RESUMO
Intrinsically photosensitive retinal ganglion cells (ipRGCs) serve as primary photoceptors by expressing the photopigment, melanopsin, and also as retinal relay neurons for rod and cone signals en route to the brain, in both cases for the purpose of non-image vision as well as aspects of image vision. So far, six subtypes of ipRGCs (M1 through M6) have been characterized. Regarding their phototransduction mechanisms, we have previously found that, unconventionally, rhabdomeric (microvillous) and ciliary signaling motifs co-exist within a given M1-, M2-, and M4-ipRGC, with the first mechanism involving PLCß4 and TRPC6,7 channels and the second involving cAMP and HCN channels. We have now examined M3-, M5-, and M6-cells and found that each cell likewise uses both signaling pathways for phototransduction, despite differences in the percentage representation by each pathway in a given ipRGC subtype for bright-flash responses (and saturated except for M6-cells). Generally, M3- and M5-cells show responses quite similar in kinetics to M2-responses, and M6-cell responses resemble broadly those of M1-cells although much lower in absolute sensitivity and amplitude. Therefore, similar to rod and cone subtypes in image vision, ipRGC subtypes possess the same phototransduction mechanism(s) even though they do not show microvilli or cilia morphologically.
Assuntos
Neurônios Retinianos , Visão Ocular , Transdução de Sinal Luminoso/fisiologia , Células Ganglionares da Retina/fisiologia , Células Fotorreceptoras Retinianas Cones/metabolismo , Neurônios Retinianos/metabolismo , Opsinas de Bastonetes/metabolismoRESUMO
Melanopsin signaling within intrinsically photosensitive retinal ganglion cell (ipRGC) subtypes impacts a broad range of behaviors from circadian photoentrainment to conscious visual perception. Yet, how melanopsin phototransduction within M1-M6 ipRGC subtypes impacts cellular signaling to drive diverse behaviors is still largely unresolved. The identity of the phototransduction channels in each subtype is key to understanding this central question but has remained controversial. In this study, we resolve two opposing models of M4 phototransduction, demonstrating that hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are dispensable for this process and providing support for a pathway involving melanopsin-dependent potassium channel closure and canonical transient receptor potential (TRPC) channel opening. Surprisingly, we find that HCN channels are likewise dispensable for M2 phototransduction, contradicting the current model. We instead show that M2 phototransduction requires TRPC channels in conjunction with T-type voltage-gated calcium channels, identifying a novel melanopsin phototransduction target. Collectively, this work resolves key discrepancies in our understanding of ipRGC phototransduction pathways in multiple subtypes and adds to mounting evidence that ipRGC subtypes employ diverse phototransduction cascades to fine-tune cellular responses for downstream behaviors.
Assuntos
Transdução de Sinal Luminoso , Células Ganglionares da Retina , Opsinas de Bastonetes , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Transdução de Sinal Luminoso/fisiologia , Células Ganglionares da Retina/fisiologia , Opsinas de Bastonetes/metabolismo , Visão Ocular , Animais , CamundongosRESUMO
Nonimage-forming vision in mammals is mediated primarily by melanopsin (OPN4)-expressing, intrinsically photosensitive retinal ganglion cells (ipRGCs). In mouse M1-ipRGCs, melanopsin predominantly activates, via Gαq,11,14, phospholipase C-ß4 to open transient receptor 6 (TRPC6) and TRPC7 channels. In M2- and M4-ipRGCs, however, a prominent phototransduction mechanism involves the opening of hyperpolarization- and cyclic nucleotide-gated channels via cyclic nucleotide, although the upstream steps remain uncertain. We report here experiments, primarily on M4-ipRGCs, with photo-uncaging of cyclic nucleotides and virally expressed CNGA2 channels to conclude that the second messenger is cyclic adenosine monophosphate (cAMP) - very surprising considering that cyclic guanosine monophosphate (cGMP) is used in almost all cyclic nucleotide-mediated phototransduction mechanisms across the animal kingdom. We further found that the upstream G protein is likewise Gq, which via its Gßγ subunits directly activates adenylyl cyclase (AC). Our findings are a demonstration in a native cell of a cross-motif GPCR signaling pathway from Gq directly to AC with a specific function.
