Contribution of hydroxycinnamates and flavonoids to epidermal shielding of UV‐A and UV‐B radiation in developing rye primary leaves as assessed by ultraviolet‐induced chlorophyll fluorescence measurements

Epidermally located ultraviolet (UV)‐absorbing phenolic compounds, flavonoids and hydroxycinnamic acid esters (HCAs), can shield the underlying tissues in plants against harmful UV‐radiation. The relative importance of the two different classes of phenolic compounds for UV‐screening was a matter of recent debate. Using a non‐invasive method based on chlorophyll fluorescence measurements to estimate epidermal UV transmittance, the relationship between epidermal UV shielding and the content of the two different groups of secondary phenolic compounds in the epidermal layers and the underlying photosynthetic mesophyll of developing rye primary leaves grown under supplementary UV‐B radiation was investigated. From the fourth to the tenth day after sowing, epidermally located flavonoids increased in an age‐ and irradiation‐dependent manner, whereas mesophyll flavonoids and epidermal HCAs, mainly ferulic acid and p‐coumaric acid esters, were constitutively present and did not vary in their contents over the observed time period. There was an excellent correlation between epidermal UV‐A and UV‐B absorbances as assessed by chlorophyll fluorescence measurements and contents of epidermal flavonoids. However, HCAs showed an additional contribution to UV‐B shielding. In contrast, mesophyll flavonoids did not seem to play a respective role. When absorbances of the abaxial and adaxial epidermal layers were compared, it became apparent that in fully expanded primary leaves epidermal tissues from both sides were equally effective in absorption of UV‐radiation. However, the earlier and more UV‐exposed abaxial epidermis of young unrolling leaves showed a significantly higher absorption. It is shown that in early stages of development the epidermal HCAs are the dominant UV‐B protective compounds of the primary leaf. This function is increasingly replaced by the epidermal flavonoids during leaf development and acclimation. The application of chlorophyll fluorescence measurements has been proven to be a useful tool for estimating relative contents of these compounds in epidermal tissue.     

Interaction of COP1 and UVR8 regulates UV‐B‐induced photomorphogenesis and stress acclimation in Arabidopsis

The ultraviolet‐B (UV‐B) portion of the solar radiation functions as an environmental signal for which plants have evolved specific and sensitive UV‐B perception systems. The UV‐B‐specific UV RESPONSE LOCUS 8 (UVR8) and the multifunctional E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) are key regulators of the UV‐B response. We show here that uvr8‐null mutants are deficient in UV‐B‐induced photomorphogenesis and hypersensitive to UV‐B stress, whereas overexpression of UVR8 results in enhanced UV‐B photomorphogenesis, acclimation and tolerance to UV‐B stress. By using sun simulators, we provide evidence at the physiological level that UV‐B acclimation mediated by the UV‐B‐specific photoregulatory pathway is indeed required for survival in sunlight. At the molecular level, we demonstrate that the wild type but not the mutant UVR8 and COP1 proteins directly interact in a UV‐B‐dependent, rapid manner in planta. These data collectively suggest that UV‐B‐specific interaction of COP1 and UVR8 in the nucleus is a very early step in signalling and responsible for the plant's coordinated response to UV‐B ensuring UV‐B acclimation and protection in the natural environment.     

