Phytochrome evolution: Phytochrome genes in ferns and mosses

We have isolated phytochrome genes from the moss Physcomitrella , the fern Psilotum and PCR-generated phytochrome sequences from a few other ferns. The phytochrome gene of the moss Physcomitrella turned out not to contain the aberrant C-terminal third of the phytochrome from the moss Ceratodon , but the transmitter module-like sequences found in other phytochromes. A series of different phytochrome genes was detected in Psilotum . Differences between the amino acid sequences derived from them ranged from about 5 to more than 22%. Some of these genes are likely pseudogenes. Analysis by phylogenetic tree constructions revealed that higher and lower plant phytochromes evolved with different velocities. Lower plant phytochromes form a separate family characterized by a high degree of similarity. The amino acid differences between phytochrome types detected in a single species of higher plants are about two-fold higher than the differences between phytochromes of species of lower plants belonging to different divisions ( Physcomitrella and Selaginella ). Future studies on phytochrome sequences may eventually also throw light on the significance of Psilotum in the evolution of vascular plants.     

The Aspergillus nidulans phytochrome FphA represses sexual development in red light

Phytochrome photoreceptors sense red and far-red light through photointerconversion between two stable conformations, a process mediated by a linear tetrapyrrole chromophore. Originally, phytochromes were thought to be confined to photosynthetic organisms including cyanobacteria, but they have been recently discovered in heterotrophic bacteria and fungi, where little is known about their functions. It was shown previously in the ascomycetous fungus Aspergillus nidulans that asexual sporulation is stimulated and sexual development repressed by red light. The effect was reminiscent of a phytochrome response, and indeed phytochrome-like proteins were detected in several fungal genomes. All fungal homologs are more similar to bacterial than plant phytochromes and have multifunctional domains where the phytochrome region and histidine kinase domain are combined in a single protein with a C-terminal response-regulator domain. Here, we show that the A. nidulans phytochrome FphA binds a biliverdin chromophore, acts as a red-light sensor, and represses sexual development under red-light conditions. FphA-GFP is cytoplasmic and excluded from the nuclei, suggesting that red-light photoperception occurs in the cytoplasm. This is the first phytochrome experimentally characterized outside the plant and bacterial kingdoms and the second type of fungal protein identified that functions in photoperception.     

Right place,right time: Spatiotemporal light regulation of plant growth and development

Recent advances in our understanding of the roles of photoreceptors in light-dependent regulation of plant growth and development have been rapid and significant. Developments have been reported for numerous plant photoreceptor signaling pathways, yet researchers have made the most progress in increasing our comprehension of the roles of phytochrome family members, as well as the intracellular roles of phytochromes and phytochrome-interacting proteins in light-dependent signaling. An understudied, but vitally important, area of phytochrome biology centers on the roles phytochromes play in intercellular and interorgan signaling at the molecular level that results in the coordination of growth responses between distinct tissues and organs. This frontier of research into the spatiotemporal roles of phytochromes, and more generally plant photoreceptors, which is only beginning to be investigated and understood at the molecular genetic level, has a rich history of physiological data.Key words: cryptochrome, photomorphogenesis, photoreception, photoreceptor, phytochrome, spatiotemporal     

Mutations in the gene for the red/far-red light receptor phytochrome B alter cell elongation and physiological responses throughout Arabidopsis development.

Phytochromes are a family of plant photoreceptors that mediate physiological and developmental responses to changes in red and far-red light conditions. In Arabidopsis, there are genes for at least five phytochrome proteins. These photoreceptors control such responses as germination, stem elongation, flowering, gene expression, and chloroplast and leaf development. However, it is not known which red light responses are controlled by which phytochrome species, or whether the different phytochromes have overlapping functions. We report here that previously described hy3 mutants have mutations in the gene coding for phytochrome B (PhyB). These are the first mutations shown to lie in a plant photoreceptor gene. A number of tissues are abnormally elongated in the hy3(phyB) mutants, including hypocotyls, stems, petioles, and root hairs. In addition, the mutants flower earlier than the wild type, and they accumulate less chlorophyll. PhyB thus controls Arabidopsis development at numerous stages and in multiple tissues.     

Roles of different phytochromes in Arabidopsis photomorphogenesis

The red/far-red light-absorbing phytochromes play fundamental roles in photoperception of the light environment and the subsequent adaptation of plant growth and development. Higher plants possess multiple, discrete phytochromes, the apoproteins of which are the products of a family of divergent (PHY) genes. Arabidopsis thaliana has at least five PHY genes, encoding the apoproteins of phytochromes A-E. Through the analysis of mutants that are deficient in phytochrome A or B and the corresponding double mutant, it is becoming clear that these phytochromes perform both discrete and overlapping roles throughout plant development. Through analysis of the phyA phyB double mutant, it has been possible to define several responses that are mediated by other members of the phytochrome family. This article reviews some of the recent progress in the study of phytochrome-deficient mutants of the model plant Arabidopsis thaliana.     