Assuntos
Adenilil Ciclases , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP , Transdução de Sinal Luminoso , Células Ganglionares da Retina , Animais , Camundongos , Adenilil Ciclases/genética , Adenilil Ciclases/metabolismo , Transdução de Sinal Luminoso/fisiologia , Mamíferos/metabolismo , Nucleotídeos Cíclicos/metabolismo , Células Ganglionares da Retina/metabolismo , Células Ganglionares da Retina/fisiologia , Opsinas de Bastonetes/metabolismo , Transdução de Sinais/fisiologia , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/metabolismoRESUMO
Retinal photoreceptor cells, rods and cones, convert photons of light into chemical and electrical signals as the first step of the visual transduction cascade. Although the chemical processes in the phototransduction system are very similar to each other in these photoreceptors, the light sensitivity and time resolution of the photoresponse in rods are functionally different than those in the photoresponses of cones. To systematically investigate how photoresponses are divergently regulated in rods and cones, we have developed a detailed mathematical model on the basis of the Hamer model. The current model successfully reconstructed light intensity-, ATP- and GTP-dependent changes in concentrations of phosphorylated visual pigments (VPs), activated transducins (Tr*s) and phosphodiesterases (PDEs) in rods and cones. In comparison to rods, the lower light sensitivity of cones was attributed not only to the lower affinity of activated VPs for Trs but also to the faster desensitization of the VPs. The assumption of an intermediate inactive state, MIIi, in the thermal decay of activated VPs was essential for inducing faster inactivation of VPs in rods, and possibly also in cones.
Assuntos
Fotofobia , Células Fotorreceptoras Retinianas Bastonetes , Humanos , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Células Fotorreceptoras Retinianas Cones/metabolismo , Transdução de Sinal Luminoso/fisiologia , Transducina/metabolismoRESUMO
The requirement of vitamin A for the synthesis of the visual chromophore and the light-sensing pigments has been studied in vertebrate and invertebrate model organisms. To identify the molecular mechanisms that orchestrate the ocular response to vitamin A deprivation, we took advantage of the fact that Drosophila melanogaster predominantly requires vitamin A for vision, but not for development or survival. We analyzed the impacts of vitamin A deficiency on the morphology, the lipidome, and the proteome of the Drosophila eye. We found that chronic vitamin A deprivation damaged the light-sensing compartments and caused a dramatic loss of visual pigments, but also decreased the molar abundance of most phototransduction proteins that amplify and transduce the visual signal. Unexpectedly, vitamin A deficiency also decreased the abundances of specific subunits of mitochondrial TCA cycle and respiratory chain components but increased the levels of cuticle- and lens-related proteins. In contrast, we found no apparent effects of vitamin A deficiency on the ocular lipidome. In summary, chronic vitamin A deficiency decreases the levels of most components of the visual signaling pathway, but also affects molecular pathways that are not vision-specific and whose mechanistic connection to vitamin A remains to be elucidated.
Assuntos
Proteínas de Drosophila , Deficiência de Vitamina A , Animais , Drosophila , Proteínas de Drosophila/genética , Drosophila melanogaster , Transdução de Sinal Luminoso/fisiologia , Proteoma , Vitamina ARESUMO
Green rods (GRs) represent a unique type of photoreceptor to be found in the retinas of anuran amphibians. These cells harbor a cone-specific blue-sensitive visual pigment but exhibit morphology of the outer segment typical for classic red rods (RRs), which makes them a perspective model object for studying cone-rod transmutation. In the present study, we performed detailed electrophysiological examination of the light sensitivity, response kinetics and parameters of discrete and continuous dark noise in GRs of the two anuran species: cane toad and marsh frog. Our results confirm that anuran GRs are highly specialized nocturnal vision receptors. Moreover, their rate of phototransduction quenching appeared to be about two-times slower than in RRs, which makes them even more efficient single photon detectors. The operating intensity ranges for two rod types widely overlap supposedly allowing amphibians to discriminate colors in the scotopic region. Unexpectedly for typical cone pigments but in line with some previous reports, the spontaneous isomerization rate of the GR visual pigment was found to be the same as for rhodopsin of RRs. Thus, our results expand the knowledge on anuran GRs and show that these are even more specialized single photon catchers than RRs, which allows us to assign them a status of "super-rods".
