Functional and physical interaction of blue- and red-light sensors in Aspergillus nidulans

Light sensing is very important for organisms in all biological kingdoms to adapt to changing environmental conditions. It was discovered recently that plant-like phytochrome is involved in light sensing in the filamentous fungus Aspergillus nidulans[1]. Here, we show that phytochrome (FphA) is part of a protein complex containing LreA (WC-1) and LreB (WC-2) [2, 3], two central components of the Neurospora crassa blue-light-sensing system. We found that FphA represses sexual development and mycotoxin formation, whereas LreA and LreB stimulate both. Surprisingly, FphA interacted with LreB and with VeA, another regulator involved in light sensing and mycotoxin biosynthesis. LreB also interacted with LreA. All protein interactions occurred in the nucleus, despite cytoplasmic subfractions of the proteins. Whereas the FphA-VeA interaction was dependent on the presence of the linear tetrapyrrole in FphA, the interaction between FphA and LreB was chromophore independent. These results suggest that morphological and physiological differentiations in A. nidulans are mediated through a network consisting of FphA, LreA, LreB, and VeA acting in a large protein complex in the nucleus, sensing red and blue light.     

Mapping the interaction sites of <Emphasis Type="Italic">Aspergillus nidulans</Emphasis> phytochrome FphA with the global regulator VeA and the White Collar protein LreB

Aspergillus nidulans senses red and blue-light and employs a phytochrome and a Neurospora crassa White Collar (WC) homologous system for light perception and transmits this information into developmental decisions. Under light conditions it undergoes asexual development and in the dark it develops sexually. The phytochrome FphA consists of a light sensory domain and a signal output domain, consisting of a histidine kinase and a response regulator domain. Previously it was shown that the phytochrome FphA directly interacts with the WC-2 homologue, LreB and another regulator, VeA. In this paper we mapped the interaction of FphA with LreB to the histidine kinase and the response regulator domain at the C-terminus in vivo using the bimolecular fluorescence complementation assay and in vitro by co-immunoprecipitation. In comparison, VeA interacted with FphA only at the histidine kinase domain. We present evidence that VeA occurs as a phosphorylated and a non-phosphorylated form in the cell. The phosphorylation status of the protein was independent of the light receptors FphA, LreB and the WC-1 homologue LreA.     

The fungal phytochrome FphA from Aspergillus nidulans

The red light-sensing photoreceptor FphA from Aspergillus nidulans is involved in the regulation of developmental processes in response to light. Here we present extended biochemical and spectroscopic characterization of recombinant FphA using a synthetic gene with host-adapted codon usage. The recombinant photosensory domain FphAN753 was shown to display all features of a bona fide phytochrome. It covalently binds biliverdin as chromophore and undergoes red/far-red light-inducible photoconversion with both parent states being protonated. The large N-terminal variable extension of FphA exerts a stabilizing effect on the active Pfr state. Upon substitution of the highly conserved histidine 504, involved in the hydrogen-bonding network of the protein moiety and the chromophore, chromophore attachment and photoreversibility were completely impaired. FphA is a functional sensor histidine kinase with a strong red-light-dependent autophosphorylation activity. Furthermore, intermolecular trans-phosphorylation to the response regulator domain of a second monomer could be demonstrated. Interestingly, co-incubation of FphA and FphA variants led to enhanced autophosphorylation, including the "inactive" Pr form. The latter observed phenomenon might suggest that auto- and trans-phosphorylation activity is modulated by additional interaction partners leading to variable phosphorylation events that trigger a specific output response.     

Genome-wide analyses of light-regulated genes in Aspergillus nidulans reveal a complex interplay between different photoreceptors and novel photoreceptor functions

Fungi sense light of different wavelengths using blue-, green-, and red-light photoreceptors. Blue light sensing requires the “white-collar” proteins with flavin as chromophore, and red light is sensed through phytochrome. Here we analyzed genome-wide gene expression changes caused by short-term, low-light intensity illumination with blue-, red- or far-red light in Aspergillus nidulans and found that more than 1100 genes were differentially regulated. The largest number of up- and downregulated genes depended on the phytochrome FphA and the attached HOG pathway. FphA and the white-collar orthologue LreA fulfill activating but also repressing functions under all light conditions and both appear to have roles in the dark. Additionally, we found about 100 genes, which are red-light induced in the absence of phytochrome, suggesting alternative red-light sensing systems. We also found blue-light induced genes in the absence of the blue-light receptor LreA. We present evidence that cryptochrome may be part of this regulatory cue, but that phytochrome is essential for the response. In addition to in vivo data showing that FphA is involved in blue-light sensing, we performed spectroscopy of purified phytochrome and show that it responds indeed to blue light.     

