UVA Stress Research Articles

The Proteasome Is an Integral Part of Solar Ultraviolet A Radiation-induced Gene Expression

Solar ultraviolet (UV) A radiation is a well known trigger of signaling responses in human skin fibroblasts. One important consequence of this stress response is the increased expression of matrix metalloproteinase-1 (MMP-1), which causes extracellular protein degradation and thereby contributes to photoaging of human skin. In the present study we identify the proteasome as an integral part of the UVA-induced, intracellular signaling cascade in human dermal fibroblasts. UVA-induced singlet oxygen formation was accompanied by protein oxidation, the cross-linking of oxidized proteins, and an inhibition of the proteasomal system. This proteasomal inhibition subsequently led to an accumulation of c-Jun and phosphorylated c-Jun and activation of activator protein-1, i.e. transcription factors known to control MMP-1 expression. Increased transcription factor activation was also observed if the proteasome was inhibited by cross-linked proteins or lactacystin, indicating a general mechanism. Most importantly, inhibition of the proteasome was of functional relevance for UVA-induced MMP-1 expression, because overexpression of the proteasome or the protein repair enzyme methionine sulfoxide reductase prevented the UVA-induced induction of MMP-1. These studies show that an environmentally relevant stimulus can trigger a signaling pathway, which links intracellular and extracellular protein degradation. They also identify the proteasome as an integral part of the UVA stress response.

Cellular and sub-cellular responses to UVA in relation to carcinogenesis

Purpose: UVA radiation (315–400 nm) contributes to skin aging and carcinogenesis. The aim of this review is to consider the mechanisms that underlie UVA-induced cellular damage, how this damage may be prevented or repaired and the signal transduction processes that are elicited in response to it.Results: Exposure to ultraviolet (UV) light is well-established as the causative factor in skin cancer. Until recently, most work on the mechanisms that underlie skin carcinogenesis focused on shorter wavelength UVB radiation (280–315 nm), however in recent years there has been increased interest in the contribution made by UVA. UVA is able to cause a range of damage to cellular biomolecules including lipid peroxidation, oxidized protein and DNA damage, such as 8-oxoguanine and cyclobutane pyrimidine dimers. Such damage is strongly implicated in both cell death and malignant transformation and cells have a number of mechanisms in place to mitigate the effects of UVA exposure, including antioxidants, DNA repair, and stress signalling pathways.Conclusions: The past decade has seen a surge of interest in the biological effects of UVA exposure as its significance to the process of photo-carcinogenesis has become increasingly evident. However, unpicking the unique complexity of the cellular response to UVA, which is only now becoming apparent, will be a major challenge for the field of photobiology in the 21st century.

The effect of UV-A (352 nm) stress on chlorophyll fluorescence, chlorophyll a content, thickness of upper cortex and determinate DNA damage in Physcia semipinnata

In the present study, Physcia semipinnata samples were exposed to UV-A (352 nm) in 20 J m −2 for 24 h, 48 h and 72 h to seek the alterations in the PSII photosynthetic quantum yield, thallus anatomy and DNA mutation rate in response to radiation. The Fv/Fm ratio decreased in P. semipinnata following exposure to UV-A for 24 h, 48 h and 72 h. The data of the present study reported that Chl a degradation occurred by exposition of UV-A for 24 h, 48 h and 72 h. Photobiont size and thickness of upper cortex layer also showed a gradually decrease in P. semipinnata thallus sections during 24 h, 48 h and 72 h. The thickness of the upper cortex layer, exposed to UV-A for 72 h, of P. semipinnata thallus reduced 64%, compared with control. The random amplified polymorphic DNA (RAPD) technique was used to detect DNA damage. The main changes observed in the RAPD profiles resulted in both appearance and disappearance of different bands and variation of their intensity. According to RAPD assay, the genetic distance between the control group and 24 h, 48 h and 72 h UV-A exposed, groups was found 56%, 78% and 84%, respectively.

