LIGHT QUALITY AND PHOTOREACTIVATION OF PLANTS AND VIRUSES

Visible light of different spectral regions was tested for its ability to reverse three effects of ultra-violet radiation, namely, injury (glazing) of French bean leaves, increased resistance of French bean leaves to infection by the Rothamsted tobacco necrosis virus, and inactivation of potato virus X. The different spectral regions were obtained with colorimetric filters and the filtered and unfiltered light from fluorescent tubes; all three effects were reversed only by regions of wave-lengths shorter than 4700 Å. Thirty minutes of illumination at 300–380 f.c. gave substantial photo-reactivation, but irradiated potato virus X did not become affected by visible light until 30 min. after tobacco leaves were inoculated.     

RESULTS OF IRRADIATING SACCHAROMYCES WITH MONOCHROMATIC ULTRA-VIOLET LIGHT : II. THE INFLUENCE OF MODIFYING FACTORS

Possible variation in the probability that absorbed quanta of ultra-violet energy will produce observable inhibitory and lethal effects in the yeast cell, due to non-uniformity in sensitivity of the different regions of the cell, may be further modified by the reproductive stage of the cell at the time of irradiation. Tests of the survival of yeast cells of 15 day and 24 hour cultures indicate that the older resting cells are more resistant to ultra-violet irradiation effects than cells undergoing rapid cell division. The effects of temperature changes within the range of normal growth are evidently small, as judged from the temperature coefficient (1.10). Possible inhibitory effects due to the action of ultra-violet radiation on the malt agar medium and to toxic substances diffused from cells killed by irradiation were not found under the conditions of the experiments. Tests of the validity of the Bunsen-Roscoe reciprocity law for variation in the intensity of the incident ultra-violet radiation up to 30 per cent indicate that for this range the rate of absorption of quanta by the cell does not produce any marked change in the lethal effects observed.     

THEORY AND MEASUREMENT OF VISUAL MECHANISMS : VI. WAVE-LENGTH AND FLASH DURATION IN FLICKER

For spectral regions associated with violet, blue, green, and red the relation between mean critical flash intensity I_m for visual flicker and the flash frequency F is modified as already found with white light when the light time fraction t_L in the flash cycle is changed. For a square image 6.13° on a side, foveally fixated, the "rod" and "cone" contributions to the duplex contour are analyzed in the way already used for white. It is pointed out that several customary qualitative criteria for cone functioning do not necessarily give concordant results. The analysis shows that the three parameters of the probability summations giving the "rod" and "cone" curves are changed independently as a function of wave-length composition of the light, and of the light time fraction. The correlation of these changes, and of those found in the associated variability functions, can be understood in terms of differences in (1) the numbers of neural units potentially excitable and (2) in the numbers of elements of neural effect obtained from them. In a multivariate situation of this kind it is necessary to compare intensities of luminous flux required to activate half the total population of potentially available elements when this total size is held constant for the different conditions. The results of this comparison, for the filtered lights used, are discussed in relation to certain aspects of excitation vs. wave-length. The problem is a general one, arising where the effects produced as a function of a particular variable are concerned. In the distinction between (1) units excited and (2) the actions they produce may be found the clue for the curious fact that with certain wave-lengths the critical intensities are lower than for white. The extension of the observations to other parts of the retina may be expected to further this analysis.     

STUDIES ON BIOLUMINESCENCE : XVII. FLUORESCENCE AND INHIBITION OF LUMINESCENCE IN CTENOPHORES BY ULTRA-VIOLET LIGHT.

1. Small dumps of the luminous cells of Mnemiopsis cannot readily be stimulated mechanically but will luminesce on treatment with saponin solution. Larger groups of luminous cells (such as are connected with two paddle plates) luminesce on mechanical stimulation. This suggests that mechanical stimulation to luminesce occurs chiefly through a nerve mechanism which has been broken up in the small dumps of luminous tissue. 2. The smallest bits of luminous tissue, even cells freed from the animal by agitation, that will pass through filter paper, lose their power to luminesce in daylight and regain it (at least partially) in the dark. 3. Luminescence of the whole animal and of individual cells is suppressed by near ultra-violet light (without visible light). 4. Inhibition in ultra-violet light is not due to stimulation (by the ultra-violet light) of the animal to luminesce, thereby using up the store of photogenic material. 5. Animals stimulated mechanically several times and placed in ultra-violet light show a luminescence along the meridians in the same positions as the luminescence that appears on stimulation. This luminescence in the ultra-violet or "tonic luminescence," is not obtained with light adapted ctenophores and is interpreted to be a fluorescence of the product of oxidation of the photogenic material. 6. Marked fluorescence of the luminous organ of the glowworm (Photuris) and of the luminous slime of Chatopterus may be observed in ultra-violet but no marked fluorescence of the luminous substances of Cypridina is apparent. 7. Evidence is accumulating to show a close relation between fluorescent and chemiluminescent substances in animals, similar to that described for unsaturated silicon compounds and the Grignard reagents.     

