Many aspects of plant photomorphogenesis are controlled by the phytochrome (Phy) family of bilin-containing photoreceptors that detect red and far-red light by photointerconversion between a dark-adapted Pr state and a photoactivated Pfr state. Whereas 3D models of prokaryotic Phys are available, models of their plant counterparts have remained elusive. Here, we present the crystal structure of the photosensing module (PSM) from a seed plant Phy in the Pr state using the PhyB isoform from Arabidopsis thaliana. The PhyB PSM crystallized as a head-to-head dimer with strong structural homology to its bacterial relatives, including a 5(Z)syn, 10(Z)syn, 15(Z)anti configuration of the phytochromobilin chromophore buried within the cGMP phosphodiesterase/adenylyl cyclase/FhlA (GAF) domain, and a well-ordered hairpin protruding from the Phy-specific domain toward the bilin pocket. However, its Per/Arnt/Sim (PAS) domain, knot region, and helical spine show distinct structural differences potentially important to signaling. Included is an elongated helical spine, an extended β-sheet connecting the GAF domain and hairpin stem, and unique interactions between the region upstream of the PAS domain knot and the bilin A and B pyrrole rings. Comparisons of this structure with those from bacterial Phys combined with mutagenic studies support a toggle model for photoconversion that engages multiple features within the PSM to stabilize the Pr and Pfr end states after rotation of the D pyrrole ring. Taken together, this Arabidopsis PhyB structure should enable molecular insights into plant Phy signaling and provide an essential scaffold to redesign their activities for agricultural benefit and as optogenetic reagents.G iven the importance of sunlight to their survival and growth, plants have adopted a collection of photoreceptors and interconnected signaling cascades to optimize their photosynthetic potential and synchronize their lifecycles with circadian and seasonal rhythms. Chief among these are the phytochromes (Phys), a family of bilin (or open-chain tetrapyrrole)-containing red/far-red light-absorbing photoreceptors that provides spatial and time-dependent information by sensing the fluence rate, direction, duration, and color of a plant's light environment (1, 2). This information then regulates nearly all aspects of plant growth and development from seed germination to senescence. Notably, seed plants typically express three Phy isoforms (PhyA, PhyB, and PhyC) that control distinct and overlapping photoresponses, with PhyB having a dominant role in green tissues (2, 3).Phys are homodimers with each sister polypeptide divided into an N-terminal photosensory module (PSM) that absorbs light followed by an output module (OPM) that promotes dimerization and presumably, relays the light signals (1, 4). The PSM sequentially contains a Per/Arnt/Sim (PAS) domain of unknown function, a cGMP phosphodiesterase/adenylyl cyclase/FhlA (GAF) domain that cradles the bilin, and a Phy-specific (PHY) domain that stabilizes the photoactivated ...

Background: Phytochromes are dimeric bili-proteins central to photoperception by plants and microorganisms. Results: An informative perspective on photoconversion from the dark-adapted to the photoactivated state was provided by crystallographic and electron microscopic analyses of a bacterial version. Conclusion: Light-induced conformational changes in the bilin induce a large scale reorientation of the sister output modules. Significance: The structures offer a model for signal transmission by phytochromes.

Phytochromes (Phys) encompass a diverse collection of bilin-containing photoreceptors that help plants and microorganisms perceive light through photointerconversion between red light (Pr) and far-red light (Pfr)-absorbing states. In addition, Pfr reverts thermally back to Pr via a highly enthalpic process that enables temperature sensation in plants and possibly other organisms. Through domain analysis of the Arabidopsis PhyB isoform assembled recombinantly, coupled with measurements of solution size, photoconversion, and thermal reversion, we identified both proximal and distal features that influence all three metrics. Included are the downstream C-terminal histidine kinase-related domain known to promote dimerization and a conserved patch just upstream of an N-terminal Period/Arnt/Sim (PAS) domain, which upon removal dramatically accelerates thermal reversion. We also discovered that the nature of the bilin strongly influences Pfr stability. Whereas incorporation of the native bilin phytochromobilin into PhyB confers robust Pfr → Pr thermal reversion, that assembled with the cyanobacterial version phycocyanobilin, often used for optogenetics, has a dramatically stabilized Pfr state. Taken together, we conclude that Pfr acquisition and stability are impacted by a collection of opposing allosteric features that inhibit or promote photoconversion and reversion of Pfr back to Pr, thus allowing Phys to dynamically measure light, temperature, and possibly time.

Photoreceptors are biologically important for sensing changes in the color and intensity of ambient light and, for photosynthetic organisms, processing this light information to optimize food production through photosynthesis. Cyanobacteria are an evolutionarily and ecologically important prokaryotic group of oxygenic photosynthesizers that contain cyanobacteriochrome (CBCR) photoreceptors, whose family members sense nearly the entire visible spectrum of light colors. Some cyanobacteria contain 12 to 15 different CBCRs, and many family members contain multiple lightsensing domains. However, the complex interactions that must be occurring within and between these photoreceptors remain unexplored. Here we describe the regulation and photobiology of a unique CBCR called IflA (influenced by far-red light), demonstrating that a second CBCR called RcaE strongly regulates IflA abundance and that IflA uses two distinct photosensory domains to respond to four different light colors: blue, green, red, and farred. The absorption of red or far-red light by one domain affects the conformation of the other domain, and the rate of relaxation of one of these domains is influenced by the conformation of the other. Deletion of iflA results in delayed growth at low cell density, suggesting that IflA accelerates growth under this condition, apparently by sensing the ratio of red to far-red light in the environment. The types of complex photobiological interactions described here, both between unrelated CBCR family members and within photosensory domains of a single CBCR, may be advantageous for species using these photoreceptors in aquatic environments, where light color ratios are influenced by many biotic and abiotic factors. chromatic acclimation | chromatic adaptation | phytochrome | signal transduction | tetrapyrrole

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