The mechanism of signal reception for the plant hormone strigolactone

Abstract

Strigolactones (SLs) are a class of hormones that play an important role in the regulation of many plant developmental processes, especially branching. In petunia, SL signalling is mediated by the receptor, DAD2, which functions as an enzyme-receptor that hydrolyses SL and binds to its hydrolysis product. DAD2 undergoes a conformational change and interacts with its signal transduction partner PhMAX2A and target protein PhD53A to activate a signalling cascade. The goal of this research was to dissect the roles of catalysis, conformational changes of DAD2 and interaction with PhMAX2A and PhD53A in the activation of SL signal transduction. Random and site-directed mutagenesis approaches were used to address whether the catalytic activity of DAD2 could be uncoupled from its ability to interact with PhMAX2A. DAD2 mutants were characterised for enzyme activity, thermal stability and interactions with PhMAX2A and PhD53A. Among all of the mutants tested, the N242I mutant, having a mutation affecting a residue residing on the surface of the core α/β hydrolase fold, showed increased interaction with PhMAX2A in the absence of SL, suggesting that catalysis can be separated from the DAD2-PhMAX2A interaction. Despite having enzymatic activity similar to wild-type DAD2, the N242I mutant was thermally less stable than wild-type DAD2, suggesting that the mutation may have altered the stability/flexibility of the protein and thus allowing this mutant to interact with PhMAX2A even without SL hydrolysis. However, the N242I mutant still required SL to interact with PhD53A, indicating that the change in stability/flexibility of the protein caused by the N242I mutation was insufficient to facilitate the interaction with PhD53A. A transgenesis approach was aimed at testing the biological functions of selected DAD2 mutants in Arabidopsis d14 mutants. However, the lack of complementation by wild-type DAD2 due to no detectable levels of DAD2 protein in the Arabidopsis transgenic lines prevented further in plant a investigation of the DAD2mutants. Overall, the in vitro findings support the hypothesis that conformational change in DAD2 is the crucial mechanism that activates SL signalling by allowing interaction with PhMAX2A, and catalysis could merely be a process that induces the conformational change.