Assuntos
Transdução de Sinal Luminoso/fisiologia , Células Fotorreceptoras Retinianas Cones/metabolismo , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Animais , Anuros/anatomia & histologia , Isomerismo , Cinética , Luz , Visão Noturna/fisiologia , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/fisiologia , Retina/anatomia & histologia , Retina/metabolismo , Células Fotorreceptoras Retinianas Cones/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Rodopsina , Opsinas de Bastonetes , Visão Ocular/fisiologiaRESUMO
Bioluminescence, the emission of light catalysed by luciferases, has evolved in many taxa from bacteria to vertebrates and is predominant in the marine environment. It is now well established that in animals possessing a nervous system capable of integrating light stimuli, bioluminescence triggers various behavioural responses and plays a role in intra- or interspecific visual communication. The function of light emission in unicellular organisms is less clear and it is currently thought that it has evolved in an ecological framework, to be perceived by visual animals. For example, while it is thought that bioluminescence allows bacteria to be ingested by zooplankton or fish, providing them with favourable conditions for growth and dispersal, the luminous flashes emitted by dinoflagellates may have evolved as an anti-predation system against copepods. In this short review, we re-examine this paradigm in light of recent findings in microorganism photoreception, signal integration and complex behaviours. Numerous studies show that on the one hand, bacteria and protists, whether autotrophs or heterotrophs, possess a variety of photoreceptors capable of perceiving and integrating light stimuli of different wavelengths. Single-cell light-perception produces responses ranging from phototaxis to more complex behaviours. On the other hand, there is growing evidence that unicellular prokaryotes and eukaryotes can perform complex tasks ranging from habituation and decision-making to associative learning, despite lacking a nervous system. Here, we focus our analysis on two taxa, bacteria and dinoflagellates, whose bioluminescence is well studied. We propose the hypothesis that similar to visual animals, the interplay between light-emission and reception could play multiple roles in intra- and interspecific communication and participate in complex behaviour in the unicellular world.
Assuntos
Transdução de Sinal Luminoso/fisiologia , Proteínas Luminescentes/metabolismo , Células Fotorreceptoras/fisiologia , Animais , Bactérias/metabolismo , Comunicação , Dinoflagellida/metabolismo , Luz , Luciferases/metabolismo , Medições Luminescentes , Células Fotorreceptoras/metabolismo , Plâncton/metabolismo , Comportamento PredatórioRESUMO
Computational functionality has been implemented successfully on chemical reactions in living systems. In the case of Belousov-Zhabotinsky (BZ) reaction, this was achieved by using collision-based techniques and by exploiting the light sensitivity of BZ. In order to unveil the computational capacity of the light sensitive BZ medium and the possibility to implement re-configurable logic, the design of multiple logic gates in a fixed BZ reservoir was investigated. The three basic logic gates (namely NOT, OR and AND) were studied to prove the Turing completeness of the architecture. Namely, all possible Boolean functions can be implemented as a combination of these logic gates. Nonetheless, a more complicated logic function was investigated, aiming to illustrate further capabilities of a fixed size BZ reservoir. The experiments executed within this study were implemented with a Cellular Automata (CA)-based model of the Oregonator equations that simulate excitation and wave propagation on a light sensitive BZ thin film. Given that conventional or von Neumann architecture computations is proved possible on the proposed configuration, the next step would be the realization of unconventional types of computation, such as neuromorphic and fuzzy computations, where the chemical substrate may prove more efficient than silicon.