Phytochrome gene diversity

The structures and functions of the phytochrome apoprotein genes (the PHY genes), their diversity across the plant kingdom, and their evolution are central concerns in the study of red-light sensing in plants. We summarize here recent advances in two areas relating to these topics: (1) the characteristics of the PHY gene family in Arabidopsis thaliana, the higher plant species for which the most extensive information on these genes is available, and (2) the similarity relationships, phylogeny, and evolutionary implications of PHY gene sequences and partial sequences which have been described from various plants. Together, these two areas of study, one directed at understanding in detail the phytochromes present in a single species and the other directed at a much broader understanding of PHY gene relatedness and distribution, are producing an increasingly clear picture of the diversity and evolution of plant red-light photoreceptors. Moreover, they suggest that the complexity of the phytochrome family has increased as land plants have evolved novel morphologies.     

Phytochrome and photoperiodic induction

The photoreceptor phytochrome has been extensively characterized at the chromophore, protein and gene level. It consists of a family of red/far-red reversible molecules and the genes for three members have been sequenced. Phytochromes are chromoproteins, which probably exist as dimers in vivo. Photoperiodism in higher plants involves the interaction of phytochrome with an endogenous timekeeping system. The interaction is complex, and several distinct actions of light can be distinguished. The possible involvement of different phytochromes in different actions of light in both long-day plants and short-day plants is discussed. Potential roles for different members of the phytochrome family and homo-and hetero-dimers of phytochrome are proposed.     

Action and function of phytochrome family members revealed through the study of mutant and transgenic plants

The adaptation of plant growth and development to changes in the light environment is dependent upon photoperception by information transducing photoreceptors. The red/far-red light-absorbing phytochromes are perhaps the best characterized of these regulatory photoreceptors. Higher plants possess multiple, discrete phytochromes, the apoprotein components of which are the products of a small, divergent gene family. Different phytochromes have different biochemical and physiological properties, and are differentially expressed in the growing plant. This has led to the proposal that different phytochromes have different physiological roles. Mutations that disrupt the normal perception of light signals have proved to be a valuable resource in assigning physiological roles to different phytochromes as well as in identifying residues/domains critical for phytochrome function and in attempting to elucidate the signal transduction pathway(s) downstream of phytochromes. This article reviews some recent progress in these areas from the study of conventional and transgenic photomorphogenic mutants.     

Arabidopsis phytochromes C and E have different spectral characteristics from those of phytochromes A and B

The red/far-red light absorbing phytochromes play a major role as sensor proteins in photomorphogenesis of plants. In Arabidopsis the phytochromes belong to a small gene family of five members, phytochrome A (phyA) to E (phyE). Knowledge of the dynamic properties of the phytochrome molecules is the basis of phytochrome signal transduction research. Beside photoconversion and destruction, dark reversion is a molecular property of some phytochromes. A possible role of dark reversion is the termination of signal transduction. Since Arabidopsis is a model plant for biological and genetic research, we focussed on spectroscopic characterization of Arabidopsis phytochromes, expressed in yeast. For the first time, we were able to determine the relative absorption maxima and minima for a phytochrome C (phyC) as 661/725 nm and for a phyE as 670/724 nm. The spectral characteristics of phyC and E are strictly different from those of phyA and B. Furthermore, we show that both phyC and phyE apoprotein chromophore adducts undergo a strong dark reversion. Difference spectra, monitored with phycocyanobilin and phytochromobilin as the apoprotein's chromophore, and in vivo dark reversion of the Arabidopsis phytochrome apoprotein phycocyanobilin adducts are discussed with respect to their physiological function.     

Phytochromes and photomorphogenesis in Arabidopsis.

Plants have evolved exquisite sensory systems for monitoring their light environment. The intensity, quality, direction and duration of light are continuously monitored by the plant and the information gained is used to modulate all aspects of plant development. Several classes of distinct photoreceptors, sensitive to different regions of the light spectrum, mediate the developmental responses of plants to light signals. The red-far-red light-absorbing, reversibly photochromic phytochromes are perhaps the best characterized of these. Higher plants possess a family of phytochromes, the apoproteins of which are encoded by a small, divergent gene family. Arabidopsis has five apophytochrome-encoding genes, PHYA-PHYE. Different phytochromes have discrete biochemical and physiological properties, are differentially expressed and are involved in the perception of different light signals. Photoreceptor and signal transduction mutants of Arabidopsis are proving to be valuable tools in the molecular dissection of photomorphogenesis. Mutants deficient in four of the five phytochromes have now been isolated. Their analysis indicates considerable overlap in the physiological functions of different phytochromes. In addition, mutants defining components acting downstream of the phytochromes have provided evidence that different members of the family use different signalling pathways.     

