Deactivation of the Arabidopsis BRASSINOSTEROID INSENSITIVE 1 (BRI1) receptor kinase by autophosphorylation within the glycine-rich loop

Abstract

The activity of the dual-specificity receptor kinase, brassinosteroid insensitive 1 (BRI1), reflects the balance between phosphorylationdependent activation and several potential mechanisms for deactivation of the receptor. In the present report, we elucidate a unique mechanism for deactivation that involves autophosphorylation of serine-891 in the ATP-binding domain. Serine-891 was identified previously as a potential site of autophosphorylation by mass spectrometry, and sequence-specific antibodies and mutagenesis studies now unambiguously establish phosphorylation of this residue. In vivo, phosphorylation of serine-891 increased slowly with time following application of brassinolide (BL) to Arabidopsis seedlings, whereas phosphorylation of threonine residues increased rapidly and then remained constant. Transgenic plants expressing the BRI1(S891A)-Flag-directed mutant have increased hypocotyl and petiole lengths, relative to wild-type BRI1-Flag (both in the bri1-5 background), and accumulate higher levels of the unphosphorylated form of the BES1 transcription factor in response to exogenous BL. In contrast, plants expressing the phosphomimetic S891D-directed mutant are severely dwarfed and do not accumulate unphosphorylated BES1 in response to BL. Collectively, these results suggest that autophosphorylation of serine-891 is one of the deactivation mechanisms that inhibit BRI1 activity and BR signaling in vivo. Many arginine-aspartate (RD)-type leucine-rich repeat receptor-like kinases have a phosphorylatable residue within the ATP-binding domain, suggesting that this mechanism may play a broad role in receptor kinase deactivation.T he receptor-like kinase (RLK) family in Arabidopsis contains more than 600 members, of which more than 400 are structurally and functionally similar to animal receptor kinases but are evolutionarily distinct (1). Animal receptor kinases are predominantly tyrosine kinases, whereas plant receptor kinases are generally classified as Ser/Thr kinases, although recent work suggests that some plant receptor kinases are dual-specificity kinases that can also autophosphorylate on tyrosine residues (2-5). One of the best-studied plant receptor kinases is BRASSINOSTEROID INSENSITIVE 1 (BRI1), which functions with its coreceptor, BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1), in brassinosteroid (BR) signaling (6-11). Current thinking is that BRI1 and BAK1 are in their unphosphorylated forms and inactive in the absence of BR, whereas in the presence of the BR ligand, BRI1 and BAK1 heterodimerize and become activated via auto-and transphosphorylation (12). The BRI1 KINASE INHIBITOR 1 (BKI1) and the BR-signaling kinase 1 (BSK1) may be two of the immediate downstream components that are first phosphorylated by BRI1 (13,14). BSK then activates the BRI1 SUPPRESSOR 1 (BSU1) phosphatase (15), which in turn inhibits the glycogen synthase 3-like protein kinase, BRASSINOSTEROID INSENSITIVE 2 (BIN2), by dephosphorylation of an essential phosphotyrosine residue (16). The net result is that the transcription fac...

Two SERK Receptor-Like Kinases Interact with EMS1 to Control Anther Cell Fate Determination

“…Conversely, The T872A mutation in the BRI1 juxtamembrane domain increases BRI1 kinase activity (Wang et al, 2005). BRI1 S981A (in the ATP-binding domain), which has a normal kinase activity, functions as the wild-type BRI1 (Wang et al, 2005;Oh et al, 2012). However, the phosphorylationmimic mutation in Ser-891 deactivates BRI1 phosphorylation and impairs BRI1 biological function (Oh et al, 2012).…”

“…BRI1 S981A (in the ATP-binding domain), which has a normal kinase activity, functions as the wild-type BRI1 (Wang et al, 2005;Oh et al, 2012). However, the phosphorylationmimic mutation in Ser-891 deactivates BRI1 phosphorylation and impairs BRI1 biological function (Oh et al, 2012). The S938A mutation in FLS2 abolishes immune responses (Cao et al, 2013).…”

A Mathematical Model for the Coreceptors SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE1 and SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE3 in BRASSINOSTEROID INSENSITIVE1-Mediated Signaling

“…The protein phosphatase 2A stimulates BRI1 signaling by dephosphorylating the downstream BRI1 transcriptional regulator BRASSINAZOLE-RESISTANT1 but is also required for the degradation of BRI1 via dephosphorylation of the activated receptor (Di Rubbo et al, 2011;Tang et al, 2011;Wu et al, 2011). Attenuation of the BRI1 signal is regulated by the phosphorylation of specific Ser and Thr residues (Oh et al, 2012). Thus, there appears to be evidence for a role of the SERKs as amplifiers of the entire signaling pathway, rather than having a specific effect on only one element.…”

Meta-Analysis of Arabidopsis thaliana Phospho-Proteomics Data Reveals Compartmentalization of Phosphorylation Motifs

“…Many protein kinases are themselves regulated by phosphorylation, either through autophosphorylation or by upstream kinases (Bögre et al, 2003;Park et al, 2011;Oh et al, 2012bOh et al, , 2012c. Because these kinases are central in phosphorylation networks, we evaluated all 8141 identified p-proteins for the presence of a predicted PFAM domain for pKinase or pKinase_Tyr using a threshold of E<0.01 (Supplemental Data Set 3).…”