UV‐A/BLUE LIGHT–INDUCED REACTIVATION OF SPORE GERMINATION IN UV‐B IRRADIATED ULVA PERTUSA (CHLOROPHYTA)1

Recent reduction in the ozone shield due to manufactured chlorofluorocarbons raised considerable interest in the ecological and physiological consequences of UV‐B radiation (λ=280–315 nm) in macroalgae. However, early life stages of macroalgae have received little attention in regard to their UV‐B sensitivity and UV‐B defensive mechanisms. Germination of UV‐B irradiated spores of the intertidal green alga Ulva pertusa Kjellman was significantly lower than in unexposed controls, and the degree of reduction correlated with the UV doses. After exposure to moderate levels of UV‐B irradiation, subsequent exposure to visible light caused differential germination in an irradiance‐ and wavelength‐dependent manner. Significantly higher germination was found at higher photon irradiances and in blue light compared with white and red light. The action spectrum for photoreactivation of germination in UV‐B irradiated U. pertusa spores shows a major peak at 435 nm with a smaller but significant peak at 385 nm. When exposed to December sunlight, the germination percentage of U. pertusa spores exposed to 1 h of solar radiation reached 100% regardless of the irradiation treatment conditions. After a 2‐h exposure to sunlight, however, there was complete inhibition of germination in PAR+UV‐A+UV‐B in contrast to 100% germination in PAR or PAR+UV‐A. In addition to mat‐forming characteristics that would act as a selective UV‐B filter for settled spores under the parental canopy, light‐driven repair of germination after UV‐B exposure could explain successful continuation of U. pertusa spore germination in intertidal settings possibly affected by intense solar UV‐B radiation.     

UV‐B INDUCTION OF UV‐B PROTECTION IN ULVA PERTUSA (CHLOROPHYTA)1

The green macroalga Ulva pertusa Kjellman produced UV‐B absorbing compounds with a prominent absorption maximum at 294 nm in response only to UV‐B, and the amounts induced were proportional to the UV‐B doses. Under a 12:12‐h light:dark regime, the production of UV‐absorbing compounds occurred only during the exposure periods with little turnover in the dark. There was significant reduction in growth in parallel with the production of UV‐B absorbing compounds. The polychromatic action spectrum for the induction of UV‐B absorbing compounds in U. pertusa exhibits a major peak at 292 nm with a smaller peak at 311.5 nm. No significant induction was detected above 354.5 nm, and radiation below 285 nm caused significant reduction in the levels of UV‐B absorbing compounds. After UV‐B irradiation at 1.0 W·m^?2 for 9 h, the optimal photosynthetic quantum yield of the samples with UV‐B absorbing compounds slightly increased relative to the initial value, whereas that of thalli lacking the compounds declined to 30%–34% of the initial followed by subsequent recovery in dim light of up to 84%–85% of the initial value. There was a positive and significant relationship between the amount of UV‐B absorbing compounds with antioxidant activity as determined by the α,α‐diphenyl‐β‐picrylhydrazyl scavenging assay. In addition to mat‐forming characteristics and light‐driven photorepair, the existence and antioxidant capacity of UV‐B absorbing compounds may confer U. pertusa a greater selective advantage over other macroalgae, thereby enabling them to thrive in the presence of intense UV‐B radiation.     

Aphid orientation and performance in glasshouses under different UV‐A/UV‐B radiation regimes

Visual cues leading to host selection and landing are of major importance for aphids and evidence suggests that flight activity is very dependent on ultraviolet (UV)‐A radiation in the environment. At the same time research on insect plant hosts suggest that the UV‐B component can deter some pests via changes in secondary metabolite chemistry. Here, we examine the potential of UV (UV‐A/UV‐B) radiation to control insect pests in the glasshouse environment. We first examined artificial exposure to UV‐B and the potential to trigger morphological and biochemical modifications in pepper (Capsicum annuum L., Solanaceae) with implications for the fitness of green peach aphid, Myzus persicae Sulzer (Hemiptera: Aphididae). UV‐B caused accumulation of leaf secondary metabolites and soluble carbohydrates, and stimulated photosynthetic pigments. However, UV‐B did not impact on foliar protein content and aphid performance was unaffected. Next, we studied how altering the UV‐A/UV‐B ratio environment affected aphid orientation and spatial distribution over time, either directly or by exposing plants to supplemental UV before insect introduction. Aphids directly settled and dispersed on their host pepper plants more readily in the presence of supplemental UV‐A and UV‐B. In the control treatment with ambient glasshouse UV‐A and UV‐B, insects remained more aggregated. Furthermore, insects were less attracted to peppers pre‐exposed to supplemental UV‐A and UV‐B radiation. Our results suggest that suppression of UV‐A and UV‐B inside the protected environment reduces aphid colonization and dispersal. Furthermore, application of moderate exposure of young pepper plants to supplemental UV‐B radiation could aid in protection from the colonization by phytophagous insects.     

