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Additional evidence for the cyclic GMP signaling pathway resulting in the photophobic behavior of Stentor coeruleus.

We report that exo- and endogenous guanosine 3',5'-cyclic monophosphate (cGMP) specifically influenced the photophobic response. In behavioral experiments the slowly hydrolyzable and membrane-permeable analogs of cGMP (8-bromo-cGMP [Br-cGMP] and N6,2'-o-dibutyryl-cGMP) dramatically prolonged the time for ciliary stop response and decreased the duration of ciliary reversal in a dose-dependent manner. When analogs of adenosine 3',5'-cyclic monophosphate (cAMP) (8-bromo-cAMP or N6,2'-o-dibutyryl-cAMP) were used, no essential effects were detected on the kinetics of the photophobic response. Both nonspecific cyclic nucleotide phosphodiesterase (PDE) activity inhibitors (3-isobutyl-1-methylxanthine [IBMX] and 1,3-dimethylxanthine [theophylline]) and the highly specific cGMP-PDE activity inhibitor 1,4-dihydro-5-[2-propoxyphenyl]-7H-1,2,3-triazolo[4,5-d]pyrimidine-7-one (zaprinast) mimicked the effects of cGMP analogs. Treatment of cells with an inhibitor of guanylate cyclase activity (6-anilino-5,8-quinolinedione [LY 83583]) exerted an effect opposite to that of cGMP analogs and PDE activity inhibitors. The positive physiological effect of LY 83583 was significantly diminished in ciliates that were treated simultaneously with Br-cGMP. In an assay of cell cyclic nucleotide content, the exposure of dark-adapted Stentor to light evoked a transient decrease in the basal level of intracellular cGMP. Alterations in internal cGMP levels were more distinct when the intensity of applied illumination was increased. In the presence of IBMX or theophylline the basal content of cGMP was markedly enhanced, and the photoinduced changes in cGMP level were less pronounced. In this paper the possible whole molecular mechanism by which the ciliary orientation in Stentor is controlled by light is presented.

Animals↗

Dictyostelium RasD is required for normal phototaxis, but not differentiation.

RasD, a Dictyostelium homolog of mammalian Ras, is maximally expressed during the multicellular stage of development. Normal Dictyostelium aggregates are phototactic and thermotactic, moving towards sources of light and heat with great sensitivity. We show that disruption of the gene for rasD causes a near-total loss of phototaxis and thermotaxis in mutant aggregates, without obvious effects on undirected movement. Previous experiments had suggested important roles for RasD in development and cell-type determination. Surprisingly, rasD(-) cells show no obvious changes in these processes. These cells represent a novel class of phototaxis mutant, and indicate a role for a Ras pathway in the connections between stimuli and coordinated cell movement.

Animals↗

Journey towards the centre of the earth: plant gravitropism.

The study of gravitropism dates back about two hundred years, and although significant progress has been made in this field, the central question in gravitropism research is still unanswered. How is a physical stimulus transduced into a biochemical signal which ultimately elicits a response (curvature) in a plant? Simply put, how does a plant 'know' which way is down?

Eukaryota↗

Serum response factor: discovery, biochemistry, biological roles and implications for tissue injury healing.

Serum response factor (SRF) is a transcription factor, which binds to a serum response element (SRE) associated with a variety of genes including immediate early genes such as c-fos, fosB, junB, egr-1 and -2, neuronal genes such as nurr1 and nur77 and muscle genes such as actins and myosins. By regulating expression of these genes, SRF controls cell growth and differentiation, neuronal transmission as well as muscle development and function. SRF can be activated by a variety of agents, including serum, lysophosphatidic acid (LPA), lipopolysaccharide (LPS), 12-O-tetradecanoylphorbol-13-acetate (TPA), cytokines, tumor necrosis factor-alpha (TNFalpha), agents that increase intracellular Ca2+, T-cell virus1 activator protein, hepatitis B virus activator proteins pX, activated oncogenes and protooncogenes as well as extracellular stimuli such as antioxidant and UV light. SRF itself is regulated by both cellular signal transduction pathways and interaction with other transcription factors e.g. Sp1, ATF6 and myogenic regulatory factors. Its biological function is best elucidated for myocardium. Specific cardiac SRF transgenesis demonstrated that overexpression of SRF caused hypertrophic cardiomyopathy in mouse and the mouse died of heart failure within 6 months after birth. Other transgenic data suggested that sufficient SRF was needed for embryogenesis and early development. Since SRF is important regulator of numerous genes involved in cell growth and differentiation, including muscle and neural components, SRF may also play a crucial role in tissue injury and ulcer healing, e.g. healing of gastrointestinal ulcers.

