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Quantitative Profiling of WNT-3A Binding to All Human Frizzled Paralogues in HEK293 Cells by NanoBiT/BRET Assessments.
Kozielewicz, Pawel; Shekhani, Rawan; Moser, Stefanie; Bowin, Carl-Fredrik; Wesslowski, Janine; Davidson, Gary; Schulte, Gunnar.
Afiliação
  • Kozielewicz P; Section of Receptor Biology & Signaling, Dept. Physiology & Pharmacology, Karolinska Institutet, S-17165, Stockholm, Sweden.
  • Shekhani R; Section of Receptor Biology & Signaling, Dept. Physiology & Pharmacology, Karolinska Institutet, S-17165, Stockholm, Sweden.
  • Moser S; Institute of Biological and Chemical Systems-Functional Molecular Systems (IBCS-FMS), Karlsruhe Institute of Technology (KIT), 76131, Karlsruhe, Germany.
  • Bowin CF; Section of Receptor Biology & Signaling, Dept. Physiology & Pharmacology, Karolinska Institutet, S-17165, Stockholm, Sweden.
  • Wesslowski J; Institute of Biological and Chemical Systems-Functional Molecular Systems (IBCS-FMS), Karlsruhe Institute of Technology (KIT), 76131, Karlsruhe, Germany.
  • Davidson G; Institute of Biological and Chemical Systems-Functional Molecular Systems (IBCS-FMS), Karlsruhe Institute of Technology (KIT), 76131, Karlsruhe, Germany.
  • Schulte G; Section of Receptor Biology & Signaling, Dept. Physiology & Pharmacology, Karolinska Institutet, S-17165, Stockholm, Sweden.
ACS Pharmacol Transl Sci ; 4(3): 1235-1245, 2021 Jun 11.
Article em En | MEDLINE | ID: mdl-34151213
ABSTRACT
The WNT signaling system governs critical processes during embryonic development and tissue homeostasis, and its dysfunction can lead to cancer. Details concerning selectivity and differences in relative binding affinities of 19 mammalian WNTs to the cysteine-rich domain (CRD) of their receptors-the ten mammalian Frizzleds (FZDs)-remain unclear. Here, we used eGFP-tagged mouse WNT-3A for a systematic analysis of WNT interaction with every human FZD paralogue in HEK293A cells. Employing HiBiT-tagged full-length FZDs, we studied eGFP-WNT-3A binding kinetics, saturation binding, and competition binding with commercially available WNTs in live HEK293A cells using a NanoBiT/BRET-based assay. Further, we generated receptor chimeras to dissect the contribution of the transmembrane core to WNT-CRD binding. Our data pinpoint distinct WNT-FZD selectivity and shed light on the complex WNT-FZD binding mechanism. The methodological development described herein reveals yet unappreciated details of the complexity of WNT signaling and WNT-FZD interactions, providing further details with respect to WNT-FZD selectivity.

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Idioma: En Revista: ACS Pharmacol Transl Sci Ano de publicação: 2021 Tipo de documento: Article País de afiliação: Suécia

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Idioma: En Revista: ACS Pharmacol Transl Sci Ano de publicação: 2021 Tipo de documento: Article País de afiliação: Suécia