Role of salicylic acid in regulating ultraviolet radiation-induced oxidative stress in pepper leaves

The effect of salicylic acid (SA) counteracting the UV-A, UV-B, and UV-C-induced action on pepper (Capsicum annuum L.) plants was studied. For this purpose, the activities of antioxidant enzymes (peroxidase, polyphenol oxidase, ascorbate peroxidase, catalase, and glutathione reductase) were measured. Plants were sprayed with SA and treated with UV-A (320–390 nm), UV-B (312 nm), and UV-C (254 nm) radiation with a density of 6.1, 5.8, and 5.7 W/m2. The activities of antioxidant enzymes were enhanced in leaves in response to UV-B and UV-C radiation. SA treatment moderated an increase in the activities of some antioxidant enzymes (peroxidase, ascorbate peroxidase, catalase, and glutathione reductase) in plants that were treated with UV radiation. The activity of antioxidant enzyme polyphenol oxidase in plants that were treated with UV-B, UV-C, and SA was significantly increased. The aim of the present study was to investigate the possible protective effect of SA treatment on UV-A, UV-B, and UV-C stress.

The role of external polyamines on photosynthetic responses, lipid peroxidation, protein and Chlorophyll a content under the UV-A (352 nm) stress in Physcia semipinnata

Physcia semipinnata was exposed to UV-A (352 nm) and visible light (210, 800 and 2000 mW/cm 2) for 30 min, 1, 2, 24, and 48 h to seek the alterations in the PSII photosynthetic quantum yield, in response to radiation. Chlorophyll a fluorescence did not influence exposure to light, 210, 800 and 2000 mW/cm 2. Significant alterations of the photosynthetic quantum yield ratio occurred in response to increase in UV-A exposure time. The photosynthetic quantum yield ratio decreased in P. semipinnata following exposure to UV-A for 24 and 48 h. The thalli of P. semipinnata treated with 1 mM polyamine were not influenced during the exposure to UV-A for 24 and 48 h. It was also found that exogenously spd added samples had higher chla content than spm and put added samples. In this study, we showed that lipid peroxidation levels between UV-A-treated samples and exogenously polyamine treated samples that were previously exposed to UV-A for 24 and 48 h were significantly decreased. This result is the first record to indicate that external polyamines might have some protective role on photosystem II and membrane against UV-A stress.

Age-dependent DNA repair and cell cycle distribution of human skin fibroblasts in response to UVA irradiation

Ageing process in cells is associated with oxidative stress. Ultraviolet A produces reactive oxygen species responsible for accumulation of DNA and cellular damage. After the evaluation of antioxidant enzyme activities and oxidative stress markers at the basal state, we have studied the responses to UVA stress of coetaneous fibroblasts, isolated from different male donors (2–88 years, n = 23) in terms of cytotoxicity, genotoxicity and DNA repair capacities. For this purpose, we have determined level of DNA damage using the comet assay (single strand breaks and alkali-labile sites) and the cell cycle distribution after a 5 J/cm 2 irradiation. No differences with age were observed for antioxidant enzyme activities and oxidative stress markers. DNA strand breaks after UVA irradiation (5–20 J/cm 2), was found to be age-dependent. DNA repair was slow and also significantly affected by ageing. The cell cycle distribution analysis showed that high repair correlated with high proliferative capacities at basal level. Twenty-four hours after the stress, fraction of young fibroblasts blocked in G1 phase was significantly increased whereas significant modifications concerned the G2-M phase for adult and older fibroblasts. These results indicate an age-dependent decline in the DNA repair capacities correlated with modifications of the cell cycle parameters.

Ectoin: An Effective Natural Substance to Prevent UVA-Induced Premature Photoaging

With the help of a new ‘UVA stress model’, it was shown that Ectoin protects the skin from the effects of UVA-induced cell damage in a number of different ways. Using cell cultures, high-performance thin-layer chromatography, gel electrophoresis mobility shift assays, reverse transcriptase polymerase chain reaction, ion exchange chromatography and UV spectroscopy, it was demonstrated that the UVA-induced second messenger release, transcription factor AP-2 activation, intercellular adhesion molecule-1 expression and mitochondrial DNA mutation could be prevented. The results obtained clearly demonstrate that Ectoin counteracts the effects of UVA-induced and accelerated skin aging at different cell levels.