Assuntos
Autômato Celular , Simulação por Computador , Transdução de Sinal Luminoso/fisiologia , Luz , Lógica , Animais , Fenômenos Químicos , HumanosRESUMO
Plants acclimate to shade by sensing light signals such as low photosynthetic active radiation (PAR), low blue light (BL) levels and low red-to-far red ratios (R:FR) trough plant photoreceptors cross talk. We previously demonstrated that grapevine is irresponsive to variations in R:FR and that BL-attenuation mediates morphological and architectural responses to shade increasing light interception and absorption efficiencies. However, we wondered if grapevine respond to low R:FR when BL is attenuated at the same time. Our objective was to evaluate if morphological, architectural and hydraulic acclimation to shade is mediated by low R:FR ratios and BL attenuation. To test this, we carried out experiments under natural radiation, manipulating light quality by selective sunlight exclusion and light supplementation. We grew grapevines under low PAR (LP) and four high PAR (HP) treatments: HP, HP plus FR supplementation (HPâ¯+â¯FR), HP with BL attenuation (HP-B) and HP with BL attenuation plus FR supplementation (HP-Bâ¯+â¯FR). We found that plants grown under HP-B and HP-Bâ¯+â¯FR had similar morphological (stem and petiole length, leaf thickness and area), architectural (laminae' angles) and anatomical (stomatal density) traits than plants grown under LP. However, only LP plants presented lower stomata differentiation, lower δ13C and hence lower water use efficiency. Therefore, even under a BL and R:FR attenuated environment, morphological and architectural responses were modulated by BL but not by variation in R:FR. Meanwhile water relations were affected by PAR intensity but not by changes in light quality. Knowing grapevine responses to light quantity and quality are indispensable to adopt tools or design new cultural management practices that manipulate irradiance in the field intending to improve crop performance.
Assuntos
Aclimatação/fisiologia , Transdução de Sinal Luminoso/fisiologia , Luz , Fotorreceptores de Plantas/fisiologia , Transpiração Vegetal/fisiologia , Vitis/anatomia & histologia , Vitis/crescimento & desenvolvimento , Produtos Agrícolas/anatomia & histologia , Produtos Agrícolas/crescimento & desenvolvimentoRESUMO
A growing body of evidence supports the modulation of pain by light exposure. As such, phototherapy is being increasingly utilized for the management of a variety of pain conditions. The modes of delivery, and hence applications of phototherapy, vary by wavelength, intensity, and route of exposure. As such, differing mechanisms of action exist depending upon those parameters. Cutaneous application of red light (660 nm) has been shown to reduce pain in neuropathies and complex regional pain syndrome-I, whereas visual application of the same wavelength of red light has been reported to exacerbate migraine headache in patients and lead to the development of functional pain in animal models. Interestingly visual exposure to green light can result in reduction in pain in variety of pain conditions such as migraine and fibromyalgia. Cutaneous application typically requires exposure on the order of minutes, whereas visual application requires exposure on the order of hours. Both routes of exposure elicit changes centrally in the brainstem and spinal cord, and peripherally in the dorsal root ganglia and nociceptors. The mechanisms of photobiomodulation of pain presented in this review provide a foundation in furtherance of exploration of the utility of phototherapy as a tool in the management of pain. PERSPECTIVE: This review synopsizes the pathways and mechanisms through which light modulates pain and the therapeutic utility of different colors and exposure modalities of light on pain. Recent advances in photobiomodulation provide a foundation for understanding this novel treatment for pain on which future translational and clinical studies can build upon.