Sub‐lethal UV‐C radiation induces callose,hydrogen peroxide and defence‐related gene expression in Arabidopsis thaliana

Exposure of plants to UV‐C irradiation induces gene expression and cellular responses that are commonly associated with wounding and pathogen defence, and in some cases can lead to increased resistance against pathogen infection. We examined, at a physiological, molecular and biochemical level, the effects of and responses to, sub‐lethal UV‐C exposure on Arabidopsis plants when irradiated with increasing dosages of UV‐C radiation. Following UV‐C exposure plants had reduced leaf areas over time, with the severity of reduction increasing with dosage. Severe morphological changes that included leaf glazing, bronzing and curling were found to occur in plants treated with the 1000 J·m^?2 dosage. Extensive damage to the mesophyll was observed, and cell death occurred in both a dosage‐ and time‐dependent manner. Analysis of H₂O₂ activity and the pathogen defence marker genes PR1 and PDF1.2 demonstrated induction of these defence‐related responses at each UV‐C dosage tested. Interestingly, in response to UV‐C irradiation the production of callose (β‐1,3‐glucan) was identified at all dosages examined. Together, these results show plant responses to UV‐C irradiation at much lower doses than have previously been reported, and that there is potential for the use of UV‐C as an inducer of plant defence.     

EFFECTS OF INCREASING UV‐B RADIATION DUE TO OZONE DEPLETION ON PHOTOSYNTHESIS OF ANTARCTIC CYANOBACTERIA

Ground level ultraviolet‐B (UV‐B; 290–320 nm) fluxes in Antarctica have been increasing due to stratospheric ozone depletion. Although mat‐forming cyanobacteria are major component of freshwater algal biomass in Antarctica, little is known about their response to increasing ultraviolet radiation (UVR). The present study evaluated the sensitivity to UVR of two strains of mat‐forming cyanobacteria with different cell size, Phormidium murrayi (6.0 x 3.2 μm) and Schizothrix calcicola (2.2 x 2.3 μm). Cyanobacterial photosynthesis was measured under different UV spectral quality and quantity achieved by polychromatic filters with different cutoff wavelengths and neutral density screens. The productivity and irradiance data were used to generate biological weighting functions (BWF) for the assessment of UV inhibition on photosynthesis. The kinetics of UV inhibition, as determined by PAM fluorometry, differed between the two species so that inhibition of P. murrayi and S. calcicola were modeled based on UV‐irradiance and cumulative exposure, respectively. After a one hour exposure, BWF's did not differ between the two isolates of cyanobacteria despite their differences in cell size. To evaluate the negative impact of increased UV‐B exposure due to ozone depletion on cyanobacteria, the BWF's were applied to two solar spectra obtained from McMurdo Station, one on a day when the ozone hole was prominent (O₃ = 170 Dobson units; DU = 10‐3 cm O₃), and the other on a day with high ozone concentration (O₃ = 328 DU). The decrease in ozone level would reduce productivity by 3–8%. Seasonal variation of UVR has a bigger impact on cyanobacterial productivity than ozone depletion.     

PHOTOSYNTHETIC ADAPTATION OF A BLOOM‐FORMING CYANOBACTERIUM MICROCYSTIS AERUGINOSA (CYANOPHYCEAE) TO PROLONGED UV‐B EXPOSURE1

