Study of light dependent Arabidopsis phytochrome A signal transduction through FHY1 and its downstream gene expression regulation

PhyA (phytochrome A) is the primary photoreceptor responsible for continuous far-red light response, yet the mechanism of phyA signaling transduction remains largely unknown. FHY1 (FAR-RED ELONGATED HYPOCOTYL 1) defines a positive regulator in the phyA signaling pathway. It has been reported to be essential for light-regulated phyA nuclear accumulation through direct physical interaction with phyA, however, little is known about how FHY1 is involved in phyA signal transduction. Thus, the first goal of this dissertation research was to investigate how FHY1 transduces phyA signal under different light conditions. Western blot was used to show that FHY1 protein was phosphorylated rapidly after exposure to red light. Subsequent exposure to far-red light after the red light pulse reversed FHY1 phosphorylation. Such a phenomenon represented a classical red/far-red reversible low fluence response. The phosphorylation of FHY1 depended on functioning phyA but not on other phytochromes and cryptochromes. Furthermore, bimolecular fluorescence complementation (BiFC) and co-immunoprecipitation techniques were used to show that FHY1 directly interacts and interacts more stably with the Pr form of phyA in Arabidopsis seedlings. Finally, in vitro kinase assays confirmed that a recombinant phyA is able to robustly phosphorylate FHY1.;In the second part of this dissertation, far-red light regulation of subtilase gene expression mediated by phyA was studied. Arabidopsis thaliana has 57 subtilases, however the roles that these proteases play in the plants are largely unknown. Previous study revealed that more than 20 of the subtilase gene expressions were light dependent. It has been suggested that far-red light is one of the factors that controls expression of the subtilase genes. The goal of the second part of this dissertation was to investigate gene expression of the entire multigene family of subtilases through phyA-mediated far-red light response. Gene-expression-based bio-informatics approach was adopted to analyze the gene expressions of 57 Arabidopsis subtilase in wild type (WT), phyA and fhy1 under different light conditions, arriving at a functional classification of subtilases based on their expression behavior under white light and far-red light. The subtilases were classified into seven subgroups. The expression of At2g39850 and At5g59130, showed sole dependence on phyA under far-red light.