Assuntos
Manejo da Dor , Dor/etiologia , Fototerapia , Humanos , Transdução de Sinal Luminoso/fisiologia , Vias Neurais/fisiologia , Dor/fisiopatologia , Dor/psicologiaRESUMO
Photoreceptors mediate the first step of vision, transducing light and passing signals to retinal neurons that ultimately send signals along the optic nerve to the brain. A functional deficiency in the photoreceptors, due to either congenital or acquired disease, can significantly affect an individual's sight and quality of life. Methods for quantifying the health and function of photoreceptors are essential for understanding both the progression of disease and the efficacy of treatment. Given that emerging treatments such as gene, stem cell, and small molecule therapy are designed to operate at the cellular scale, it is desirable to monitor function at the commensurate resolution of individual photoreceptors. Previously, non-invasive imaging methods for visualizing photoreceptor mosaic structure have been used to infer photoreceptor health, but these methods do not assess function directly. Conversely, most functional techniques, such as ERG and conventional microperimetry, measure function by aggregating the effects of signals from many photoreceptors. We have previously shown that stimulus-evoked intrinsic changes in intensity can be measured reliably in populations of cone photoreceptors in the intact human eye, a measurement we refer to more generally as the cone optoretinogram. Here we report that we can resolve the intensity optoretinogram at the level of individual cones. Moreover, we show that the individual cone optoretinogram exhibits two key signatures expected of a functional measure. First, responses in individual cones increase systematically as a function of stimulus irradiance. Second, we can use the amplitude of the functional response to middle wavelength (545 nm) light to separate the population of short-wavelength-sensitive (S) cones from the population of middle- and long-wavelength-sensitive (L and M) cones. Our results demonstrate the promise of optoretinography as a direct diagnostic measure of individual cone function in the living human eye.
Assuntos
Transdução de Sinal Luminoso/fisiologia , Óptica e Fotônica , Células Fotorreceptoras Retinianas Cones/citologia , Células Fotorreceptoras Retinianas Cones/fisiologia , Humanos , OftalmoscopiaRESUMO
Phototransduction reactions in the rhabdomeric photoreceptor are profoundly stochastic due to the small number of participating molecules and small reaction space. The resulting quantum bumps (QBs) vary in their timing (latency), amplitudes and durations, and these variabilities within each cell are not correlated. Using modeling and electrophysiological recordings, we investigated how the QB properties depend on the cascade speed and how they influence signal transfer. Parametric analysis in the model supported by experimental data revealed that faster cascades elicit larger and narrower QBs with faster onsets and smaller variabilities than slower cascades. Latency dispersion was stronger affected by modification of upstream than downstream activation parameters. The variability caused by downstream modifications closely matched the experimental variability. Frequency response modeling showed that corner frequency is a reciprocal function of the characteristic duration of the multiphoton response, which, in turn, is a non-linear function of QB duration and latency dispersion. All QB variabilities contributed noise but only latency dispersion slowed and spread multiphoton responses, lowering the corner frequency. Using the discovered QB correlations, we evaluated transduction noise for dissimilar species and two extreme adaptation states, and compared it to photon noise. The noise emitted by the cascade was non-additive and depended non-linearly on the interaction between the QB duration and the three QB variabilities. Increased QB duration strongly suppressed both noise and corner frequency. This trade-off might be acceptable for nocturnal but not diurnal species because corner frequency is the principal determinant of information capacity. To offset the increase in noise accompanying the QB narrowing during light adaptation and the response-expanding effect of latency dispersion, the cascade accelerates. This explains the widespread evolutionary tendency of diurnal fliers to have fast phototransduction, especially after light adaptation, which thus appears to be a common adaptation to contain stochasticity, improve SNR and expand the bandwidth.
