.Kaur, G.V.Samant, K.Pramanik, P.W.Loscombe, M.L.Pendrak, D.D.Roberts, and R.Ramchandran (2008).
Silencing of directional migration in roundabout4 knockdown endothelial cells.
  BMC Cell Biol, 9, 61.  
17901255 A.Schmandke, A.Schmandke, and S.M.Strittmatter (2007).
ROCK and Rho: biochemistry and neuronal functions of Rho-associated protein kinases.
  Neuroscientist, 13, 454-469.  
17848514 C.Morlot, N.M.Thielens, R.B.Ravelli, W.Hemrika, R.A.Romijn, P.Gros, S.Cusack, and A.A.McCarthy (2007).
Structural insights into the Slit-Robo complex.
  Proc Natl Acad Sci U S A, 104, 14923-14928.
PDB codes: 2v9q 2v9r 2v9s 2v9t
17704564 C.Morlot, W.Hemrika, R.A.Romijn, P.Gros, S.Cusack, and A.A.McCarthy (2007).
Production of Slit2 LRR domains in mammalian cells for structural studies and the structure of human Slit2 domain 3.
  Acta Crystallogr D Biol Crystallogr, 63, 961-968.
PDB code: 2v70
17192264 H.M.Kim, S.C.Oh, K.J.Lim, J.Kasamatsu, J.Y.Heo, B.S.Park, H.Lee, O.J.Yoo, M.Kasahara, and J.O.Lee (2007).
Structural diversity of the hagfish variable lymphocyte receptors.
  J Biol Chem, 282, 6726-6732.
PDB codes: 2o6q 2o6r 2o6s
17517123 N.Matsushima, T.Tanaka, P.Enkhbayar, T.Mikami, M.Taga, K.Yamada, and Y.Kuroki (2007).
Comparative sequence analysis of leucine-rich repeats (LRRs) within vertebrate toll-like receptors.
  BMC Genomics, 8, 124.  
17567741 T.Huyton, and C.Wolberger (2007).
The crystal structure of the tumor suppressor protein pp32 (Anp32a): structural insights into Anp32 family of proteins.
  Protein Sci, 16, 1308-1315.
PDB codes: 2je0 2je1
17553100 W.D.Andrews, M.Barber, and J.G.Parnavelas (2007).
Slit-Robo interactions during cortical development.
  J Anat, 211, 188-198.  
17029581 B.J.Dickson, and G.F.Gilestro (2006).
Regulation of commissural axon pathfinding by slit and its Robo receptors.
  Annu Rev Cell Dev Biol, 22, 651-675.  
16341215 J.R.Bermingham, H.Shearin, J.Pennington, J.O'Moore, M.Jaegle, S.Driegen, A.van Zon, A.Darbas, E.Ozkaynak, E.J.Ryu, J.Milbrandt, and D.Meijer (2006).
The claw paw mutation reveals a role for Lgi4 in peripheral nerve development.
  Nat Neurosci, 9, 76-84.  
17001097 L.Banci, I.Bertini, S.Cusack, Jong, U.Heinemann, E.Y.Jones, F.Kozielski, K.Maskos, A.Messerschmidt, R.Owens, A.Perrakis, A.Poterszman, G.Schneider, C.Siebold, I.Silman, T.Sixma, G.Stewart-Jones, J.L.Sussman, J.C.Thierry, and D.Moras (2006).
First steps towards effective methods in exploiting high-throughput technologies for the determination of human protein structures of high biomedical value.
  Acta Crystallogr D Biol Crystallogr, 62, 1208-1217.  
17062560 S.A.Hussain, M.Piper, N.Fukuhara, L.Strochlic, G.Cho, J.A.Howitt, Y.Ahmed, A.K.Powell, J.E.Turnbull, C.E.Holt, and E.Hohenester (2006).
A molecular mechanism for the heparan sulfate dependence of slit-robo signaling.
  J Biol Chem, 281, 39693-39698.  
16481322 S.Kaur, M.D.Castellone, V.M.Bedell, M.Konar, J.S.Gutkind, and R.Ramchandran (2006).
Robo4 signaling in endothelial cells implies attraction guidance mechanisms.
  J Biol Chem, 281, 11347-11356.  
16015381 A.Chédotal, G.Kerjan, and C.Moreau-Fauvarque (2005).
The brain within the tumor: new roles for axon guidance molecules in cancers.
  Cell Death Differ, 12, 1044-1056.  
15979925 M.Klagsbrun, and A.Eichmann (2005).
A role for axon guidance receptors and ligands in blood vessel development and tumor angiogenesis.
  Cytokine Growth Factor Rev, 16, 535-548.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.