Animals↗

Nonlinearity and facilitation in phosphoinositide signaling studied by the use of caged inositol trisphosphate in Xenopus oocytes.

The phosphoinositide signaling pathway, which mediates neurotransmitter responses, was studied in Xenopus oocytes by recording membrane currents evoked using lightflash photolysis of caged inositol trisphosphate (caged IP3) to produce rapid and reproducible transients in intracellular IP3 levels. Photolysis of caged IP3 evoked currents which were carried largely by chloride ions and depended upon intracellular, but not extracellular, calcium. A given light flash evoked larger responses when the amount of caged IP3 loaded into the oocyte was increased, and illumination of the vegetal hemisphere gave larger responses than the animal. Long (10 sec) light exposures produced oscillatory currents, resembling responses to serotonin and other agonists, which became larger, more transient, and of shorter latency as the light intensity was increased. Brief (ca. 100 msec) flashes evoked a single "spike" of current. The caged IP3 response showed a threshold, in that light flashes had to be greater than a certain intensity and duration before currents could be detected. Associated with this, sub- and suprathreshold light flashes caused a long-lasting (seconds or minutes) potentiation of responses to subsequent test flashes. The lightflash response was also potentiated by a preceding intracellular injection of IP3 and by extracellular application of an agonist thought to induce IP3 liberation. However, intracellular injections of calcium depressed the response. We conclude that the liberation of calcium from intracellular stores varies nonlinearly with the intracellular level of IP3. This phenomenon may explain earlier observations, including the long latency of currents evoked by low doses of agonists such as acetylcholine and serotonin, and the nonlinear facilitation seen between these agonists. Further, it suggests a mechanism for "chemical integration," which may be important in the functioning of neurons and other cells which use IP3 as an intracellular messenger.

Animals↗

Poly ADP ribosylation as a possible mechanism of microwave--biointeraction.

Electromagnetic fields (EMFs) affect the metabolism of the body including the nervous, endocrine, cardiovascular, hematological as well as the reproductive system. EMFs are environmental pollutants, thus posing a health hazard which can cause steric changes in the molecule located at the cell surface. Microwaves are known to cause chromosomal abberations and act as tumor promoters. The process involves a stream of signals from cell membrane to nucleus and other organelles. The present investigations aim to understand the mechanism of biological effects of microwaves (2.45 GHz). The effect was studied on poly ADP-ribosylation, which is a post translational modification of chromatin protein catalysed by the enzyme poly ADPR polymerase using NAD+ as the substrate. Poly ADP-ribosylation has been shown to be involved in several aspects of chromatin structure and function. Twenty-three days old rats weighing 42-48 gms were exposed at a microwave dose level of 1.0 mW/cm2. After exposure for sixty days the animals were sacrificed and an estimation of poly ADPR polymerase activity was undertaken in different organs of these animals. There was an increase of 20% in its activity in liver, 35% in testis, whereas brain showed a 53% decrease in diencephalon and 20% decrease in the cortex in the exposed animals as compared to their respective controls. There was no change in enzyme activity in spleen and kidney. This was accompanied by concomitant changes in NAD+ levels. The above results may be cited as important events in carcinogenesis and tumor promotion related to microwave exposure and the signal transduction mechanism involved. The goal is to shed light on complex ecogenetic interactions leading to cancer modulation of gene expression by epigenetic mechanism.

Animals↗

Ca2+ signaling in the pancreatic acinus.

Change in cytosolic free calcium ion concentration [Ca2+]c, resulting from receptor activation by an appropriate agonist, functions as a cardinal intracellular signaling in the stimulus-secretion coupling in a wide variety of secretory cells including the acini of the pancreas. Ratiometric imaging of [Ca2+]c dynamics by UV-laser scanning confocal microscopy led us to conclude that in the cholecystokinin (CCK)-8-induced recurrent [Ca2+]c, spiking increases initially in the basolateral margin of the acinus and propagates to the luminal margin. [Ca2+]c in this initial cell increased rapidly and uniformly to the maximum level. The decrease in [Ca2+]c in the initial cell coincided with a small increase in [Ca2+]c in the luminal regions of the bilateral neighboring cells followed by uniform maximal increase in [Ca2+]c in these neighboring cells. A series of [Ca2+]c dynamics was repeated to form recurrent Ca2+ spiking. The temporal sequences of [Ca2+]c dynamics recorded during continuous stimulation with CCK-8 at a physiologic concentration in individual acinar cells forming the acinus were displayed on the identical time scale. The figure indicates that the signaling is not synchronous in cells forming an acinus. From these and other results, we proposed a model in which CCK-8 at a low physiologic concentration binds to highly sensitive CCK receptor interacting with heterotrimeric guanosine 5'-triphosphate-binding proteins of the Gq class, generate Ins 1,4,5-P3, and recurrent [Ca2+]c, spiking. The recurrent Ca2+ spiking maintains a sustained secretory response, recurrent exocytosis of zymogen granules, and concomitant secretion of isotonic NaCl.

