title: Expression, purification and functional characterization of Wnt signaling co-receptors LRP5 and LRP6.
abstract: Activation of the Wnt signaling cascade plays a pivotal role during development and in various disease states. Wnt signals are transduced by seven-transmembrane Frizzled (Fz) proteins and the single-transmembrane LDL-receptor-related proteins 5 or 6 (LRP5/6). Genetic mutations resulting in a loss or gain of function of LRP5 in humans lead to osteopenia and bone formation, respectively. These findings demonstrate the genetic link between LRP5 signaling and the regeneration of bone mass. Herein we describe for the first time the production and characterization of soluble ectodomains of LRP5 and LRP6, (EC-LRP5, EC-LRP6). We have produced these proteins in amounts that are compatible with both in vitro and cell-based assays to study their binding properties. Purified EC-LRP5 and EC-LRP6 were able to interact with Wnt signaling components Dkk1 and Dkk2 and their functionality was confirmed in cell-based Wnt signaling assays. Hence, tools are now available to explore LRP5/6 interaction with other proteins and to screen for synthetic or natural compounds and biologics that might be novel therapeutics targeting the Wnt pathway.
authors.affiliation: Protein Sciences, Galapagos, 102 route Gaston Roussel, F-93230 Romainville, France.
authors.forename: Damien
authors.initials: D
authors.lastname: Fleury
authors.forename: Béatrice
authors.initials: B
authors.lastname: Vayssiere
authors.forename: Robert
authors.initials: R
authors.lastname: Touitou
authors.forename: Corinne
authors.initials: C
authors.lastname: Gillard
authors.forename: Hélène
authors.initials: H
authors.lastname: Lebhar
authors.forename: Georges
authors.initials: G
authors.lastname: Rawadi
authors.forename: Patrick
authors.initials: P
authors.lastname: Mollat
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D036341:Intercellular Signaling Peptides and Proteins / Q000302:isolation & purification / Q000378:metabolism; D026502:LDL-Receptor Related Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D060492:Low Density Lipoprotein Receptor-Related Protein-5; D060488:Low Density Lipoprotein Receptor-Related Protein-6; D015398:Signal Transduction; D051153:Wnt Proteins / Q000378:metabolism*
chemical_list: C501127:Dkk1 protein, mouse; C405060:Dkk2 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D026502:LDL-Receptor Related Proteins; C555482:LRP5 protein, human; C555475:LRP6 protein, human; D060492:Low Density Lipoprotein Receptor-Related Protein-5; D060488:Low Density Lipoprotein Receptor-Related Protein-6; C555483:Lrp5 protein, mouse; C555476:Lrp6 protein, mouse; D051153:Wnt Proteins