Assuntos
Insetos/fisiologia , Transdução de Sinal Luminoso/fisiologia , Modelos Biológicos , Células Fotorreceptoras de Invertebrados/fisiologia , Animais , Evolução Biológica , Biologia Computacional , Simulação por Computador , Fenômenos Eletrofisiológicos , Cinética , Microvilosidades/fisiologia , Dinâmica não Linear , Periplaneta/fisiologia , Células Fotorreceptoras de Vertebrados/fisiologia , Teoria Quântica , Razão Sinal-Ruído , Processos EstocásticosRESUMO
Retinal rod and cone photoreceptors mediate vision in dim and bright light, respectively, by transducing absorbed photons into neural electrical signals. Their phototransduction mechanisms are essentially identical. However, one difference is that, whereas a rod visual pigment remains stable in darkness, a cone pigment has some tendency to dissociate spontaneously into apo-opsin and retinal (the chromophore) without isomerization. This cone-pigment property is long known but has mostly been overlooked. Importantly, because apo-opsin has weak constitutive activity, it triggers transduction to produce electrical noise even in darkness. Currently, the precise dark apo-opsin contents across cone subtypes are mostly unknown, as are their dark activities. We report here a study of goldfish red (L), green (M), and blue (S) cones, finding with microspectrophotometry widely different apo-opsin percentages in darkness, being â¼30% in L cones, â¼3% in M cones, and negligible in S cones. L and M cones also had higher dark apo-opsin noise than holo-pigment thermal isomerization activity. As such, given the most likely low signal amplification at the pigment-to-transducin/phosphodiesterase phototransduction step, especially in L cones, apo-opsin noise may not be easily distinguishable from light responses and thus may affect cone vision near threshold.
Assuntos
Escuridão , Transdução de Sinal Luminoso/fisiologia , Opsinas/metabolismo , Células Fotorreceptoras Retinianas Cones/fisiologia , Animais , Carpa Dourada , Modelos Animais , Técnicas de Patch-Clamp , Estimulação Luminosa/métodos , Células Fotorreceptoras Retinianas Cones/efeitos da radiação , Análise de Célula ÚnicaRESUMO
Numerous rhodopsin mutations have been implicated in night blindness and retinal degeneration, often with unclear etiology. D190N-rhodopsin (D190N-Rho) is a well-known inherited human mutation causing retinitis pigmentosa. Both higher-than-normal spontaneous-isomerization activity and misfolding/mistargeting of the mutant protein have been proposed as causes of the disease, but neither explanation has been thoroughly examined. We replaced wild-type rhodopsin (WT-Rho) in RhoD190N/WT mouse rods with a largely "functionally silenced" rhodopsin mutant to isolate electrical responses triggered by D190N-Rho activity, and found that D190N-Rho at the single-molecule level indeed isomerizes more frequently than WT-Rho by over an order of magnitude. Importantly, however, this higher molecular dark activity does not translate into an overall higher cellular dark noise, owing to diminished D190N-Rho content in the rod outer segment. Separately, we found that much of the degeneration and shortened outer-segment length of RhoD190N/WT mouse rods was not averted by ablating rod transducin in phototransduction-also consistent with D190N-Rho's higher isomerization activity not being the primary cause of disease. Instead, the low pigment content, shortened outer-segment length, and a moderate unfolded protein response implicate protein misfolding as the major pathogenic problem. Finally, D190N-Rho also provided some insight into the mechanism of spontaneous pigment excitation.
Assuntos
Degeneração Retiniana/metabolismo , Rodopsina/metabolismo , Animais , Linhagem Celular , Modelos Animais de Doenças , Células HEK293 , Humanos , Transdução de Sinal Luminoso/fisiologia , Camundongos , Mutação/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Retinose Pigmentar/metabolismo , Segmento Externo da Célula Bastonete/metabolismoRESUMO
Photoreceptors initiate vision by converting photons to electrical activity. The onset of the phototransduction cascade is marked by the isomerization of photopigments upon light capture. We revealed that the onset of phototransduction is accompanied by a rapid (<5 ms), nanometer-scale electromechanical deformation in individual human cone photoreceptors. Characterizing this biophysical phenomenon associated with phototransduction in vivo was enabled by high-speed phase-resolved optical coherence tomography in a line-field configuration that allowed sufficient spatiotemporal resolution to visualize the nanometer/millisecond-scale light-induced shape change in photoreceptors. The deformation was explained as the optical manifestation of electrical activity, caused due to rapid charge displacement following isomerization, resulting in changes of electrical potential and surface tension within the photoreceptor disc membranes. These all-optical recordings of light-induced activity in the human retina constitute an optoretinogram and hold remarkable potential to reveal the biophysical correlates of neural activity in health and disease.