The bloom‐forming cyanobacterium Microcystis aeruginosa Kütz 854 was cultured with 1.05 W·m^?2 UV‐B for 3 h every day, and its growth, pigments, and photosynthesis were investigated. The specific growth rates represented by chl a concentration and OD₇₅₀ were inhibited 8% and 9% by UV‐B exposure, respectively. Six days of UV‐B treatment significantly reduced cellular contents of phycocyanin and allophycocyanin by 32% and 62%, respectively, and markedly increased the carotenoid content by 27%, but had little effect on the chl a content. The initial values of optimal photosynthetic efficiency for UV‐B treated samples were, respectively, 52%, 87%, and 93% of controls on days 4, 7, and 10 of growth. The light‐saturated photosynthetic rates at day 6 were significantly lower than controls grown without UV‐B. The probability of electron transfer beyond Q_A decreased during UV‐B exposure, and this indicated that the acceptor side of PSII was one of main damage sites. The adaptation of M. aeruginosa 854 to UV‐B radiation could be observed from light‐saturated photosynthetic rates on day 13 and diurnal changes of chl fluorescence during the late growth phase. When both exposed to higher UV‐B, samples cultured under 1.05 W·m^?2 UV‐B for 9 days recovered faster than controls. It is suggested that M. aeruginosa 854 had at least three adaptive strategies to cope with the enhanced UV‐B: increasing the synthesis of carotenoids to counteract reactive oxidants caused by UV‐B exposure, degrading phycocyanin and allophycocyanin to avoid further damage to DNA and reaction centers, and enhancing the repair of UV‐B induced damage to the photosynthetic apparatus.     

Water- and temperature-dependence of DNA damage and repair in the fruticose lichen Cladonia arbuscula ssp. mitis exposed to UV-B radiation

The induction of cyclobutane pyrimidine dimers (CPDs) by ultraviolet‐B radiation (UV‐B, 280–315 nm) and repair mechanisms were studied in the lichen Cladonia arbuscula ssp. mitis exposed to different temperatures and water status conditions. In addition, the development and repair of CPDs were studied in relation to the different developmental stages of the lichen thallus podetial branches. Air‐dried lichen thalli exposed to UV‐B radiation combined with relatively high visible light (HL, 800 μmol m^?2 s^?1; 400–700 nm) for 7 days showed a progressive increase of CPDs with no substantial repair, although HL was present during and after irradiation with UV‐B. Fully hydrated lichen thalli, that had not been previously exposed to UV‐B radiation for 7 days, were given short‐term UV‐B radiation treatment at 25°C, and accumulated DNA lesions in the form of CPDs, with repair occurring when they were exposed to photoreactivating conditions (2 h of 300 μmol m^?2 s^?1, 400–700 nm). A different pattern was observed when fully hydrated thalli were exposed to short‐term UV‐B radiation at 2°C, in comparison with exposure at 25°C. High levels of CPDs were induced at 2°C under UV‐B irradiation, without significant repair under subsequent photoreactivating light. Likewise, when PAR (300 μmol m^?2 s^?1) and UV‐B radiation were given simultaneously, the CPD levels were not lowered. Throughout all experiments the youngest, less differentiated parts of the lichen thallus – namely ‘tips’, according to our arbitrary subdivision – were the parts showing the highest levels of CPD accumulation and the lowest levels of repair in comparison with the older thallus tissue (‘stems’). Thus the experiments showed that Cladonia arbuscula ssp. mitis is sensitive to UV‐B irradiation in the air‐dried state and is not able to completely repair the damage caused by the radiation. Furthermore, temperature plays a role in the DNA damage repairing capacity of this lichen, since even when fully hydrated, C. arbuscula ssp. mitis did not repair DNA damage at the low temperatures.     

Nitric oxide is involved in integration of UV‐B absorbing compounds among parts of clonal plants under a heterogeneous UV‐B environment

In nature, ultraviolet‐B (UV‐B) radiation is highly heterogeneous, both spatially and temporally. Plants exposed to UV‐B radiation produce UV‐B absorbing compounds that function as a protective filter. For clonal plants under heterogeneous UV‐B radiation conditions, integration among ramets can allow irradiated ramets to benefit un‐irradiated ramets by causing them to increase their UV‐B absorbing compounds content. In this study, we evaluated integration between pairs of clonal ramets of Glechoma longituba under heterogeneous or homogeneous UV‐B conditions. We determined the levels of UV‐B absorbing compounds, nitric oxide (NO) and hydrogen peroxide (H₂O₂) and measured the activity of phenylalanine ammonia‐lyase (PAL) in connected ramet pairs under homogeneous or heterogeneous UV‐B conditions. Under heterogeneous UV‐B conditions, the UV‐B absorbing compounds content increased in leaves of irradiated and un‐irradiated ramets, but not in the connecting stolons. The NO content increased in irradiated and un‐irradiated leaves and stolons, but the H₂O₂ content did not. Application of NO synthesis inhibitors and an NO blocker to irradiated ramets blocked the increase in UV‐B absorbing compounds and PAL activity in un‐irradiated ramets. These results suggested that NO is involved in the integration process for UV‐B absorbing compounds among ramets. Our findings suggested that a UV‐B‐induced increase in NO transmits a signal to un‐irradiated ramets via the stolon, leading to an increase in PAL activity and UV‐B absorbing compounds content. The internal translocation of signal enables members of clonal networks to function as a whole unit and to mount an efficient defensive response to localized UV‐B radiation.     