A phosphorylation code of the Aspergillus nidulans global regulator VelvetA (VeA) determines specific functions

The velvet protein VeA is a global fungal regulator for morphogenetic pathways as well as for the control of secondary metabolism. It is found exclusively in filamentous fungi, where it fulfills conserved, but also unique functions in different species. The involvement of VeA in various morphogenetic and metabolic pathways is probably due to spatially and timely controlled specific protein–protein interactions with other regulators such as phytochrome (FphA) or velvet‐like proteins (VelB). Here we present evidence that Aspergillus nidulans VeA is a multi‐phosphorylated protein and hypothesize that at least four specific amino acids (T167, T170, S183 and Y254) undergo reversible phosphorylation to trigger development and sterigmatocystin biosynthesis. Double mutation of T167 to valine and T170 to glutamic acid exerted the largest effects with regards to sexual development and veA gene expression. In comparison with wild‐type VeA, which shuttles out of the nuclei after illumination this VeA variant showed stronger nuclear accumulation than the wild type, independent of the light conditions. The interaction between VeA and VelB or FphA, respectively, was affected in the T167V‐T170E mutant. Our results suggest complex regulation of the phosphorylation status of the VeA protein.     

Regulation of conidiation by light in Aspergillus nidulans

Light regulates several aspects of the biology of many organisms, including the balance between asexual and sexual development in some fungi. To understand how light regulates fungal development at the molecular level we have used Aspergillus nidulans as a model. We have performed a genome-wide expression analysis that has allowed us to identify >400 genes upregulated and >100 genes downregulated by light in developmentally competent mycelium. Among the upregulated genes were genes required for the regulation of asexual development, one of the major biological responses to light in A. nidulans, which is a pathway controlled by the master regulatory gene brlA. The expression of brlA, like conidiation, is induced by light. A detailed analysis of brlA light regulation revealed increased expression after short exposures with a maximum after 60 min of light followed by photoadaptation with longer light exposures. In addition to brlA, genes flbA-C and fluG are also light regulated, and flbA-C are required for the correct light-dependent regulation of the upstream regulator fluG. We have found that light induction of brlA required the photoreceptor complex composed of a phytochrome FphA, and the white-collar homologs LreA and LreB, and the fluffy genes flbA-C. We propose that the activation of regulatory genes by light is the key event in the activation of asexual development by light in A. nidulans.     

The protein kinase ImeB is required for light-mediated inhibition of sexual development and for mycotoxin production in Aspergillus nidulans

Spore formation is a common process in the developmental cycle of fungi. In the yeast Saccharomyces cerevisiae, Ime2 is a key protein kinase for the meiotic cell cycle, which precedes ascospore formation. Here, we analysed the IME2 -related imeB gene of the filamentous ascomycete Aspergillus nidulans. imeB deletion strains are retarded in growth and overproduce fertile sexual fruiting bodies in the presence of light, which normally represses sexual development. imeB mutants also display abnormal differentiation of sexual Hülle cells in submerged cultures. Increased sexual development of imeB mutants is dependent on VeA, a component of the heterotrimeric velvet complex. A combined deletion of imeB with the phytochrome fphA , a red light receptor, results in a complete loss of light response, suggesting that ImeB and FphA cooperate in light-mediated inhibition of sexual development. Furthermore, we found that imeB mutants fail to produce the mycotoxin sterigmatocystin, an aflatoxin precursor, and show that ImeB is needed for expression of the sterigmatocystin gene cluster. ImeB contains a TXY motif conserved in mitogen-activated protein kinases. This sequence element is essential for ImeB function. We conclude that ImeB is a mitogen-activated protein kinase-related protein kinase required for the co-ordinated control of light-dependent development with mycotoxin production.     

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.     

In vitro analysis of His-Asp phosphorelays in Aspergillus nidulans: the first direct biochemical evidence for the existence of His-Asp phosphotransfer systems in filamentous fungi

His-Asp phosphorelays are widespread signal transduction mechanisms in bacteria, fungi, and higher plants. In order to investigate a His-Asp phosphorelay network in filamentous fungi, which has been genetically characterized in part, we attempted to construct an in vitro phosphotransfer network in Aspergillus nidulans comprising all the necessary components. As a first step, we established an in vitro phosphotransfer system with a histidine-containing phosphotransmitter YpdA, a response regulator SrrA, and a bacterial histidine kinase ArcB as a phosphate donor. We demonstrated the phosphotransfer from ArcB to A. nidulans YpdA and the subsequent transfer from YpdA to SrrA. This is the first direct biochemical evidence for the presence of the phosphotransfer system in filamentous fungi. Furthermore, a retrograde phosphorylation from YpdA to FphA, a histidine kinase similar to bacterial phytochrome, was found. The overall picture of the His-Asp phosphorelays in A. nidulans is discussed based on the results of the in vitro study.     

