THE ROLE OF LRR RECEPTOR-LIKE KINASES IN ABA DEPENDENT TPK1 CHANNEL REGULATION

The increasing world population demands increased food production, but several abiotic factors, for example, drought is hampering increased food production. The problem of drought is increasing and further limiting the productivity of many crops. There could be a range of potential strategies that may help plants to tolerate and avoid drought conditions, for example, to produce stress tolerant plants, to understand and utilize signalling mechanisms in plants to adapt them to drought, to minimize yield loss in dryland areas and to reduce the water needs.
ABA (abscisic acid) is produced during drought stress causing the closure of the stomatal pore to prevent water loss. K+ (potassium) release from the vacuole through K+ transporters localised to the tonoplast, during these conditions this is one of the important steps. However, the mechanism of coupling of ABA to the tonoplast is not known yet. The AtTPK1 channel (Arabidopsis thaliana two pore potassium channel) is localised to the tonoplast and has been shown to have a role in the vacuolar K+ release and stomatal closure. AtTPK1 is activated by phosphorylation and binding of 14-3-3 proteins. The surface of the plasma membrane of plants contains receptor-like kinases (RLKs) that are known to be involved in the early steps of osmotic-stress signalling. Binding of a ligand to the extracellular domain of the RLK activates the intracellular kinase domain, resulting in bringing extracellular environment signals into the intracellular targets. LRRs (Leucine-rich repeat) are a possible mechanism to link external ABA with TPK1 and because LRRs had been found to impact in TPK1 current, for example, two LRR receptor kinase candidates, KINASE1 and KINASE2 (At3g02880 and At4g21410) were shown by patch clamp studies to affect TPK1 current stimulation. The BiFC (Bimolecular fluorescence complementation) studies also showed interaction of these kinases with TPK1 when they were treated with ABA. Therefore, it was hypothesized that they are involved in the activation of TPK1 (Isner et al., unpublished data). The kinase mutant lines were selected for further characterization in comparison to tpk1 and WT (wild type) in different media conditions.
All the knockout lines showed shorter root lengths and lower fresh weights as compared to the wild type in K+-deficient, higher K+ and osmotic stress conditions. Lower fresh weights for the KO (knock out) lines as compared to the wild type were also observed in soil in control and moderate stress conditions. The lower growth of the tpk1 and kinase KO lines as compared to the wild type may be because of the lack of TPK1 activity. The lack of kinase proteins may lead to the inactivation of TPK1 channels and thus it leads to the comparable results between the kinase KO lines and tpk1 KO line. These results suggest a link between these kinases and TPK1 channel activity. These lines were also tested for the stomatal conductance under various ABA treatments applied to the excised leaves. The kinase KO lines and tpk1 KO led to a delayed response in stomatal closure after exposure to different concentration of ABA (1 µM, 10 µM and 100 µM).
The similarities in phenotype between the kinases KO and channel KO mutants suggest there may be a relation between these kinases and the TPK1 channel. Combined with other, as yet unpublished data, the data from this report support the idea of the involvement of these kinases in ABA dependent regulation of TPK1.