Dissecting brassinosteroid signaling mediated regulation of root growth angle in wheat

RGA of different root-types (primary, seminal and crown) are often distinct to limit competition and are referred to as gravitropic setpoint angle (GSA). The vertical GSA of primary roots is maintained by the positive gravitropic mechanism, while non-vertical GSA in other root-types is determined by competing gravitropic and anti-gravitropic offset (AGO) mechanisms. Although past research in the model plant Arabidopsis unraveled gravitropic mechanisms, understanding of AGO mechanisms remains limited as crucial root-types (seminal and crown) do not exist in Arabidopsis. Moreover, these mechanisms remain largely unexplored in crops due to limited genetic resources.
Wheat, an important crop for the UK and world, represents an ideal system for studying RGA as all root-types are easily distinguiishable and have available genetic and genomic resources. Recently, we screened various wheat phytohormone signaling mutants (auxin, ethylene, brassinosteroid and bibberellin) for RGA and observed that BR signaling mutants (prominently for tabri1) show significantly steeper seminal and crown roots compared to wildtype.
Aim:
In this project, we will determine the molecular mechanisms by which brassinosteroids control RGA in wheat.
Work Plan:
Do other Brassinosteroid pathway genes regulate RGA? (Year1)
tabri1 (A-homeologue) showed >30% steeper seminal and crown RGA than the wildtype. We will phenotype remaining single tabri1 mutants (of B- and D-homeologues) and their combinatorial double and triple mutants and assess if the observed tabri1 pheotype is specific to A-homeolgoue or enhanced by stacking mutations in other TaBRI1 homeologues. Additionally, we will screen other available brassinosteroid signaling, biosynthetic and receptor mutants for RGA phenotype using vertical and 90 gravistmulus.
Where does TaBR1-dependant RGA regulation takes place? (Year1-2)
We will investigate where TaBRI1 is made in the wheat root and functions to control RGA by tagging the BRI1 protein with green fluorescent protein (GFP), then putting this transgene into tabri1 plants to test if it can rescue the mutant and reveal its location of expression.
Explore the genetic diversity of BR signaling and RGA in Watkins collection (Year3)
We are currently screening >800 lines from a wheat population for RGA phenotype. Using bioinformatics pipeline, we will investigate how much natural variation of RGA is explained by the polymorphism within the TaBR1. This analyss will help link the ancestory, habitat and environmental conditions from which these lines were collected and generate further testable hypothesis.
What downstream components TaBR1 regulate to control RGA? (Year3-4)
Extract RNA from root samples of WT and single or higher order Tabri1 mutants growing under vertical and gravistimulus conditions and perform RNAseq analysis to determine key differentially expressed genes and their enriched processes involved during BRI1 mediated RGA control.
What is the relevane of TaBR1 mediated RGA? (Year4)
Nutrient, water and heat stresses are known to influence RGA to optimise capture of soil resources. We will perform growth-room, glasshouse and field experiments on mutants and WT under control and stress coditions and assess the RGA, root and shoot biomass and grain yield.
Output:
Improved understanding of molecular mechanisms controlling RGA will guide selection of alleles that can be exploited through DFW breeders toolkit.