Assuntos
Transdução de Sinal Luminoso , Células Fotorreceptoras Retinianas Cones , Humanos , Transdução de Sinal Luminoso/fisiologia , Retina/fisiologia , Células Fotorreceptoras Retinianas Cones/fisiologia , Tomografia de Coerência Óptica , Visão OcularRESUMO
Rod photoreceptors are composed of a soma and an inner segment (IS) connected to an outer segment (OS) by a thin cilium. OSs are composed of a stack of â¼800 lipid discs surrounded by the plasma membrane where phototransduction takes place. Intracellular calcium plays a major role in phototransduction and is more concentrated in the discs, where it can be incorporated and released. To study calcium dynamics in rods, we used the fluorescent calcium dye CaSiR-1 AM working in the near-infrared (NIR) (excitation at 650 and emission at 664 nm), an advantage over previously used dyes. In this way, we investigated calcium dynamics with an unprecedented accuracy and most importantly in semidark-adapted conditions. We observed light-induced drops in [Ca2+]i with kinetics similar to that of photoresponses recorded electrophysiologically. We show three properties of the rods. First, intracellular calcium and key proteins have concentrations that vary from the OS base to tip. At the OS base, [Ca2+]i is â¼80 nM and increases up to â¼200 nM at the OS tip. Second, there are spontaneous calcium flares in healthy and functional rod OSs; these flares are highly localized and are more pronounced at the OS tip. Third, a bright flash of light at 488 nm induces a drop in [Ca2+]i at the OS base but often a flare at the OS tip. Therefore, rod OSs are not homogenous structures but have a structural and functional gradient, which is a fundamental aspect of transduction in vertebrate photoreceptors.
Assuntos
Cálcio/metabolismo , Transdução de Sinal Luminoso/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Animais , Cálcio/fisiologia , Membrana Celular/metabolismo , Citoplasma/metabolismo , Feminino , Cinética , Masculino , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Segmento Externo da Célula Bastonete/fisiologia , Xenopus laevisRESUMO
Phototropins are photoreceptor proteins that regulate blue light-dependent biological processes for efficient photosynthesis in plants and algae. The proteins consist of a photosensory domain that responds to the ambient light and an output module that triggers cellular responses. The photosensory domain of phototropin from Chlamydomonas reinhardtii contains two conserved LOV (light-oxygen-voltage) domains with flavin chromophores. Blue light triggers the formation of a covalent cysteine-flavin adduct and upregulates the phototropin kinase activity. Little is known about the structural mechanism that leads to kinase activation and how the two LOV domains contribute to this. Here, we investigate the role of the LOV1 domain from C. reinhardtii phototropin by characterizing the structural changes occurring after blue light illumination with nano- to millisecond time-resolved X-ray solution scattering. By structurally fitting the data with atomic models generated by molecular dynamics simulations, we find that adduct formation induces a rearrangement of the hydrogen bond network from the buried chromophore to the protein surface. In particular, the change in conformation and the associated hydrogen bonding of the conserved glutamine 120 induce a global movement of the ß-sheet, ultimately driving a change in the electrostatic potential on the protein surface. On the basis of the change in the electrostatics, we propose a structural model of how LOV1 and LOV2 domains interact and regulate the full-length phototropin from C. reinhardtii. This provides a rationale for how LOV photosensor proteins function and contributes to the optimal design of optogenetic tools based on LOV domains.
