Elongating roots for efficient nitrogen foraging via altered brassinosteroid and auxin synthesis and signaling

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Plants adapt continuously to spatiotemporal nutrient fluctuations in soils by sensing available nutrient forms and modulating their root system architecture to sustain adequate nutrition. Such dynamic architectural responses of roots allow plants to optimize spatially defined soil exploration for limiting nutrients. When plants grow under suboptimal external nitrogen (N) levels that induce mild deficiency, they increase the elongation of primary and lateral roots. This stimulation of root expansion under mild N deficiency is of particular interest as it increases the soil volume that can be explored by roots for nutrient acquisition. In our previous work, we found in natural accessions of the model species Arabidopsis allelic variation in brassinosteroid and auxin biosynthesis or signalling genes (YUC8 and BSK3), which alters root elongation under mild N deficiency. The ultimate goal of the 6-years project is to understand the hormonal regulation of the root foraging response to low N in plants and to exploit this knowledge for improving soil exploration – and ultimately N uptake efficiency – in barley. The two major objectives of the first 3 years are i) to determine in Arabidopsis the role of YUC8-dependent auxin biosynthesis and its relation to brassinosteroids in the root foraging response to mild N deficiency, and ii) to explore in a translational approach in barley the contribution of allelic variation in brassinosteroid as well as auxin biosynthesis and signaling genes to the root foraging response to mild N deficiency. First, we will verify in Arabidopsis the molecular mechanism behind the identified allelic variation in the YUC8 gene and its role in regulating the root foraging response. We will determine the lateral root elongation response in single and multiple yuc mutant lines and in lines complemented with different YUC8 alleles. Using these and other mutant and reporter lines, we will investigate the crosstalk between brassinosteroids and auxin in the root elongation response to low N. In a translational approach, we will introduce the "strong" BSK3 allele from Arabidopsis into barley, which carries in all accession investigated so far only a "weak" BSK3 allele. For this purpose, we will combine Cas endonuclease-mediated knockout of the weak BSK3 endogene with complementation by the strong transgene in just one step. Finally, we will create an inventory of N deficiency-regulated genes in barley roots by an RNA sequencing approach, which will serve as resource for the selection of further barley genes that will be targeted by CRISPR-Cas-mediated gene deletion and by ectopic expression using a root-specific promoter. All lines will be subjected to phenotypic analysis of root architectural responses to mild N deficiency.