Development of the Casparian strip is delayed by blue light in pea stems

To understand the regulatory mechanisms involved in tissue development by light, the kinetics of regulation of Casparian strip (CS) development in garden pea stems was studied. We found that short-term irradiation with white light delayed the development of the CS and used this delay to assess the quantitative effect of light on CS development. We examined the effect of the duration and fluence rates of white light treatment on CS development and observed a significant relationship between fluence and the delay in CS development indicating that the Bunsen–Roscoe law of reciprocity holds for this response. The effect of white light irradiation was not inhibited in the presence of a photosynthetic inhibitor, DCMU, or a carotenoid biosynthesis inhibitor, Norflurazon, indicating that the delay in CS development by light is a photomorphogenetic response rather than a subsidiary effect mediated by photosynthetic activity. An action spectrum for the response displayed a major peak in the blue-light region, suggesting a dominant role for blue-light receptors. A minor peak in the red-light region also suggested the possible involvement of phytochromes. Although phytochromes are known to contribute to blue-light responses, phytochrome-deficient mutants showed a normal delay of CS development in response to blue light, indicating that the response is not mediated by phytochrome and suggesting a role for one or more specific blue-light receptors.     

Effects of light-deprivation on development of photopositive behavior in Xenopus laevis tadpoles

The development of amphibian sensory systems and behavior is generally considered to proceed normally without reference to sensory experience during embryonic or larval stages. Most of the supporting research, however, has concentrated on the retinotectal (visual) systems of anurans and has ignored behaviors directed by other sensory systems. We demonstrate that early exposure to light is necessary for the development of photopositive behavior in Xenopus laevis tadpoles, a behavior probably directed by the pineal complex. Light-deprivation during the tadpoles' first 10 days of development results in a long-lasting reduction in the tadpoles' light preference. The development of a strong light preference is not influenced by light-deprivation before the tadpoles are 2 days old or after the tadpoles are 10 days old, but light-deprived tadpoles recover a weak light preference after subsequent days of rearing in the light. Lengthening the tadpoles' exposure to light during the first 10 days of development produces increasingly strong light preferences. Considering the important role of the pineal complex in guiding phototactic behaviors in anurans, we suggest that light-deprivation alters photopositive behavior in Xenopus tadpoles by altering the development of the pineal complex.     

Darkness during early postnatal development is required for normal circadian patterns in the adult rat

Early light experience influences the brain during development. Perinatal light exposure has an important effect on the development of the circadian system, although the role of quantity versus quality of light in this process is still unclear. We tested the development of the circadian rhythm of locomotor activity under constant bright light from the day of weaning, of six groups of rats raised under different light conditions during suckling. Results indicated that when rats received daily darkness during suckling (rats reared under constant darkness or light-dark cycles with dim or bright light) became arrhythmic when exposed to continuous bright light after weaning. However, those rats reared in the absence of darkness (constant dim or bright light, or alternating dim and bright light) developed a circadian rhythm, which was stronger and had a shorter period depending on the quantity of light received during suckling. Vasointestinal polypeptide immunoreactivity in the suprachiasmatic nucleus (SCN) was higher in those rats with weaker rhythms. However, no apparent differences among these groups were found in the melanopsin-expressing retinal ganglion cells, which provide the SCN with light input in the photoentrainment process. When bright light was shifted to dim light in three of the groups on day 57 after weaning, all of them generated a circadian rhythm with a longer period in those rats previously arrhythmic. Our results indicate the importance of the amount of light received at the early stages of life in the development of the circadian system and suggest that darkness is needed for the normal development of circadian behaviour.     

Characterisation of the interaction between cryptochromes and phytochromes in blue light-induced coiling and prehaustoria development of dodder (Cuscuta campestris) seedlings

Previous studies demonstrated that blue, near far‐red and a mixture of red and far‐red light stimulate, while red light inhibits coiling and prehaustoria development in young excised de‐etiolated dodder seedlings. In this study, evidence was obtained for the involvement of cryptochromes in the mediation of coiling and prehaustoria development in response to blue light. The results indicated that coiling and prehaustoria are affected by the addition of red or far‐red light and therefore are dependent on the state of phytochromes. Results also suggest the occurrence of a synergistic interaction between phytochromes (Pr) and cryptochromes and an antagonistic interaction between phytochromes (Pfr) and cryptochromes in mediating coiling and prehaustoria development. Gabaculine significantly reduced coiling and prehaustoria development under far‐red or a mixture of red and far‐red light but not under blue or a mixture of blue and far‐red light, indicating that cryptochromes are involved in blue‐induced coiling and prehaustoria development.     