QUANTITATIVE STUDIES ON THE PHOTOLETHAL EFFECTS OF QUARTZ ULTRA-VIOLET RADIATION UPON PARAMECIUM

Paramecia grown under controlled conditions were irradiated at known intensities of light of wave-lengths 2537, 2654, 2804, 3025, and 3130 A. The approximate absorption of the light by the Parmecia was found to be greatest and of the same order of magnitude at the three shortest wave-lengths, considerably less at 3025, and least at 3130 A. Paramecia did not die when irradiated with high dosages of intense light of wave-length 3130 A. At the other wave-lengths 50 per cent vesiculation occurred when between 10¹² and 10¹³ quanta had been absorbed by a Paramecium. This would indicate that a very large number of molecules in a Paramecium are affected before vesiculation occurs.     

Effect of calcium channel blockers on the action of various antitumour agents in the yeast Saccharomyces cerevisiae

The cytotoxic effects of a 22 h treatment with four antineoplastic agents in the yeast Saccharomyces cerevisiae ATCC 2366 were investigated. Two agents, doxorubicin and 5-fluorouracil (5-FU), were effective in decreasing the colony-forming ability of yeast cells. Following examination under the light microscope, the effect of doxorubicin appeared to be, at least partially, due to killing of yeast cells whereas the effect of 5-FU was rather due to changes in cell structure leading to abnormal bud formation. For amsacrine (AMSA) and melphalan, cytotoxicity was totally absent. In the presence of diltiazem the above described effects were not significantly changed. When verapamil was added in the culture medium the cytotoxic activity of doxorubicin and 5-FU did not change. However, following treatment with AMSA in combination with verapamil, cell survival was significantly decreased whereas the presence of verapamil increased the yeast survival which was observed after melphalan treatment.     

THE ABSORPTION OF ULTRA-VIOLET RADIATION BY CRYSTALLINE PEPSIN

Determination of the absorption spectra of pure preparations of Northrop''s crystalline pepsin inactivated by irradiation with ultra-violet light shows that the total absorption in the ultra-violet region of the spectrum increases with the degree of inactivation. This increase is especially marked between 2400 and 2750 Å.u. The rate of photoinactivation is shown to be sensitive to changes in pH, increasing with lower values, and evidently bears a one-quantum relationship to the energy flux. Tests of the rate of inactivation of pepsin exposed to several different bands of the ultra-violet spectrum, in relation to the absorbed energy, indicate that the destruction spectrum of the enzyme agrees essentially with its absorption spectrum and is similar to that of urease.     

Effect of cobalt and morphogenic autoregulatory substance (MARS) on morphogenesis ofCandida albicans

Candida albicans yeast cells produce a factor, designated morphogenic autoregulatory substance (MARS), which suppresses yeast-to-mycelial conversion under conditions that support germination. In experiments designed to characterize MARS, the effect of various metals on MARS activity was tested. Cobalt but not zinc, iron, or other metals enhanced the ability of MARS to inhibit germination. Furthermore, uptake of cobalt by cells exposed to MARS was enhanced over cells in control medium if the cells were first incubated in a medium containing MARS for several hours prior to addition of cobalt to the medium. Uptake of zinc under similar conditions was not affected by MARS. The enhanced uptake of cobalt by MARS-exposed cells appears to be related to germination because such enhancement did not occur in strains of yeasts incapable of germination even in the absence of MARS.     