ATR and MKP1 play distinct roles in response to UV‐B stress in Arabidopsis

Ultraviolet‐B (UV‐B) stress activates MAP kinases (MAPKs) MPK3 and MPK6 in Arabidopsis. MAPK activity must be tightly controlled in order to ensure an appropriate cellular outcome. MAPK phosphatases (MKPs) effectively control MAPKs by dephosphorylation of phosphothreonine and phosphotyrosine in their activation loops. Arabidopsis MKP1 is an important regulator of MPK3 and MPK6, and mkp1 knockout mutants are hypersensitive to UV‐B stress, which is associated with reduced inactivation of MPK3 and MPK6. Here, we demonstrate that MPK3 and MPK6 are hyperactivated in response to UV‐B in plants that are deficient in photorepair, suggesting that UV‐damaged DNA is a trigger of MAPK signaling. This is not due to a block in replication, as, in contrast to atr, the mkp1 mutant is not hypersensitive to the replication‐inhibiting drug hydroxyurea, hydroxyurea does not activate MPK3 and MPK6, and atr is not impaired in MPK3 and MPK6 activation in response to UV‐B. We further show that mkp1 leaves and roots are UV‐B hypersensitive, whereas atr is mainly affected at the root level. Tolerance to UV‐B stress has been previously associated with stem cell removal and CYCB1;1 accumulation. Although UV‐B‐induced stem cell death and CYCB1;1 expression are not altered in mkp1 roots, CYCB1;1 expression is reduced in mkp1 leaves. We conclude that the MKP1 and ATR pathways operate in parallel, with primary roles for ATR in roots and MKP1 in leaves.     

Dimer/monomer status and in vivo function of salt‐bridge mutants of the plant UV‐B photoreceptor UVR8

UV RESISTANCE LOCUS8 (UVR8) is a photoreceptor for ultraviolet‐B (UV‐B) light that initiates photomorphogenic responses in plants. UV‐B photoreception causes rapid dissociation of dimeric UVR8 into monomers that interact with CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1) to initiate signal transduction. Experiments with purified UVR8 show that the dimer is maintained by salt‐bridge interactions between specific charged amino acids across the dimer interface. However, little is known about the importance of these charged amino acids in determining dimer/monomer status and UVR8 function in plants. Here we evaluate the use of different methods to examine dimer/monomer status of UVR8 and show that mutations of several salt‐bridge amino acids affect dimer/monomer status, interaction with COP1 and photoreceptor function of UVR8 in vivo. In particular, the salt‐bridges formed between arginine 286 and aspartates 96 and 107 are key to dimer formation. Mutation of arginine 286 to alanine impairs dimer formation, interaction with COP1 and function in vivo, whereas mutation to lysine gives a weakened dimer that is functional in vivo, indicating the importance of the positive charge of the arginine/lysine residue for dimer formation. Notably, a UVR8 mutant in which aspartates 96 and 107 are conservatively mutated to asparagine is strongly impaired in dimer formation but mediates UV‐B responses in vivo with a similar dose–response relationship to wild‐type. The UV‐B responsiveness of this mutant does not correlate with dimer formation and monomerisation, indicating that monomeric UVR8 has the potential for UV‐B photoreception, initiating signal transduction and responses in plants.     

