TY - UNPB
T1 - NRBP1 pseudokinase binds to and activates the WNK pathway in response to osmotic stress
AU - Amnekar, Ramchandra
AU - Dite, Toby Dite
AU - Lis, Paweł
AU - Bell, Sebastian
AU - Brown, Fiona
AU - Johnson, Clare
AU - Wilkinson, Stuart G
AU - Raggett, Samantha L.
AU - Dorward, Mark
AU - Wightman, Melanie
AU - Macartney, Thomas
AU - Filipe Soares, Renata
AU - Lamoliatte, Fred
AU - Alessi, Dario
N1 - The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.
PY - 2024/12/20
Y1 - 2024/12/20
N2 - WNK family kinases are regulated by osmotic stress and control ion homeostasis by activating SPAK and OXSR1 kinases. Using a proximity ligation approach, we found that osmotic stress promotes the association of WNK1 with the NRBP1 pseudokinase and TSC22D2/4 adaptor proteins, results that are confirmed by immunoprecipitation and mass spectrometry and immunoblotting studies. NRBP1 pseudokinase is closely related to WNK isoforms and contains a RΦ-motif binding conserved C-terminal (CCT) domain, similar to the CCT domains in WNKs, SPAK and OXSR1. Knockdown or knock-out of NRBP1 markedly inhibited sorbitol-induced activation of WNK1 and downstream components. We demonstrate recombinant NRBP1 can directly induce the activation of WNK4 in vitro. AlphaFold-3 modelling predicts that WNK1, SPAK, NRBP1, and TSC22D4 form a complex, in which two TSC22D4 RΦ-motifs interact with the CCTL1 domain of WNK1 and the CCT domain of NRBP1. Our data indicates NRBP1 functions as an upstream activator of the WNK pathway.
AB - WNK family kinases are regulated by osmotic stress and control ion homeostasis by activating SPAK and OXSR1 kinases. Using a proximity ligation approach, we found that osmotic stress promotes the association of WNK1 with the NRBP1 pseudokinase and TSC22D2/4 adaptor proteins, results that are confirmed by immunoprecipitation and mass spectrometry and immunoblotting studies. NRBP1 pseudokinase is closely related to WNK isoforms and contains a RΦ-motif binding conserved C-terminal (CCT) domain, similar to the CCT domains in WNKs, SPAK and OXSR1. Knockdown or knock-out of NRBP1 markedly inhibited sorbitol-induced activation of WNK1 and downstream components. We demonstrate recombinant NRBP1 can directly induce the activation of WNK4 in vitro. AlphaFold-3 modelling predicts that WNK1, SPAK, NRBP1, and TSC22D4 form a complex, in which two TSC22D4 RΦ-motifs interact with the CCTL1 domain of WNK1 and the CCT domain of NRBP1. Our data indicates NRBP1 functions as an upstream activator of the WNK pathway.
U2 - 10.1101/2024.12.12.628181
DO - 10.1101/2024.12.12.628181
M3 - Preprint
BT - NRBP1 pseudokinase binds to and activates the WNK pathway in response to osmotic stress
PB - BioRxiv
ER -