Synchronous Activation of Cell Division by Light or Temperature Stimuli in the Dimorphic Yeast Schizosaccharomyces japonicus

Many fungi respond to light and regulate fungal development and behavior. A blue light-activated complex has been identified in Neurospora crassa as the product of the wc-1 and wc-2 genes. Orthologs of WC-1 and WC-2 have hitherto been found only in filamentous fungi and not in yeast, with the exception of the basidiomycete pathogenic yeast Cryptococcus. Here, we report that the fission yeast Schizosaccharomyces japonicus responds to blue light depending on Wcs1 and Wcs2, orthologs of components of the WC complex. Surprisingly, those of ascomycete S. japonicus are more closely related to those of the basidiomycete. S. japonicus reversibly changes from yeast to hyphae in response to environmental stresses. After incubation at 30°C, a colony of yeast was formed, and then hyphal cells extended from the periphery of the colony. When light cycles were applied, distinct dark- and bright-colored hyphal cell stripes were formed because the growing hyphal cells had synchronously activated cytokinesis. In addition, temperature cycles of 30°C for 12 h and 35°C for 12 h or of 25°C for 12 h and 30°C for 12 h during incubation in the dark induced a response in the hyphal cells similar to that of light. The stripe formation of the temperature cycles was independent of the wcs genes. Both light and temperature, which are daily external cues, have the same effect on growing hyphal cells. A dual sensing mechanism of external cues allows organisms to adapt to daily changes of environmental alteration.     

Three-state denaturation of alpha-lactalbumin by guanidine hydrochloride.

The reversible unfolding of α-lactalbumin by guanidine hydrochloride has been studied at 25.0 °C by means of ultraviolet circular dichroism measurements. The non-coincidence of the apparent transition curves obtained from the ellipticity changes at far (222 nm) and at near (270 nm and 296 nm) ultraviolet wave-lengths demonstrates the presence of at least one intermediate in the denaturation process. The aromatic residues which contribute to the Cotton effects at 270 nm and at 296 nm appear to be exposed to solvent in the first stage of a two-stage process, while the helical regions of the polypeptide chain appear to be destroyed in the second stage. Earlier work has demonstrated an acid transition between two compact forms of α-lactalbumin, a native (neutral pH) form and an acid form. Results presented here suggest that the acid form is produced as an intermediate in the first stage of total unfolding at neutral pH.Lysozyme and α-lactalbumin are known to have similar primary structures and are expected to have similar tertiary structures, but several differences in their properties have been described. The comparison of the unfolding transitions of α-lactalbumin and lysozyme provides a result compatible with similar tertiary structures, although the free energy of stabilization of the native state is 3 to 5 kcal/mol smaller for α-lactalbumin than for lysozyme. The pH dependence of the unfolding reaction can be described in terms of abnormal histidyl and carboxyl residues. The presence of a stable intermediate in the denaturation process may cause a difference in dynamic character in the native state between the two proteins and thus provide a reasonable interpretation for their known differences in chemical reactivity.     

Blue and Green Light-Induced Phototropism in Arabidopsis thaliana and Lactuca sativa L. Seedlings

Exposure time-response curves for blue and green light-induced phototropic bending in hypocotyls of Arabidopsis thaliana (L.) Heynh. and Lactuca sativa L. seedlings are presented. These seedlings show significant phototropic sensitivity up to 540 to 550 nanometers. Since wave-lengths longer than 560 nanometers do not induce phototropic bending, it is suggested that the response to 510 to 550 nanometers light is mediated by the specific blue light photoreceptor of phototropism. We advise care in the use of green `safelights' for studies of phototropism.     

INCREASED IRRITABILITY IN NITELLA DUE TO GUANIDINE

Guanidine applied to Nitella may lower the threshold of E.M.F. required to produce electrical stimulation and may give rise to trains of action currents. Its effect thus appears to be somewhat similar to that observed in animals. Rapid action currents are produced as well as "square topped" action curves and transitional forms. These effects may be due in part to increased protoplasmic conductivity produced by the penetration of guanidine.     