Animals↗

Cytoskeletal assembly and ATP release regulate astrocytic calcium signaling.

We have studied the role of actin fiber assembly on calcium signaling in astrocytes. We found that (1) after astrocytes have been placed in culture, it takes several hours for organization of the definitive actin cytoskeleton. Actin organization and the number of cells engaged in calcium signaling increased in parallel. (2) Disruption of the actin cytoskeleton attenuated the calcium wave propagation; cytochalasin D treatment reduced the number of astrocytes engaged in calcium signaling. (3) Propagation of calcium waves depends on cytoskeletal function; inhibition of myosin light chain kinase suppressed wave activity. (4) Astrocytic calcium signaling is mediated by release of ATP and purinergic receptor stimulation, because agents that interfere with this cascade attenuated or reduced calcium signaling. Because purinergic receptors are fully functional shortly after plating and not affected by cytochalasin D, these observations indicate that cytoskeleton organization is a prerequisite for interastrocytic calcium signaling mediated by release of ATP.

Actins↗

Vascular endothelial growth factor (VEGF) and its receptors.

Vascular endothelial growth factor (VEGF) is a highly specific mitogen for vascular endothelial cells. Five VEGF isoforms are generated as a result of alternative splicing from a single VEGF gene. These isoforms differ in their molecular mass and in biological properties such as their ability to bind to cell-surface heparan-sulfate proteoglycans. The expression of VEGF is potentiated in response to hypoxia, by activated oncogenes, and by a variety of cytokines. VEGF induces endothelial cell proliferation, promotes cell migration, and inhibits apoptosis. In vivo VEGF induces angiogenesis as well as permeabilization of blood vessels, and plays a central role in the regulation of vasculogenesis. Deregulated VEGF expression contributes to the development of solid tumors by promoting tumor angiogenesis and to the etiology of several additional diseases that are characterized by abnormal angiogenesis. Consequently, inhibition of VEGF signaling abrogates the development of a wide variety of tumors. The various VEGF forms bind to two tyrosine-kinase receptors, VEGFR-1 (flt-1) and VEGFR-2 (KDR/flk-1), which are expressed almost exclusively in endothelial cells. Endothelial cells express in addition the neuropilin-1 and neuropilin-2 coreceptors, which bind selectively to the 165 amino acid form of VEGF (VEGF165). This review focuses on recent developments that have widened considerably our understanding of the mechanisms that control VEGF production and VEGF signal transduction and on recent studies that have shed light on the mechanisms by which VEGF regulates angiogenesis.

Alternative Splicing↗

[Mechanism of signal transduction in photoreceptors of vertebrates: identification of an intracellular transmitter].

A short account is given of the studies which resulted in identification of cGMP as intracellular transmitter which realizes the control of light-depending channels of vertebrate photoreceptors. The following problems are discussed. 1. Proofs of necessary participation of intracellular mediator in the signal transduction in the photoreception act. 2. Competition between cGMP and Ca2+ ions for the role of such mediator in the photoreceptors of vertebrates. Relationship between metabolism of cGMP and Ca2+ which makes impossible identification of the mediator in the in vivo experiments. 3. Patch-clamp-experiments which have shown that conductivity of the rod plasma membrane is controlled by cGMP rather than by Ca2+ ions. Proofs of the identity of cGMP-regulated channels recorded in patch-clamp-experiments and light-dependent channels observed in the in vivo experiments. 4. Analysis of the prospects of further studies of the control mechanism of light-dependent channels.

Animals↗

Stimulation of reactive oxygen species production by an antidepressant visible light source.