Blue light effects on rose photosynthesis and photomorphogenesis

Through its impact on photosynthesis and morphogenesis, light is the environmental factor that most affects plant architecture. Using light rather than chemicals to manage plant architecture could reduce the impact on the environment. However, the understanding of how light modulates plant architecture is still poor and further research is needed. To address this question, we examined the development of two rose cultivars, Rosa hybrida‘Radrazz’ and Rosa chinensis‘Old Blush’, cultivated under two light qualities. Plants were grown from one‐node cuttings for 6 weeks under white or blue light at equal photosynthetic efficiencies. While plant development was totally inhibited in darkness, blue light could sustain full development from bud burst until flowering. Blue light reduced the net CO₂ assimilation rate of fully expanded leaves in both cultivars, despite increasing stomatal conductance and intercellular CO₂ concentrations. In ‘Radrazz’, the reduction in CO₂ assimilation under blue light was related to a decrease in photosynthetic pigment content, while in both cultivars, the chl a/b ratio increased. Surprisingly, blue light could induce the same organogenetic activity of the shoot apical meristem, growth of the metamers and flower development as white light. The normal development of rose plants under blue light reveals the strong adaptive properties of rose plants to their light environment. It also indicates that photomorphogenetic processes can all be triggered by blue wavelengths and that despite a lower assimilation rate, blue light can provide sufficient energy via photosynthesis to sustain normal growth and development in roses.     

Light regulation of asexual development in the rice blast fungus, Magnaporthe oryzae

Light is a major environmental factor that influences many biological processes. We characterized the roles of light in asexual development (including the formation of aerial hyphae and conidiophore) in Magnaporthe oryzae, which is the causal agent of rice blast disease. Our data revealed a complex nature of light regulation in the asexual developments of M. oryzae. Asexual development of M. oryzae is suppressed by blue light in a light/dark cycling environment and asexual spore release is controlled by both blue and red light. We demonstrated that even very dim light, about 10 micromol m(-2), is sufficient to suppress spore-release behavior in M. oryzae. We also generated knockout strains of a blue light receptor, mgwc-1, the M. oryzae homolog of white collar-1 in Neurospora crassa, and demonstrated blue-light-specific regulation in the asexual development and spore release in M. oryzae. Our findings in this agriculturally important pathogen, M. oryzae, broaden our understanding of the roles of light in fungal development.     

真菌发育过程中的蓝光诱导

本文对多种真菌在发育过程中受蓝光诱导的现象进行了综述。蓝光可以诱导多种真菌的形态发生和发育,包括孢子的产生和菌丝的延伸等。此外,蓝光还可引起真菌的生理和生化改变,如细胞膜对离子的通透性的变化、类胡萝卜素的合成等。真菌也存在一个蓝光信号系统,通过蓝光受体接受蓝光信号,导致了真菌的一系列形态发生和生理生化的变化。     

Blue light sensory systems in plants

Plant development is influenced by many environmental stimuli, including light, temperature and gravity. Of these stimuli, light is of particular importance because plants depend on it for energy and, thus, for survival. Moreover, virtually all stages of plant development are regulated in part by light through the action of various photosensory systems. Examples of light-regulated processes include germination, stem growth, leaf and root development, tropic responses and flower induction. This review provides an analysis of recent investigations of blue light sensory systems in plants. Current results suggest that plants respond to blue light through a complex photosensory network that incorporates the action of multiple blue light perception systems.     

Effects of Light, pH, Temperature, and Crowding on Megaspore Germination and Sporophyte Formation in Marsilea

Megaspore germination and sporophyte formation of Marsilea vestitaH. and G. was studied under different light, pH, temperature,and crowding conditions during a 6 d experimental period. Maximumgermination and sporophyte development occurs under relativelylow light intensity. Darkness and high light intensity inhibitsporophyte development. Selected wavelengths of light (red,far-red, green, blue) and darkness reduce both megaspore germinationand sporophyte development as compared to white light. Megasporesand sporophytes show maximum development at 25 ?C in light,whereas their development is reduced at all temperatures indarkness. The optimum pH range for megaspore germination ispH 7–8 and that for sporophyte development is pH 7. Sporocarpcontents alter the pH of unbuffered acidic and basic media towarda more optimal growth condition (pH 6–8) for megasporegermination. Megaspore germination and sporophyte developmentvary inversely with conditions of crowding. Root and leaf growthon developed sporophytes is very similar in most treatments.