Investigation of the role of strigolactones in barley in response to drought

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Strigolactones (SLs) are described as a class of plant hormones that promotes symbiotic interactions in the rhizosphere and that regulates plant architecture and organ development. In barley it was recently shown that abscisic acid (ABA) represses transcripts of SL biosynthesis. On the other hand our preliminary data indicate that SL signalling mutant is hypersensitive to drought and insensitive to ABA treatment. Additional reports on interactions of SLs with ABA during drought stress in rice and tomato or osmotic stress in L japonicus suggest that SLs may play also an important role in coping with abiotic stress responses. The overall goal of the proposed project is to further dissect the ABA-SL interaction to reveal their role in the crop species barley in response to drought. In a first attempt the available barley mutant hvd14.d, defective in SL signaling, and its parental line will be grown on soil under ambient conditions and drought stress. Additional to physiological and morphological analyses immunolocalisation studies of HvD14 will be performed to increase our knowledge in regard of place of its transcription and/or possible transport of the protein via phloem or xylem during plant development and in response to drought. Complementary comparative studies with barley germplasm that display a natural variation in the sequence of the HvD14 gene will be analysed to gain knowledge about molecular mechanisms for enhanced tillering and drought tolerance. To reveal the direct or indirect interactions between SLs and ABA we will use targeted mutagenesis with CRISPR RNA/Cas9 technology and a TILLING population, to identify further mutants with modified barley genes of SL biosynthesis (D27, D17, D10, MAX1) and signaling (D3, D53, FC1). As in contrast to dicots, monocots contain several copies of MAX1 gene, the functionality of the three individual alleles of barley will be proven by specific knockout. Furthermore, differentially expressed genes and deep sequencing of transcriptome, completed with measurement of concentrations of SLs, ABA, auxins, brassinosteroids, cytokinins, gibberellins, jasmonic acid and salicylic acid will be analysed in drought-stressed wild-type and SL mutant plants to reveal the role of a possible crosstalk among the phytohormones. Moreover, selected mutant lines will be treated with selected hormones, such as ABA, brassinosteroids and cytokinins to confirm predicted interactions. Finally, results obtained in all work packages will be assembled to confirm an assumed crosstalk of SLs with other phytohormones in response to drought stress, with impact on stomata closure, and developmental processes as tillering or lateral root formation. In particular, analysis of phytohormone content and localization studies of D14 and other proteins that were identified within the studies as important factors in SL response to drought stress may further strengthen this hypothesis.