title: Targeting the cis-dimerization of LINGO-1 with low MW compounds affects its downstream signalling.
abstract: OBJECTIVE
  The transmembrane protein LINGO-1 is a negative regulator in the nervous system mainly affecting axonal regeneration, neuronal survival, oligodendrocyte differentiation and myelination. However, the molecular mechanisms regulating its functions are poorly understood. In the present study, we investigated the formation and the role of LINGO-1 cis-dimers in the regulation of its biological activity.
  
  
  METHODS
  LINGO-1 homodimers were identified in both HEK293 and SH-SY5Y cells using co-immunoprecipitation experiments and BRET saturation analysis. We performed a hypothesis-driven screen for identification of small-molecule protein-protein interaction modulators of LINGO-1 using a BRET-based assay, adapted for screening. The compound identified was further assessed for effects on LINGO-1 downstream signalling pathways using Western blotting analysis and AlphaScreen technology.
  
  
  RESULTS
  LINGO-1 was present as homodimers in primary neuronal cultures. LINGO-1 interacted homotypically in cis-orientation and LINGO-1 cis-dimers were formed early during LINGO-1 biosynthesis. A BRET-based assay allowed us to identify phenoxybenzamine as the first conformational modulator of LINGO-1 dimers. In HEK-293 cells, phenoxybenzamine was a positive modulator of LINGO-1 function, increasing the LINGO-1-mediated inhibition of EGF receptor signalling and Erk phosphorylation.
  
  
  CONCLUSIONS
  Our data suggest that LINGO-1 forms constitutive cis-dimers at the plasma membrane and that low MW compounds affecting the conformational state of these dimers can regulate LINGO-1 downstream signalling pathways. We propose that targeting the LINGO-1 dimerization interface opens a new pharmacological approach to the modulation of its function and provides a new strategy for drug discovery.
authors.affiliation: Centre de Biophysique Moléculaire, Département biologie cellulaire et cibles thérapeutiques, CNRS, UPR 4301, University of Orléans and INSERM, Orléans, France.
authors.forename: L
authors.initials: L
authors.lastname: Cobret
authors.forename: M L
authors.initials: ML
authors.lastname: De Tauzia
authors.forename: J
authors.initials: J
authors.lastname: Ferent
authors.forename: E
authors.initials: E
authors.lastname: Traiffort
authors.forename: I
authors.initials: I
authors.lastname: Hénaoui
authors.forename: F
authors.initials: F
authors.lastname: Godin
authors.forename: E
authors.initials: E
authors.lastname: Kellenberger
authors.forename: D
authors.initials: D
authors.lastname: Rognan
authors.forename: J
authors.initials: J
authors.lastname: Pantel
authors.forename: H
authors.initials: H
authors.lastname: Bénédetti
authors.forename: S
authors.initials: S
authors.lastname: Morisset-Lopez
mesh_terms: D045744:Cell Line, Tumor; D019281:Dimerization; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015394:Molecular Structure; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010643:Phenoxybenzamine / Q000737:chemistry / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects*; D013237:Stereoisomerism; D013329:Structure-Activity Relationship
chemical_list: C482293:LINGO1 protein, human; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010643:Phenoxybenzamine