BACKGROUND: The mechanism by which visible light stimulates chronobiological phase-shifting or antidepressant effects in humans is unknown. METHODS: Normal human NIH/3T3 nonpigmented fibroblasts were irradiated with a visible light source (SunRay) used in the treatment of winter seasonal depression. Electron spin resonance was assessed before and after 10 min of illumination at 2 mW/cm(2) (illuminance of 3700 lux), with and without the presence of 5 microL of 0.0214 mg/mL vitamin C. RESULTS: The fibroblasts showed evidence of production of reactive oxygen species after 10 min of irradiation. CONCLUSIONS: These in vitro data establish that an antidepressant source of visible light is capable of inducing the production of reactive oxygen species in skin. Such species may participate in signal transduction pathways leading to mood changes.

Cell Line↗

DELLAs contribute to plant photomorphogenesis.

Plant morphogenesis is profoundly influenced by light (a phenomenon known as photomorphogenesis). For example, light inhibits seedling hypocotyl growth via activation of phytochromes and additional photoreceptors. Subsequently, information is transmitted through photoreceptor-linked signal transduction pathways and used (via previously unknown mechanisms) to control hypocotyl growth. Here we show that light inhibition of Arabidopsis (Arabidopsis thaliana) hypocotyl growth is in part dependent on the DELLAs (a family of nuclear growth-restraining proteins that mediate the effect of the phytohormone gibberellin [GA] on growth). We show that light inhibition of growth is reduced in DELLA-deficient mutant hypocotyls. We also show that light activation of phytochromes promotes the accumulation of DELLAs. A green fluorescent protein (GFP)-tagged DELLA (GFP-RGA) accumulates in elongating cells of light-grown, but not dark-grown, transgenic wild-type hypocotyls. Furthermore, transfer of seedlings from light to dark (or vice versa) results in rapid changes in hypocotyl GFP-RGA accumulation, changes that are paralleled by rapid alterations in the abundance in hypocotyls of transcripts encoding enzymes of GA metabolism. These observations suggest that light-dependent changes in hypocotyl GFP-RGA accumulation are a consequence of light-dependent changes in bioactive GA level. Finally, we show that GFP accumulation and quantitative modulation of hypocotyl growth is proportionate with light energy dose (the product of exposure duration and fluence rate). Hence, DELLAs inhibit hypocotyl growth during the light phase of the day-night cycle via a mechanism that is quantitatively responsive to natural light variability. We conclude that DELLAs are a major component of the adaptively significant mechanism via which light regulates plant growth during photomorphogenesis.

Arabidopsis↗

A new type of mutation in phytochrome A causes enhanced light sensitivity and alters the degradation and subcellular partitioning of the photoreceptor.

A specific light program consisting of multiple treatments with alternating red and far-red light pulses was used to isolate mutants in phytochrome A-dependent signal transduction pathways in Arabidopsis. Because of their phenotype, the mutants were called eid for empfindlicher im dunkelroten Licht, which means hypersensitive in far-red light. One of the isolated mutants, eid4, is a novel semi-dominant allele of the phytochrome A gene that carries a missense mutation in the chromophore-binding domain. The mutation did not change the photochemical properties of the photoreceptor, but it leads to an increased stability under light conditions that induce its rapid degradation. Fusion proteins with the green fluorescent protein exhibited clear alterations in subcellular localization of the mutated photoreceptor: The fusion protein was impaired in the formation of sequestered areas of phytochrome in the cytosol, which can explain its reduced light-dependent degradation. In contrast, the mutation stabilizes nuclear speckles (NUS) that appear late under continuous far-red light, whereas the formation of early, transiently appearing NUS remained more or less unaltered.

Amino Acid Sequence↗

The evolution and function of blue and red light photoreceptors.

Photoreceptors allow living organisms to optimize perception of light in the natural environment and thus to gain information about their external world. In this review, we describe blue and red light photoreceptors in bacteria, plants, and animals in relation to their evolution. Analyses performed in different organisms have revealed wonderful examples of structural modifications of the light-sensing proteins themselves, as well as diversification of the signal transduction pathways they use in relation with their evolutionary history and function. In different organisms, the same photoreceptor may have a very conserved role (convergent evolution of function) or may modulate different responses (acquisition of new function). Multiple photoreceptors of the same family in the same organism indicate gene duplication events during evolution, with a consequent enhanced sensitivity to variations in ambient light. Conversely, two different photoreceptors may be involved in the control of the same physiological response. Genomic analysis in marine diatoms, combined with phylogenetic studies, has also revealed the presence of blue and red light photoreceptors in the marine environment. This discovery has intriguing implications for the understanding of light perception and its evolution in photosynthetic organisms. In addition, the characterization of these photoreceptors likely will add to our understanding of photoreceptor diversity as an adaptation to different habitats.

Animals↗