UC San Diego

Plants monitor their light environment with a sophisticated set of photoreceptors that includes the red/ far-red light absorbing phytochromes. To better understand the early steps of phytochrome signaling, we performed a yeast two-hybrid screen and identified PIK (Phytochrome Interactor with Kelch Motifs) as a new phytochrome interactor. PIK interacts with phyA and phyB, and morphological phenotypes of a PIK mutant (pik-1) suggest that PIK acts as a positive regulator of phytochrome signaling. Downregulation of phyA is an important event in phytochrome signaling and occurs transcriptionally and post-translationally. We show that PIK downregulates PHYA transcription in response to light, and that pik-1 phenotypes result from persistent phyA. We propose that PIK acts as an adaptor between phytochromes and transcriptional machinery to ensure proper light regulation of PHYA. Inside the nucleus, phytochromes form subnuclear foci called nuclear bodies (NBs). We recovered two hypersensitive alleles of phyB in a mutant screen to identify factors involved in NB formation. These alleles exhibit hypersensitivity in terms of NB formation and inhibition of hypocotyl elongation by light. Mutations in phyB-501 and phyB-502 result in reduced intramolecular interactions, which may account for an impaired response to end-of-day far-red treatments. phyB-501 and phyB-502 also exhibit reduced affinity towards a downstream signaling factor. These mutations identify residues that are important for intra- and intermolecular interactions, and provide further evidence that NBs play a positive role in phytochrome signaling. Genetic evidence implicates the PIF/PIL subfamily of bHLH transcription factors as important players in phytochrome signaling. However, the field lacks the tools to address their function biochemically. To that end, we generated antibodies against six members of this subfamily. We also identified T-DNA insertion alleles in five of the six genes encoding these proteins. Together, this represents a powerful set of tools for studying the role of these bHLH proteins in phytochrome signaling. Lastly, we have identified a novel zinc knuckle/PLUS3 domain protein that directly regulates expression of key growth genes in a time-of-day fashion. Identification of TZP highlights the importance of daily synchronization of growth pathways, and underscores the utility of natural variation approaches to identifying new genes in light signaling