Stress tolerance responses of two cotton cultivars exposed to ultraviolet-A (366 nm) radiation: photosynthetic performance and chemical constituents

The stress tolerance responses of two Egyptian cotton cultivars (Giza 45 and 86) exposed to various doses (40, 80, 160 and 320 min) of artificial ultraviolet-A (366 nm) radiation were investigated. The seed germination of Giza 86 was promoted at 40 min, but substantially inhibited at 80 and 160 min and completely suppressed at 320 min. However, the seed germination of Giza 45 was progressively inhibited by UV-A exposure and ceased at 160 min, so doses of 40 and 80 min were selected for further studies. In contrast to seed germination, the seedling growth of Giza 86 was negatively affected at 40 min. UV-A stress induced a great reduction in the leaf carbohydrates as well as in the viability and dry mass production of the shoots of both cultivars, but the response was comparatively higher in Giza 45. It also decreased the chorophyll (Chl) and carotenoid contents, coupled with an increase in the Chla/bratio, diminished the Hill reaction activity, and quenched the Chlafluorescence both in the presence and absence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea, suggesting an inhibitory effect on the water-splitting system (donor side) as well as on the electron transport from the primary to the secondary acceptors of PSII (acceptor side). These changes reflect a disturbance in the structure, composition and function of the photosynthetic apparatus as well as the sensitivity of PSII to UV-A stress. Nucleic acids (DNA and RNA) were markedly damaged by exposure to UV-A for 80 min, while both cultivars developed adaptive mechanisms for damage moderation. These mechanisms involved increasing the levels of flavonoids, total lipids and total soluble proteins as well as having smaller, thicker leaf blades. Since Giza 86 showed a comparatively higher level of adaptation, it tolerates UV-A stress better than Giza 45.Abbreviations:Car, carotenoids; Chl, chlorophyll; DCMU, 3-(3,4-dichlorophenyl)-1,1-dimethylurea; DCPIP, 2,6-dichlorophenol indophenol; DNA, deoxyribonucleic acid; d.m., dry mass; f.m., fresh mass; PSII, photosystem II; RNA, ribonucleic acid; TSP, total soluble proteins; UV-AR, ultraviolet-A (366 nm) radiation.

Identification of genes responsive to UV-A radiation in human lens epithelial cells using complementary DNA microarrays.

UV-A radiation produces cataract in animals, enhances photoaging of the lens and skin and increases the phototoxicity of drugs. However, the nature of genes that are activated or repressed after cellular exposure to UV-A radiation remains enigmatic. Because lens epithelial cells exposed to UV-A radiation undergo apoptosis 4 h after exposure to the stress, we sought to establish the change in gene expression in cells by UV-A radiation using gene expression profiling using complementary DNA microarrays containing about 12 000 genes. We identified 78 genes abnormally expressed in UV-A-irradiated cells (showing >2.5-fold change at P < 0.05). These genes are implicated in various biological processes, including signal transduction and nucleic acid binding, and genes encoding enzymes. A majority of the genes were downregulated. Our analysis revealed that the expression of genes for the transcription factors ATF-3 and Pilot increased four-fold, whereas the gene for the apoptosis regulator NAPOR-1 decreased five-fold. These changes were confirmed by real-time quantitative reverse transcriptase-polymerase chain reaction. The calpain large polypeptide 3 (CANP3) gene also increased nine-fold after UV-A radiation. In addition, peroxisomal biogenesis factor 7, glucocorticoid receptor-alpha and tumor-associated calcium signal transducer genes decreased three- to eight-fold. Western blot analysis further confirmed the increase in protein expression of ATF-3 and CANP3 and decreased expression of glucocorticoid receptor-alpha in the irradiated cells. Surprisingly, most of these genes had not been previously shown to be modulated by UV-A radiation. Our results show that human lens epithelial cells respond to a single dose of UV-A radiation by enhancing or suppressing functionally similar sets of genes, some of which have opposing functions, around the time at which apoptosis occurs. These studies support the intriguing concept that activation of competing pathways favoring either cell survival or death is a means to coordinate the response of cells to UV-A stress.