Assuntos
Transdução de Sinal Luminoso/fisiologia , Fototropinas/química , Fototropinas/metabolismo , Sítios de Ligação , Chlamydomonas reinhardtii , Luz , Modelos Moleculares , Simulação de Dinâmica Molecular , Fotoquímica , Conformação Proteica , Domínios Proteicos , Espalhamento de Radiação , Difração de Raios XRESUMO
Light and the circadian clock are two essential external and internal cues affecting seedling development. COLD-REGULATED GENE27 (COR27), which is regulated by cold temperatures and light signals, functions as a key regulator of the circadian clock. Here, we report that COR27 acts as a negative regulator of light signaling. COR27 physically interacts with the CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1)-SUPPRESSOR OF PHYTOCHROME A1 (SPA1) E3 ubiquitin ligase complex and undergoes COP1-mediated degradation via the 26S proteasome system in the dark. cor27 mutant seedlings exhibit shorter hypocotyls, while transgenic lines overexpressing COR27 show elongated hypocotyls in the light. In addition, light induces the accumulation of COR27. On one hand, accumulated COR27 interacts with ELONGATED HYPOCOTYL5 (HY5) to repress HY5 DNA binding activity. On the other hand, COR27 associates with the chromatin at the PHYTOCHROME INTERACTING FACTOR4 (PIF4) promoter region and upregulates PIF4 expression in a circadian clock-dependent manner. Together, our findings reveal a mechanistic framework whereby COR27 represses photomorphogenesis in the light and provide insights toward how light and the circadian clock synergistically control hypocotyl growth.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/fisiologia , Relógios Circadianos/fisiologia , Hipocótilo/crescimento & desenvolvimento , Transdução de Sinal Luminoso/fisiologia , Proteínas Repressoras/genética , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Relógios Circadianos/genética , Regulação da Expressão Gênica de Plantas , Hipocótilo/genética , Transdução de Sinal Luminoso/genética , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Repressoras/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Purpose: The primary hypotheses tested are that (1) there exist stimulus-driven intrinsic optical signals in the mouse retina similar to those previously observed in other species, and (2) these optical signals require an intact rod photoreceptor phototransduction cascade. Methods: We used 38 wild-type C57BL6J mice and 18 genetic knockout Gnat1-/- mice to study the light-evoked retinal intrinsic response. A custom mouse fundus camera delivered visual stimuli and collected mouse retinal imaging data of changes in retinal reflectance for further analysis. The retina was stimulated in the high-mesopic range with a 505-nm light-emitting diode while also being illuminated with 780-nm near-infrared light. Results: Wild-type C57BL6J mice yielded retinal imaging signals that typically showed a stimulus-driven decrease in retinal reflectance of â¼0.1%, with a time course of several seconds. The signals exhibit spatial specificity in the retina. Overall, the mouse imaging signals are similar in sign and time course to those reported in other mammalian species but are of lower amplitude. In contrast, functional retinal imaging of Gnat1-/- mice that lack a functional rod transducin yielded no such stimulus-driven signals. Conclusions: Previous studies have not shown which pathway component is essential for the generation of these imaged signals. The absence of the intrinsic signal responses in Gnat1-/- knockout mice indicates that a functional rod transducin is likely to be necessary for generating the retinal intrinsic signals. These studies, to the best of our knowledge, demonstrate for the first time in vivo mouse retinal functional imaging signals similar to those previously shown in other mammalian species.
Assuntos
Transdução de Sinal Luminoso/fisiologia , Estimulação Luminosa , Retina/fisiopatologia , Células Fotorreceptoras Retinianas Bastonetes/fisiologia , Transducina/genética , Animais , Potenciais Evocados Visuais , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Imagem Óptica/métodos , Estimulação Luminosa/instrumentação , Estimulação Luminosa/métodos , Visão Ocular/fisiologiaRESUMO
IMPACT STATEMENT: As the center of phototransduction, retinal photoreceptors are responsible for capturing and converting photon energy to bioelectric signals for following visual information processing in the retina. This article summarizes experimental observation and discusses biophysical mechanism of fast photoreceptor-intrinsic optical signal (IOS) correlated with early phase of phototransduction. Quantitative imaging of fast photoreceptor-IOS may provide objective optoretinography to advance the study and diagnosis of age-related macular degeneration, retinitis pigmentosa, diabetic retinopathy, and other eye diseases that can cause photoreceptor dysfunctions.