Effect of UV‐B radiation on the content of UV‐B absorbing compounds and photosynthetic parameters in Parmotrema austrosinense from two contrasting habitats

• We studied the resistance of Parmotrema austrosinense to UV‐B stress. We focused on the effects of a high dose UV‐B radiation on the content of chlorophylls, carotenoids and UV‐B screening compounds.
• Photosynthetic parameters were measured by chlorophyll fluorescence (potential and effective quantum yields, photochemical and non‐photochemical quenching) and evaluated in control and UV‐B‐treated lichens. Lichens from two different locations in Cordoba, Argentina, were selected: (i) high altitude and dry plots at (Los Gigantes) and (ii) lowland high salinity plots (Salinas Grandes).
• UV‐B treatment led to a decrease in the content of photosynthetic pigments and UV‐B screens (absorbance decrease in 220–350 nm) in the samples from Salinas Grandes, while in Los Gigantes samples, an increase in UV‐B screen content was observed. Chlorophyll fluorescence parameters showed a UV‐B‐induced decline in F_V/F_M, Φ_PSII and qP indicating limitation of primary photosynthetic processes in photosystem II (PSII) of symbiotic alga, more pronounced in Salinas Grandes samples. Protective mechanism of PSII were activated by the UV‐B treatment to a higher extent in samples from Salinas Grandes (NPQ 0.48) than in Los Gigantes samples (NPQ 0.26).
• We concluded that site‐related characteristics, and in particular different UV‐B radiation regimen, had a strong effect on resistance of the photosynthetic apparatus of P. austrosinense to UV‐B radiation.

     

Solar UV‐B radiation and ethylene play a key role in modulating effective defenses against Anticarsia gemmatalis larvae in field‐grown soybean

Solar UV‐B radiation has been reported to enhance plant defenses against herbivore insects in many species. However, the mechanism and traits involved in the UV‐B mediated increment of plant resistance are unknown in crops species, such as soybean. Here, we studied defense‐related responses in undamaged and Anticarsia gemmatalis larvae‐damaged leaves of two soybean cultivars grown under attenuated or full solar UV‐B radiation. We determined changes in jasmonates, ethylene (ET), salicylic acid, trypsin protease inhibitor activity, flavonoids, and mRNA expression of genes related with defenses. ET emission induced by Anticarsia gemmatalis damage was synergistically increased in plants grown under solar UV‐B radiation and was positively correlated with malonyl genistin concentration, trypsin proteinase inhibitor activity and expression of IFS2, and the pathogenesis protein PR2, while was negatively correlated with leaf consumption. The precursor of ET, aminocyclopropane‐carboxylic acid, applied exogenously to soybean was sufficient to strongly induce leaf isoflavonoids. Our results showed that in field‐grown soybean isoflavonoids were regulated by both herbivory and solar UV‐B inducible ET, whereas flavonols were regulated by solar UV‐B radiation only and not by herbivory or ET. Our study suggests that, although ET can modulate UV‐B‐mediated priming of inducible plant defenses, some plant defenses, such as isoflavonoids, are regulated by ET alone.     

INTERACTIVE EFFECTS OF PAR AND UV‐B RADIATION ON PSII ELECTRON TRANSPORT IN THE MARINE ALGA DUNALIELLA TERTIOLECTA (CHLOROPHYCEAE)1

To better understand the interactions between PAR and UV‐B radiation in microalgae, the marine chlorophyte alga Dunaliella tertiolecta was subjected to a UV‐B flux of 4.1 W·m ^{? 2} (unweighted) with varying PAR fluxes. Rate constants for damage and repair processes during UV‐B exposure increased with PAR flux. However, recovery after UV‐B exposure increased with PAR up to 300 μmol quanta·m ^{? 2}·s ^{? 1} 1 Received 17 September 2002. Accepted 19 February 2003. , beyond which photoinhibition of PSII electron transport was found to decrease recovery rates. In the absence of PAR during the post UV‐B exposure period, no recovery was seen, indicating that perhaps the lack of light available for photosynthesis depresses repair either directly or indirectly by affecting ATP synthesis. Possible mechanisms for the observed interactions between PAR and UV‐B exposure are discussed.