Bottom-up Assembly of the Phytochrome Network

Plants have developed sophisticated systems to monitor and rapidly acclimate to environmental fluctuations. Light is an essential source of environmental information throughout the plant’s life cycle. The model plant Arabidopsis thaliana possesses five phytochromes (phyA-phyE) with important roles in germination, seedling establishment, shade avoidance, and flowering. However, our understanding of the phytochrome signaling network is incomplete, and little is known about the individual roles of phytochromes and how they function cooperatively to mediate light responses. Here, we used a bottom-up approach to study the phytochrome network. We added each of the five phytochromes to a phytochrome-less background to study their individual roles and then added the phytochromes by pairs to study their interactions. By analyzing the 16 resulting genotypes, we revealed unique roles for each phytochrome and identified novel phytochrome interactions that regulate germination and the onset of flowering. Furthermore, we found that ambient temperature has both phytochrome-dependent and -independent effects, suggesting that multiple pathways integrate temperature and light signaling. Surprisingly, none of the phytochromes alone conferred a photoperiodic response. Although phyE and phyB were the strongest repressors of flowering, both phyB and phyC were needed to confer a flowering response to photoperiod. Thus, a specific combination of phytochromes is required to detect changes in photoperiod, whereas single phytochromes are sufficient to respond to light quality, indicating how phytochromes signal different light cues.     

Changes in photoperiod or temperature alter the functional relationships between phytochromes and reveal roles for phyD and phyE

The phytochromes are one of the means via which plants obtain information about their immediate environment and the changing seasons. Phytochromes have important roles in developmental events such as the switch to flowering, the timing of which can be crucial for the reproductive success of the plant. Analysis of phyB mutants has revealed that phyB plays a major role in this process. We have recently shown, however, that the flowering phenotype of the phyB monogenic mutant is temperature dependent. A modest reduction in temperature to 16 degrees C was sufficient to abolish the phyB mutant early-flowering phenotype present at 22 degrees C. Using mutants null for one or more phytochrome species, we have now shown that phyA, phyD, and phyE, play greater roles with respect to phyB in the control of flowering under cooler conditions. This change in the relative contributions of individual phytochromes appears to be important for maintaining control of flowering in response to modest alterations in ambient temperature. We demonstrate that changes in ambient temperature or photoperiod can alter the hierarchy and/or the functional relationships between phytochrome species. These experiments reveal new roles for phyD and phyE and provide valuable insights into how the phytochromes help to maintain development in the natural environment.     

Mutant analyses define multiple roles for phytochrome C in Arabidopsis photomorphogenesis

The analysis of Arabidopsis mutants deficient in the A, B, D, and E phytochromes has revealed that each of these phytochrome isoforms has both distinct and overlapping roles throughout plant photomorphogenesis. Although overexpression studies of phytochrome C (phyC) have suggested photomorphogenic roles for this receptor, conclusive evidence of function has been lacking as a result of the absence of mutants in the PHYC locus. Here, we describe the isolation of a T-DNA insertion mutant of phyC (phyC-1), the subsequent creation of mutant lines deficient in multiple phytochrome combinations, and the physiological characterization of these lines. In addition to operating as a weak red light sensor, phyC may perform a significant role in the modulation of other photoreceptors. phyA and phyC appear to act redundantly to modulate the phyB-mediated inhibition of hypocotyl elongation in red light and to function together to regulate rosette leaf morphology. In addition, phyC performs a significant role in the modulation of blue light sensing. Several of these phenotypes are supported by the parallel analysis of a quadruple mutant deficient in phytochromes A, B, D, and E, which thus contains only active phyC. Together, these data suggest that phyC has multiple functions throughout plant development that may include working as a coactivator with other phytochromes and the cryptochrome blue light receptors.     

The system of phytochromes: photobiophysics and photobiochemistry in vivo.