Specificity in stress response: epidermal keratinocytes exhibit specialized UV-responsive signal transduction pathways.

UV light, a paradigmatic initiator of cell stress, invokes responses that include signal transduction, activation of transcription factors, and changes in gene expression. Consequently, in epidermal keratinocytes, its principal and frequent natural target, UV regulates transcription of a distinctive set of genes. Hypothesizing that UV activates distinctive epidermal signal transduction pathways, we compared the UV-responsive activation of the JNK and NFkappaB pathways in keratinocytes, with the activation of the same pathways by other agents and in other cell types. Using of inhibitors and antisense oligonucleotides, we found that in keratinocytes only UVB/UVC activate JNK, while in other cell types UVA, heat shock, and oxidative stress do as well. Keratinocytes express JNK-1 and JNK-3, which is unexpected because JNK-3 expression is considered brain-specific. In keratinocytes, ERK1, ERK2, and p38 are activated by growth factors, but not by UV. UVB/UVC in keratinocytes activates Elk1 and AP1 exclusively through the JNK pathway. JNKK1 is essential for UVB/UVC activation of JNK in keratinocytes in vitro and in human skin in vivo. In contrast, in HeLa cells, used as a control, crosstalk among signal transduction pathways allows considerable laxity. In parallel, UVB/UVC and TNFalpha activate the NFkappaB pathway via distinct mechanisms, as shown using antisense oligonucleotides targeted against IKKbeta, the active subunit of IKK. This implies a specific UVB/UVC responsive signal transduction pathway independent from other pathways. Our results suggest that in epidermal keratinocytes specific signal transduction pathways respond to UV light. Based on these findings, we propose that the UV light is not a genetic stress response inducer in these cells, but a specific agent to which epidermis developed highly specialized responses.

Induction, exudation and the UV protective role of kelp phlorotannins

This study examined the ability of kelp phlorotannin exudate to attenuate artificial UVB radiation and whether exposure to UV could affect short-term phlorotannin levels within Macrocystis integrifolia Bory, the surrounding seawater or affect the seawater UV transparency. A dilution series of seawater that had contained M. integrifolia, reduced UVB radiation in an inverse relationship. Phlorotannin levels detected in this contact seawater were also inversely related to UVB exposure. Increasing concentrations of seawater previously containing M. integrifolia with phlorotannin concentration from 0.5 to 10.5 μg ml −1, significantly increased ( P<0.05) the survivorship (from 0 to 100%) of germinating Laminaria groenlandica Rosenvinge meiospores grown under an artificial UVB radiation source. Short-term exposure of M. integrifolia blades to UVA and UVB radiation significantly ( P<0.05) increased phlorotannins within tissues (∼10% of dry tissue weight) as compared to PAR or dark exposed algae (∼8.5–7.5%, respectively). UVA radiation was found to significantly ( P<0.05) increase contact seawater UVB absorbance from ∼0.01 to 0.025 (Abs 300 nm g −1 wet blade), which was mirrored, though not significantly ( P>0.05), by concentrations of phlorotannins within contact liquid. Results suggest that phlorotannins from macroalgal sources are produced and released into seawater during periods of UVA stress but degraded by UVB and can, at low concentrations (>0.84 μg ml −1), reduce the impact of UVB exposure to UV-sensitive kelp meiospores. Kelp beds are significant point sources of water soluble UV-screening compounds, and may form natural UV-refugia along coastal shores.

