Phytochromes are red/far-red photochromic biliprotein photoreceptors, which in plants regulate seed germination, stem extension, flowering time, and many other light effects. However, the structure/functional basis of the phytochrome photoswitch is still unclear. Here, we report the ground state structure of the complete sensory module of Cph1 phytochrome from the cyanobacterium Synechocystis 6803. Although the phycocyanobilin (PCB) chromophore is attached to Cys-259 as expected, paralleling the situation in plant phytochromes but contrasting to that in bacteriophytochromes, the ZZZssa conformation does not correspond to that expected from Raman spectroscopy. We show that the PHY domain, previously considered unique to phytochromes, is structurally a member of the GAF (cGMP phosphodiesterase/adenylyl cyclase/FhlA) family. Indeed, the tandem-GAF dumbbell revealed for phytochrome sensory modules is remarkably similar to the regulatory domains of cyclic nucleotide (cNMP) phosphodiesterases and adenylyl cyclases. A unique feature of the phytochrome structure is a long, tongue-like protrusion from the PHY domain that seals the chromophore pocket and stabilizes the photoactivated far-red-absorbing state (Pfr). The tongue carries a conserved PRxSF motif, from which an arginine finger points into the chromophore pocket close to ring D forming a salt bridge with a conserved aspartate residue. The structure that we present provides a framework for light-driven signal transmission in phytochromes.

Extensive studies in both lower and higher plants indicate that plant phytochrome photoreceptors signal not only by regulating transcription in the nucleus but also by acting within the cytoplasm, the latter signaling routes acting within minutes or even seconds and also providing directional information. Directional signals seem to arise from phytochromes attached anisotropically to the plasma membrane. Neochromes—phytochrome-phototropin hybrid photoreceptors probably attached to the plasma membrane—provide this signal in various ferns and perhaps certain algae but are absent from other groups. In mosses and probably higher plants too, a subpopulation of canonical phytochromes interact with phototropins at the plasma membrane and thereby steer directional responses. Phytochromes also seem able to regulate translation in the cytoplasm. This review discusses putative phytochrome functions in these contexts.

Phytochromes are red/far-red photochromic photoreceptors central to regulating plant development. Although they are known to enter the nucleus upon light activation and, once there, regulate transcription, this is not the complete picture. Various phytochrome effects are manifested much too rapidly to derive from changes in gene expression, whereas others seem to occur without phytochrome entering the nucleus. Phytochromes also guide directional responses to light, excluding a genetic signaling route and implying instead plasma membrane association and a direct cytoplasmic signal. However, to date, no such association has been demonstrated. Here we report that a phytochrome subpopulation indeed associates physically with another photoreceptor, phototropin, at the plasma membrane. Yeast two-hybrid methods using functional photoreceptor molecules showed that the phytochrome steering growth direction in Physcomitrella protonemata binds several phototropins specifically in the photoactivated Pfr state. Split-YFP studies in planta showed that the interaction occurs exclusively at the plasma membrane. Coimmunoprecipitation experiments provided independent confirmation of in vivo phy-phot binding. Consistent with this interaction being associated with a cellular signal, we found that phytochrome-mediated tropic responses are impaired in Physcomitrella phot ⁻ mutants. Split-YFP revealed a similar interaction between Arabidopsis phytochrome A and phototropin 1 at the plasma membrane. These associations additionally provide a functional explanation for the evolution of neochrome photoreceptors. Our results imply that the elusive phytochrome cytoplasmic signal arises through binding and coaction with phototropin at the plasma membrane.

Red and far-red light–sensing phytochromes are widespread in nature, occurring in plants, algae, fungi, and prokaryotes. Despite at least a billion years of evolution, their photosensory modules remain structurally and functionally similar. Conversely, nature has found remarkably different ways of transmitting light signals from the photosensor to diverse physiological responses. We summarize key features of phytochrome structure and function and discuss how these are correlated, from how the bilin environment affects the chromophore to how light induces cellular signals. Recent advances in the structural characterization of bacterial and plant phytochromes have resulted in paradigm changes in phytochrome research that we discuss in the context of present-day knowledge. Finally, we highlight questions that remain to be answered and suggest some of the benefits of understanding phytochrome structure and function.

The complete three-dimensional sensory module structures of the Pr ground state of Synechocystis 6803 Cph1 and the unusual Pfr ground state of the bacteriophytochrome PaBphP (PDB codes 2VEA and 3C2W respectively) have now been solved, revealing an asymmetrical dumbbell form made up of a PAS (Period/ARNT/Singleminded)–GAF (cGMP phosphodiesterase/adenylate cyclase/FhlA) bidomain carrying the chromophore and the smaller PHY (phytochrome-specific) domain. The PHY domain is structurally related to the GAF family, but carries an unusual tongue-like structure which contacts the larger lobe to seal the chromophore pocket. In 2VEA, the tongue makes intimate contact with the helical N-terminus; both the N-terminus and the tongue structures are quite different in 3C2W. As expected, the structures reveal ZZZssa and ZZEssa chromophore conformations in 2VEA and 3C2W respectively, associated with tautomeric differences in several nearby tyrosine residues. Two salt bridges on opposite sides of the chromophore, as well as the associations of the C-ring propionates also differ. It is still unclear, however, which of these structural differences are associated with bacteriophytochromes compared with Cph1 and plant-type phytochromes, the unusual 3C2W Pfr ground state functionality compared with the Pr ground state or the Pr compared with Pfr photoisomerism. To access the latter unambiguously, both Pr and Pfr structures of the same molecule are required. New solid-phase NMR data for Cph1 in the Pr, Pfr and freeze-trapped intermediate states reveal unexpected changes in the chromophore during Pfr→Pr photoconversion. These, together with our efforts to solve the three-dimensional structure of a complete phytochrome molecule are also described.