TTL Proteins Scaffold Brassinosteroid Signaling Components at the Plasma Membrane to Optimize Signal Transduction in Arabidopsis

Abstract

Brassinosteroids (BRs) form a group of steroidal hormones essential for plant growth, development, and stress responses. BRs are perceived extracellularly by plasma membrane receptor-like kinases that activate an interconnected signal transduction cascade, leading to the transcriptional regulation of BR-responsive genes. TETRATRICOPEPTIDE THIOREDOXIN-LIKE (TTL) genes are specific for land plants, and their encoded proteins are defined by the presence of protein-protein interaction motives, that is, an intrinsic disordered region at the N terminus, six tetratricopeptide repeat domains, and a C terminus with homology to thioredoxins. TTL proteins thus likely mediate the assembly of multiprotein complexes. Phenotypic, molecular, and genetic analyses show that TTL proteins are positive regulators of BR signaling in Arabidopsis (Arabidopsis thaliana). TTL3 directly interacts with a constitutively active BRASSINOSTEROID INSENSITIVE1 (BRI1) receptor kinase, BRI1-SUPPRESSOR1 phosphatase, and the BRASSINAZOLE RESISTANT1 transcription factor and associates with BR-SIGNALING KINASE1, BRASSINOSTEROID INSENSITIVE2 kinases, but not with BRI1-ASSOCIATED KINASE1. A functional TTL3-green fluorescent protein (GFP) shows dual cytoplasmic plasma membrane localization. Depleting the endogenous BR content reduces plasma membrane localization of TTL3-GFP, while increasing BR content causes its plasma membrane relocalization, where it strengthens the association of BR signaling components. Our results reveal that TTL proteins promote BR responses and suggest that TTL proteins may function as scaffold proteins by bringing together cytoplasmic and plasma membrane BR signaling components.

Brassinosteroids: Multidimensional Regulators of Plant Growth, Development, and Stress Responses

“…In line with the notion that scaffolding plays an important role in BR signaling, TETRATRICOPEPTIDE THIOREDOXIN-LIKE (TTL) proteins TTL1, TTL3, and TTL4 have recently been implicated in bringing BR signaling components together at the PM (Amorim-Silva et al, 2019). TTL3 forms a complex with several players in BR signaling, including BRI1, BSU1, and BZR1.…”

“…Together with its homologs TTL3/4, act to scaffold BR signaling components at the PM (Amorim-Silva et al, 2019) Modulators of BES1/BZR1 degradation and activation MAX2 At2g42620 At5g40140 At2g32950 At3g58040…”

Deviating from the Beaten Track: New Twists in Brassinosteroid Receptor Function

“…In line with this, a recent study identified TETRATRICOPEPTIDE-REPEAT THIOREDOXIN-LIKE proteins (TTLs), which combine several protein interaction motifs, i.e. have the potential for multivalency (engaging in several simultaneous protein-protein interactions), as positive regulators of BR signalling by scaffolding BR signalling components [135]. TTL proteins become membrane-associated when BR signalling is activated and interact with BRI1, BSU1, BSK1, BIN2, and the BZR1 transcription factor, tethering many of the required signalling components in multiprotein complexes [135].…”

“…have the potential for multivalency (engaging in several simultaneous protein-protein interactions), as positive regulators of BR signalling by scaffolding BR signalling components [135]. TTL proteins become membrane-associated when BR signalling is activated and interact with BRI1, BSU1, BSK1, BIN2, and the BZR1 transcription factor, tethering many of the required signalling components in multiprotein complexes [135]. Thus, rather than as a series of isolated reactions, BR signalling might be better thought of as a signalling assembly, which could serve as a mechanism ensuring signalling specificity in light of the diverse roles that several of the signalling components engage in.…”

Molecular Mechanisms of Brassinosteroid-Mediated Responses to Changing Environments in Arabidopsis

“…At the receptor level, the function of BRI1 can be post-translationally regulated by PUB12/13-directed ubiquitination, PP2A-mediated dephosphorylation, BKI1-mediated kinase inhibition, and BIK1-and BIR3-mediated competition for the coreceptor BAK1 [69][70][71][72][73]. At the BIN2 level, it has been reported that BIN2 is regulated by OCTOPUSor POLAR-mediated membrane sequestration, HDA6-mediated deacetylation, KIB1-mediated ubiquitination, TTL-enhanced interaction with BSU1, ABI1/2-mediated dephosphorylation, and ROS (reactive oxygen species)-mediated oxidation [74][75][76][77][78][79][80]. At the transcription level, it has been reported that PP2A phosphatases can promote BR signaling by dephosphorylating BES1 and BZR1, whereas 14-3-3 and BRZ-SENSITIVE-SHORT HYPOCOTYL1 (BSS1) negatively regulate BR signaling by inhibiting the translocation of BES1 and BZR1 from the cytosol to the nucleus [81][82][83].…”