The Molecular Chaperone αA-Crystallin Enhances Lens Epithelial Cell Growth and Resistance to UVA Stress

alphaA-Crystallin (alphaA) is a member of the small heat shock protein (sHSP) family and has the ability to prevent denatured proteins from aggregating in vitro. Lens epithelial cells express relatively low levels of alphaA, but in differentiated fiber cells, alphaA is the most abundant soluble protein. The lenses of alphaA-knock-out mice develop opacities at an early age, implying a critical role for alphaA in the maintenance of fiber cell transparency. However, the function of alpha-crystallin in the lens epithelium is unknown. To investigate the physiological function of alphaA in lens epithelial cells, we used the following two systems: alphaA knock-out (alphaA(-/-)) mouse lens epithelial cells and human lens epithelial cells that overexpress alphaA. The growth rate of alphaA(-/-) mouse lens epithelial cells was reduced by 50% compared with wild type cells. Cell cycle kinetics, measured by fluorescence-activated cell sorter analysis of propidium iodide-stained cells, indicated a relative deficiency of alphaA(-/-) cells in the G2/M phases. Exposure of mouse lens epithelial cells to physiological levels of UVA resulted in an increase in the number of apoptotic cells in the cultures. Four hours after irradiation the fraction of apoptotic cells in the alphaA(-/-) cultures was increased 40-fold over wild type. In cells lacking alphaA, UVA exposure modified F-actin, but actin was protected in cells expressing alphaA. Stably transfected cell lines overexpressing human alphaA were generated by transfecting extended life span human lens epithelial cells with the mammalian expression vector construct pCI-neoalphaA. Cells overexpressing alphaA were resistant to UVA stress, as determined by clonogenic survival. alphaA remained cytoplasmic after exposure to either UVA or thermal stress indicating that, unlike other sHSPs, the protective effect of alphaA was not associated with its relocalization to the nucleus. These results indicate that alphaA has important cellular functions in the lens over and above its well characterized role in refraction.

Heat shock protein 70 overexpression affects the response to ultraviolet light in murine fibroblasts. Evidence for increased cell viability and suppression of cytokine release.

To elucidate cellular concepts for protection against ultraviolet (UV) light we investigated the effect of heat shock protein 70 (hsp70) overexpression on cell viability and on the secretion of UV-inducible immunological cytokines. Transfected murine fibrosarcoma cells (WEHI-S), overexpressing hsp70 or a sham transfected control were used. Overexpression of hsp70 was sufficient to markedly increase cell viability upon treatment with UVB (290-320 nm). Since long wave UV (UVA, 320-400 nm) as well as UVB turned out to stimulate the release of O2- radicals we studied the cell viability upon oxidative stress. Hsp70 overexpression increased viability upon treatment with hydrogen peroxide or menadione, but had no influence on UV-induced O2- release. UV-light is known to upregulate immunologic and proinflammatory cytokines such as IL-1 and IL-6. Oxidative stress appeared to exert a similar effect. Hsp70 overexpression markedly decreased the release of IL-6 induced by UVA, UVB and oxidative stress. To test whether the hsp70 mediated suppression is confined to events caused by UV-light we determined IL-1-mediated effects. IL-1-induced IL-6 release was reduced by hsp70 overexpression, whereas the IL-1 mediated activation of nuclear factor kappa B was not affected. Our data suggests that hsp70 plays a central role not only in cell protection against UV-light, but also in the regulation of proinflammatory cytokine release induced by UV-exposure.

UV-A oxidative damage modified by environmental conditions in Escherichia coli.

The effect of sublethal fluences (50-200 kJ m-2) of UV-A radiation (320-400 nm) in bacterial cells is a transient growth inhibition related to photo-modified tRNA and is associated with changes in membrane structure and function. Higher UV-A fluences result in cell death due to the production of reactive oxygen species, so far undetected at sublethal doses. Oxidative mechanisms of toxicity induced by 120 kJ m-2 UV-A radiation can be recorded by ultra-weak chemiluminescence, useful in quantifying oxidative reactions. When Escherichia coli was exposed to UV-A stress at a fluence rate equivalent to that of the Sun in the biosphere (33 W m-2), chemiluminescence levels were proportional to the photodamage. Chemiluminescence and photo-damage are linearly proportional and dependent on environmental conditions of the cells. It is postulated that in addition to tRNA photo-modification, UV-A alters the membrane structure of E. coli by oxidative damage, since changes in the membrane structure under different environmental conditions play a key role in the cell's response to UV-A injury.