Sequential Transphosphorylation of the BRI1/BAK1 Receptor Kinase Complex Impacts Early Events in Brassinosteroid Signaling

Abstract

Brassinosteroids (BRs) regulate plant development through a signal transduction pathway involving the BRI1 and BAK1 transmembrane receptor kinases. The detailed molecular mechanisms of phosphorylation, kinase activation, and oligomerization of the BRI1/BAK1 complex in response to BRs are uncertain. We demonstrate that BR-dependent activation of BRI1 precedes association with BAK1 in planta, and that BRI1 positively regulates BAK1 phosphorylation levels in vivo. BRI1 transphosphorylates BAK1 in vitro on specific kinase-domain residues critical for BAK1 function. BAK1 also transphosphorylates BRI1, thereby quantitatively increasing BRI1 kinase activity toward a specific substrate. We propose a sequential transphosphorylation model in which BRI1 controls signaling specificity by direct BR binding followed by substrate phosphorylation. The coreceptor BAK1 is then activated by BRI1-dependent transphosphorylation and subsequently enhances signaling output through reciprocal BRI1 transphosphorylation. This model suggests both conservation and distinct differences between the molecular mechanisms regulating phosphorylation-dependent kinase activation in plant and animal receptor kinases.

Functional importance of BAK1 tyrosine phosphorylation in vivo

“…How the phosphorylation status of Tyr-610 affects the functions of BAK1 in various signaling pathways is not clear but there are several possibilities to consider. The first is that phosphorylation of Tyr-610 is simply required for proper interaction with certain associated receptor kinase such that efficient transphosphorylation can occur, 2 and this altered functional interaction in the Y610F directed mutant with BRI1 may explain the reduced BL-signaling observed in vivo. 3 A second possibility is that phosphorylation of Tyr-610 may be directly recognized by phosphotyrosinebinding proteins such as SH2-domain containing proteins 38 and thereby directly contribute to protein:protein interactions in the signaling complex.…”

“…29,30 In its role as coreceptor, BAK1 is thought to bind to the receptor kinase in a ligand-dependent manner, and to then autophosphorylate and also transphosphorylate sites on the receptor kinase. 2 How the various functions and interactions of BAK1 are regulated is not known but conceivably site-specific (auto) phosphorylation could play a role. In the present report, we discuss the occurrence of Tyr autophosphorylation of BAK1 and present a simplified model to describe different mechanisms that may be involved in Tyr signaling by this multifunctional receptor kinase.…”

“…That phosphorylation of BAK1 at the Tyr-610 sites plays a positive role in BR which is also known as BAK1, can multitask 24 as coreceptor with BRI1 in BR signaling, 2,25 BIR1 in control of cell death, 26 FLS2 in flg22 signaling, 27 EFR in elf18 signaling, 28 and PEPR1 and PEPR2 in Pep1 signaling. 29,30 In its role as coreceptor, BAK1 is thought to bind to the receptor kinase in a ligand-dependent manner, and to then autophosphorylate and also transphosphorylate sites on the receptor kinase.…”

“…3 Finally, the ability of the BAK1(Y610F) directed mutant to transphosphorylate the kinase inactive mBRI1 was severely impaired, 3 which provides a possible explanation for reduced BR signaling in vivo as reciprocal transphosphorylations have been shown to be essential for enhanced BR signaling. 2 Our current working model for the role of Tyr-610 phosphorylation in BR signaling is summarized in Figure 3. It will be interesting to determine whether other transphosphorylation reactions catalyzed by BAK1 are impacted by the Y610F directed mutation.…”

Mining expressed sequence tags of rapeseed (Brassica napus L.) to predict the drought responsive regulatory network

“…BR signaling is induced by binding of BR to a membrane receptor kinase, BRI1. BRI1 makes hetero oligomers with another receptor kinase, BAK1 to shape a phosphorylation string that ultimately causes functional changes of target genes (Nam and Li 2002;Wang et al 2008). The BRI1 receptor complex induces the redundant genes encoding basic helix-loop-helix (bHLH) namely BEE1, BEE2 and BEE3 (Friedrichsen et al 2002).…”