title: Interaction with both ZNRF3 and LGR4 is required for the signalling activity of R-spondin.
abstract: R-spondin proteins sensitize cells to Wnt signalling and act as potent stem cell growth factors. Various membrane proteins have been proposed as potential receptors of R-spondin, including LGR4/5, membrane E3 ubiquitin ligases ZNRF3/RNF43 and several others proteins. Here, we show that R-spondin interacts with ZNRF3/RNF43 and LGR4 through distinct motifs. Both LGR4 and ZNRF3 binding motifs are required for R-spondin-induced LGR4/ZNRF3 interaction, membrane clearance of ZNRF3 and activation of Wnt signalling. Importantly, Wnt-inhibitory activity of ZNRF3, but not of a ZNRF3 mutant with reduced affinity to R-spondin, can be strongly suppressed by R-spondin, suggesting that R-spondin primarily functions by binding and inhibiting ZNRF3. Together, our results support a dual receptor model of R-spondin action, where LGR4/5 serve as the engagement receptor whereas ZNRF3/RNF43 function as the effector receptor.
authors.affiliation: Novartis Institutes for Biomedical Research, 250 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xie
authors.forename: Raffaella
authors.initials: R
authors.lastname: Zamponi
authors.forename: Olga
authors.initials: O
authors.lastname: Charlat
authors.forename: Melissa
authors.initials: M
authors.lastname: Ramones
authors.forename: Susanne
authors.initials: S
authors.lastname: Swalley
authors.forename: Xiaomo
authors.initials: X
authors.lastname: Jiang
authors.forename: Daniel
authors.initials: D
authors.lastname: Rivera
authors.forename: William
authors.initials: W
authors.lastname: Tschantz
authors.forename: Bo
authors.initials: B
authors.lastname: Lu
authors.forename: Lisa
authors.initials: L
authors.lastname: Quinn
authors.forename: Chris
authors.initials: C
authors.lastname: Dimitri
authors.forename: Jefferson
authors.initials: J
authors.lastname: Parker
authors.forename: Doug
authors.initials: D
authors.lastname: Jeffery
authors.forename: Sheri K
authors.initials: SK
authors.lastname: Wilcox
authors.forename: Mike
authors.initials: M
authors.lastname: Watrobka
authors.forename: Peter
authors.initials: P
authors.lastname: LeMotte
authors.forename: Brian
authors.initials: B
authors.lastname: Granda
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Porter
authors.forename: Vic E
authors.initials: VE
authors.lastname: Myer
authors.forename: Andreas
authors.initials: A
authors.lastname: Loew
authors.forename: Feng
authors.initials: F
authors.lastname: Cong
mesh_terms: D020816:Amino Acid Motifs; D001665:Binding Sites; D057809:HEK293 Cells; D006801:Humans; D011485:Protein Binding; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D019699:Thrombospondins / Q000737:chemistry / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D060449:Wnt Signaling Pathway; D000098907:R-Spondins
chemical_list: D043562:Receptors, G-Protein-Coupled; D019699:Thrombospondins; D044767:Ubiquitin-Protein Ligases; C411562:LGR4 protein, human; C503243:RSPO1 protein, human; D000098907:R-Spondins; C572950:ZNRF3 protein, human