Action Spectra of Photosynthetic Activities of Chlorella ellipsoidea

• 1) Suspensions of Chlorella show an even stronger light scattering than suspensions of chloroplasts of spinach. The bands of absorption are thus broadened and, at higher concentrations, moved to lower wave-lengths. The intensity of the photosynthesis closely follows the curves of light scattering, a fact partly explaining the high efficiency of green light. Calculated per unit thermoelectrically measured incident energy the action spectrum shows bands at 660–670 nm and c. 500 nm and a comparatively high level of the whole region 500–560 nm.
• 2) Flash experiments show the existence of a steady state carotene/xanthophyll that is moved to reduction (c/x > 1) in blue and green light and to oxidation (c/x < 1) in red light. All experiments point to the existence of two light reactions, the first one involving excitation of carotenoids, with ferredoxin-TPN as acceptor, the second one involving excitation of chlorophyll, with the cytochrome system of the chloroplasts acting as donors of electrons and thus completing an energy converting circulation between pigments and enzyme systems.
• 3) The operation of combined light reactions appears also from experiments with simultaneous or succedaneous illumination with monochromatic light of different wave-lengths. Some effects may be explained from separate excitations of carotenoids and chlorophylls, others may depend on still unknown photic reactions.
• 4) The action spectrum in ultrared shows a positive band at c. 900 nm but no or only very small effects in the region 950–1400 nm. Ultrared radiation has on the other hand an enhancing effect on the light excitation in the visible spectrum. A combination of infrared and visible radiation shows a roughly linear relation between incident energy and photosynthetic effect.
• 5) All experiments were performed in the region of linear relation between intensity of incident light and O₂-production. Induced effects of combined monochromatic regions show a very rapid initial change in the steady states that in one or two minutes simmers down to a balanced state of continued photosynthesis. No change was observed in the total quantity of the pigments.

     

Life History Changes in Coral Fluorescence and the Effects of Light Intensity on Larval Physiology and Settlement in Seriatopora hystrix

Fluorescence is common in both coral adult and larval stages, and is produced by fluorescent proteins that absorb higher energy light and emit lower energy light. This study investigated the changes of coral fluorescence in different life history stages and the effects of parental light environment on larval fluorescence, larval endosymbiotic dinoflagellate abundance, larval size and settlement in the brooding coral Seriatopora hystrix. Data showed that coral fluorescence changed during development from green in larvae to cyan in adult colonies. In larvae, two green fluorescent proteins (GFPs) co-occur where the peak emission of one GFP overlaps with the peak excitation of the second GFP allowing the potential for energy transfer. Coral larvae showed great variation in GFP fluorescence, dinoflagellate abundance, and size. There was no obvious relationship between green fluorescence intensity and dinoflagellate abundance, green fluorescence intensity and larval size, or dinoflagellate abundance and larval size. Larvae of parents from high and low light treatments showed similar green fluorescence intensity, yet small but significant differences in size, dinoflagellate abundance, and settlement. The large variation in larval physiology combined with subtle effects of parental environment on larval characteristics seem to indicate that even though adult corals produce larvae with a wide range of physiological capacities, these larvae can still show small preferences for settling in similar habitats as their parents. These data highlight the importance of environmental conditions at the onset of life history and parent colony effects on coral larvae.     

Mechanism of the Synergistic Inactivation of Escherichia coli by UV-C Light at Mild Temperatures

UV light only penetrates liquid food surfaces to a very short depth, thereby limiting its industrial application in food pasteurization. One promising alternative is the combination of UV light with mild heat (UV-H), which has been demonstrated to produce a synergistic bactericidal effect. The aim of this article is to elucidate the mechanism of synergistic cellular inactivation resulting from the simultaneous application of UV light and heat. The lethality of UV-H treatments remained constant below ∼45°C, while lethality increased exponentially as the temperature increased. The percentage of synergism reached a maximum (40.3%) at 55°C. Neither the flow regimen nor changes in the dose delivered by UV lamps contributed to the observed synergism. UV-H inactivation curves of the parental Escherichia coli strain obtained in a caffeic acid selective recovery medium followed a similar profile to those obtained with uvrA mutant cells in a nonselective medium. Thermal fluidification of membranes and synergistic lethal effects started around 40 to 45°C. Chemical membrane fluidification with benzyl alcohol decreased the UV resistance of the parental strain but not that of the uvrA mutant. These results suggest that the synergistic lethal effect of UV-H treatments is due to the inhibition of DNA excision repair resulting from the membrane fluidification caused by simultaneous heating.     