Phytochrome is a key photoregulation pigment in plants which determines the strategy of their development throughout their life cycle. The major achievement in the recent investigations of the pigment is the discovery of its structural and functional heterogeneity: existence of a family of phytochromes (phyA-phyE) differing by the apoprotein was demonstrated. We approach this problem by investigating the chromophore component of the pigment with the use of the developed method of in vivo low-temperature fluorescence spectroscopy of phytochrome. In etiolated plants, phytochrome fluorescence was detected and attributed to its red-light absorbing form (Pr) and the first photoproduct (lumi-R), and a scheme of the photoreaction in phytochrome, a distinction of which is the activation barrier in the excited state, was put forward. It was found that the spectroscopic and photochemical characteristics of Pr depend on the plant species and phytochrome mutants and overexpressors used, on localization of the pigment in organs and tissues, plant age, effect of preillumination and other physiological factors. This variability of the parameters was interpreted as the existence of at least two phenomenological Pr populations, which differ by their spectroscopic characteristics and activation parameters of the Pr --> lumi-R photoreaction (in particular, by the extent of the Pr --> lumi-R photoconversion at low temperatures, gamma1): the longer-wavelength major and variable by its content in plant tissues Pr' with gamma1 = 0.5 and the shorter-wavelength minor relatively constant Pr" with gamma1 < or = 0.05. The analysis of the phytochrome mutants and overexpressors allows a conclusion that phytochrome A (phyA), which dominates in etiolated seedlings, is presented by two isoforms attributed to Pr' and Pr" (phyA' and phyA", respectively). Phytochrome B (phyB) accounts for less than 10% of the total phytochrome fluorescence and belongs to the Pr" type. It is also characterized by the relatively low extent of the Pr photoconversion into the far-red-light absorbing physiologically active phytochrome form, Pfr. Fluorescence of the minor phytochromes (phyC-phyE) is negligible. The recently discovered phytochrome of the cyanobacterium Synechocystis also belongs to the phenomenological Pr" type. PhyA' is a light-labile and soluble fraction, while phyA" is a relatively light-stable and, possibly, membrane (protein)-associated. Experiments with transgenic tobacco plants overexpressing full-length and C- and N-terminally truncated oat phytochrome A suggest that phyA' and phyA" might differ by the post-translational modification of the small N-terminal segment (amino acid residues 7-69) of the pigment. PhyA' is likely to be active in the de-etiolation processes while phyA" together with phyB, in green plants as revealed by the experiments on transgenic potato plants and phytochrome mutants of Arabidopsis and pea with altered levels of phytochromes A and B and modified phenotypes. And finally, within phyA', there are three subpopulations which are, possibly, different conformers of the chromophore. Thus, there is a hierarchical system of phytochromes which include: (i) different phytochromes; (ii) their post-translationally modified states and (iii) conformers within one molecular type. Its existence might be the rationale for the multiplicity of the photoregulation reactions in plants mediated by phytochrome.     

Arabidopsis phytochrome a is modularly structured to integrate the multiple features that are required for a highly sensitized phytochrome

Phytochrome is a red (R)/far-red (FR) light-sensing photoreceptor that regulates various aspects of plant development. Among the members of the phytochrome family, phytochrome A (phyA) exclusively mediates atypical phytochrome responses, such as the FR high irradiance response (FR-HIR), which is elicited under prolonged FR. A proteasome-based degradation pathway rapidly eliminates active Pfr (the FR-absorbing form of phyA) under R. To elucidate the structural basis for the phyA-specific properties, we systematically constructed 16 chimeric phytochromes in which each of four parts of the phytochrome molecule, namely, the N-terminal extension plus the Per/Arnt/Sim domain (N-PAS), the cGMP phosphodiesterase/adenyl cyclase/FhlA domain (GAF), the phytochrome domain (PHY), and the entire C-terminal half, was occupied by either the phyA or phytochrome B sequence. These phytochromes were expressed in transgenic Arabidopsis thaliana to examine their physiological activities. Consequently, the phyA N-PAS sequence was shown to be necessary and sufficient to promote nuclear accumulation under FR, whereas the phyA sequence in PHY was additionally required to exhibit FR-HIR. Furthermore, the phyA sequence in PHY alone substantially increased the light sensitivity to R. In addition, the GAF phyA sequence was important for rapid Pfr degradation. In summary, distinct structural modules, each of which confers different properties to phyA, are assembled on the phyA molecule.     

Phytochrome regulation of seed germination

Seed germination of many plant species is influenced by light. Of the various photoreceptor systems, phytochrome plays an especially important role in seed germination. The existence of at least five phytochrome genes has led to the proposal that different members of the family have different roles in the photoregulation of seed germination. Physiological analysis of seed germination ofArabidopsis thaliana (L.) Heynh. with phytochrome-deficient mutants showed for the first time that phytochrome A and phytochrome B modulate the timing of seed germination in distinct actions. Phytochrome A photo-irreversibly triggers the photoinduction of seed germination after irradiation with extremely low fluence light in a wide range of wavelengths, from UV-A, to visible, to far-red. In contrast, phytochrome B mediates the well-characterized photoreversible reaction, responding to red and far-red light of fluences four orders of magnitude higher than those to which PhyA responds. Wild plants, such asA. thaliana, survive under ground as dormant seeds for long periods, and the timing of seed germination is crucial for optimizing growth and reproduction. It therefore seems reasonable for plants to possess at least two different physiological systems for sensing the light environment over a wide spectral range with exquisite sensitivity of different phytochromes. This redundancy seems to enhance plant survival in a fluctuating environment.