Translational regulation of protein synthesis, in response to light, at a critical stage of Volvox development

In Volvox cultures synchronized by a light-dark cycle, juveniles containing presumptive somatic and reproductive cells are produced during the dark, but their cells do not differentiate until after the lights come on. The pattern of protein synthesis changes rapidly after the lights come on. Action spectra and effects of photosynthesis inhibitors indicate that this protein synthetic change is not simply a consequence of renewed flow of energy from illuminated chloroplasts. Actinomycin, at a level adequate to block the response to heat shock, has virtually no effect on the response of the same cells to light; furthermore, RNAs isolated from unilluminated and illuminated juveniles yield indistinguishable in vitro translation products. We conclude, therefore, that this effect of light is exerted almost exclusively at the translational level, generating one of the most striking examples of translational regulation yet described.     

SOME EFFECTS OF ULTRA-VIOLET RADIATION ON LEAVES OF FRENCH BEAN (PHASEOLUS VULGARIS L.)

Exposing the primary leaves of French bean to ultra-violet light produces a variety of effects, their magnitude depending on the extent of the exposure and the subsequent treatment of the leaves. Effects include some that are externally obvious, such as glazing and bronzing; others are detectable by changes in susceptibility to virus infection, by increased sensitivity to damage by ultra-violet, and by an increased tendency for cells to collapse when the leaves are kept in darkness. Some of the effects can be counteracted by exposing leaves to visible light. Effects of radiation are not confined to epidermal cells, and measurements with detached epidermis indicate that from a quarter to a half of incident radiation of wave-length 2536 A. may be transmitted by the epidermis.     

Control of Flowering of Xanthium pensylvanicum by Red and Far-red Light

A study was made of the effects of various durations, intensities and combinations of red and far-red light interruptions on the flowering responses of Xanthium pensylvanicum Wallr. A dual response to treatments of far-red light was observed. In short dark periods, far-red light alone did not greatly affect flowering but was able to overcome the inhibition of flowering caused by red light. In dark periods longer than 15 hours, far-red inhibited flowering and added to rather than overcame the inhibition by red light. The dark period length required for far-red inhibition remained the same whether far-red was given at the start or at the eighth hour of darkness.

In 48-hour dark periods Xanthium showed 3 responses to additions of red and far-red light breaks: A) response to red light; B) response to far-red light; and C) response to red followed by far-red light. Red light given any time in the first 30 hours of darkness overcame the inhibitory effect of far-red light given at either the start or the eighth hour of darkness. Red light given later than the thirtieth hour did not overcome the far-red effect.

Approximately the same energy of red light was required to overcome the inhibitory effect of far-red at the second hour of darkness as was required to produce maximum red light inhibition at the eighth hour. Although far-red light was most inhibitory when given early in a long dark period, approximately the same energy of far-red light was required to saturate the far-red response at the fourth, eighth and sixteenth hours.

The results are discussed in relation to other reports of far-red inhibition of flowering in short-day plants.

     

Alteration of the largest subunit of RNA polymerase II and its effect on chromosome stability in Schizosaccharomyces pombe

A mutation in the RNA polymerase II largest subunit (RpII LS) that is related to abnormal induction of sister chromatid exchange has previously been described the CHO-K1 cell mutant tsTM4. To elucidate the molecular basis of this effect we introduced the mutation into the homologous site in the Schizosaccharomyces pombe rpb1 gene, which encodes RpII LS. Since the tsTM4 mutant exhibited a decrease in the rate of DNA synthesis in cells arrested in S phase at the nonpermissive temperature, we focussed on the study of growth, the cell cycle, and chromosome stability at various temperatures. First, we examined the effects of the mutation on haploid yeast cells. The mutant showed slower growth than the wild type, but cell growth was not arrested at the nonpermissive temperature. When growing cells were shifted to the nonpermissive temperature, an accumulation of cells in G1 and/or G0 was observed. Tetrad analysis suggested that these phenotypes were associated with the mutation. In diploid cells, chromosome instability was detected by loss of intragenic complementation between two alleles of the ade6 gene. An abnormal fraction of cells containing an intermediate DNA content was also observed by FACS analysis. The accumulation of this fraction may reflect the fact that a large number of cells are in S phase or have an abnormal DNA content as a result of chromosome instability. These observations demonstrate that the S. pomberpb1 mutant exhibits a phenotype very similar to that of the CHO-K1 cell mutant tsTM4.