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Year 2011 noted the first definable ozone “hole” in the Arctic region, serving as an indicator to the continued threat of dangerous ultraviolet radiation (UVR) exposure caused by the deterioration of stratospheric ozone in the northern hemisphere. Despite mandates of the Montreal Protocol to phase out the production of ozone depleting chemicals (ODCs), the relative stability of ODCs validates popular notions of persistent stratospheric ozone for several decades. Moreover, increased UVR exposure through stratospheric ozone depletion is occurring within a larger context of physiological stress and climate change across the biosphere. In this review, we provide commentaries on stratospheric ozone depletion with relative comparisons between the well-known Antarctic ozone hole and the newly defined ozone hole in the Arctic. Compared to the Antarctic region, increased UVR exposure in the Northern Hemisphere poses a threat to denser human populations across North America, Europe and Asia. In this context, we discuss emerging targets of UVR exposure that can potentially offset normal biological rhythms in terms of taxonomically conserved photoperiod dependent seasonal signaling and entrainment of circadian clocks. Consequences of seasonal shifts during critical life history stages can alter the fitness and condition, while circadian disruption is increasingly becoming associated as a causal link to increased carcinogenesis. We further review the significance of genomic alterations via UVR induced modulations of phase I and phase II transcription factors, the aryl hydrocarbon receptor (AhR) and the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2), with emphasis on mechanism that can lead to metabolic shifts and cancer. While concern for adverse health consequences due to increased UVR exposure are longstanding, recent advances in biochemical research suggest that AhR and Nrf2 transcriptional regulators are likely targets for UVR mediated dysregulations of rhymicity and homeostasis among animals, including humans.
Anthropogenic induced climate change poses a significant risk to long term ecological sustainability and human health. Depletion of stratospheric ozone at the poles is recognized as one of the most significant environmental threats characterized in the past several decades. Moreover, the response and establishment of the Montreal Protocol in 1987 (1) with the goal to curtail this glooming threat is championed as a success story of international environmental policy; however, the threat of detrimental exposure to ultraviolet radiation (UVR) still persists.
Until 2011, the depletion of stratospheric ozone associated with the Antarctic polar vortex was the largest and most characterized ozone threat. During the period associated with the spring equinox (March) of 2011, Arctic ozone depletion was recorded at a record high and comparable to that of the Antarctic ozone hole. National Aeronautics and Space Administration (NASA) research has indicated that record cold temperatures in the arctic stratosphere during the winter of 2011 prompted a record breaking amount of ozone depletion in the arctic north (2). The 2011 arctic ozone layer was approximately ~2 million kilometers2 (~ 2x size of Ontario, Canada).
The atmosphere encircles the earth, allowing photons of light energy to exergonically fuel the biosynthesis of earth’s primary chemical food source (i.e. glucose), while filtering the sun’s most harmful UVR from entering the biosphere. Gases in the upper atmosphere absorb virtually all of the sun’s UVC waves (100 to 290 nm), while the ozone layer, located in the stratosphere (~ 10–50 kilometers above sea level) absorbs a majority of the UVB (290 to 320 nm) and UVA (320 to 400 nm) waves. Industrial sourced nitrogen oxides, halogenated hydrocarbons, chloroflorocarbons (CFCs), and bromines released into the atmosphere have been identified as leading ozone depleting chemicals (ODCs) in the stratosphere, causing increased UVR penetration in recent decades (Kyoto Protocol 1997, Montreal Protocol 1987) (1, 3).
Numerous ecological and environmental health risks are associated with excessive UVR exposure, particularly UVB. At the ecological scale, increased exposure of UVR is attributed to global shifts in primary production (4, 5). While rainforests are the most productive ecosystems, oceans dominate the earth’s surface area and therefore validate concerns of UVR mediated alterations of marine ecosystems (4). At the organismal level for aquatic and terrestrial species, both plants and animals are at risk, and the most pronounced physiological alterations induced by excessive UVR exposure are the production of reactive oxygen species (ROS) (6, 7, 8), DNA mutations (6), and elevated carcinogenicity (9, 10). Gene expression can be altered by UVR in both plants (7, 11, 12) and animals (13). Overall, from a life history standpoint, any UVR mediated biochemical or physiological alteration has the potential to reduce individual fitness, thereby leading to reduced populations and negatively impacting ecological integrity.
Herein, we will combine a brief comparative characterization of the Antarctic and Arctic scenarios regarding the global threat to loss of stratospheric ozone and the subsequent increased in UVR exposure, with a commentary on emerging routes of genomic dysregulation and their potential deleterious physiological effects. Specifically, we will consider UVR modulation of phase I and phase II transcription factors, the aryl hydrocarbon receptor (AhR) and the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and address the potential for dysregulation of coordinated physiological processes, including cross talk between AhR and Nrf2. We underscore the reliance of sunlight among biological systems for the adaptive timing of seasonal and circadian rhythms, and emphasize the potential for UVR exposure to cause a loss of fitness among target organisms and address risks to human health.
The ozone layer is formed naturally by two possible reactions in the lower altitudes of the stratosphere 1) (O2 + (UV-C λ <242.4 nm) → O + O; 2) O + O2 + (N2, or O2) → O3 + N2 or O2). Stratospheric ozone levels from Antarctica have been monitored by British run Antarctic Survey stations since the late 1950’s and trend analyses of these data reported a weight of evidence demonstrating the occurrence of springtime (September) depletions of stratospheric ozone in the Antarctic region (14, 15). Methyl bromides and halogenated hydrocarbons, particularly chlorofluorocarbons (CFCs) which were historically characterized to undergo photodissociation via a series of reactions (CF2Cl2 + UV → Cl. + CCIF.2) (16) are identified as the main exogenous sources of ozone depletion.
The industrial utility of CFCs and various halogenated hydrocarbons is due to their stability. CFC compounds found great utility as refrigerants and solvents, while bromines were heavily used as fumigants (17, 18); however, their stability is a major threat due to environmental persistence. (> 100 years). Due to the threat of halogenated hydrocarbon mediated ozone destruction, the Montreal Protocol (1987, and later amendments) formulated as an international treaty to differentially delineate guidelines for developed and developing countries to eradicate the production of halogenated hydrocarbons known to cause stratospheric ozone destruction (17). While existing CFCs and other ozone depleting substances will persist for decades, the effort of the Montreal Protocol is regarded as a victorious achievement of environmental global policy.
The ability of CFCs and other ODCs to induce ozone destruction at the poles is driven by multiple factors, including the coupling of extreme cold winter temperatures, followed by springtime solar radiance (19). Stratospheric ozone accumulation at the poles is largely driven by Brewer-Dobson circulation patterns that naturally draw ozone from the tropics towards the polar regions (20). During cold winter months, circular wind patterns at the poles lead to the formation polar stratospheric clouds (PSC) and the formation of stratospheric polar vortices (SPV) (21) which further concentrate reactive atmospheric chemicals such as nitric acid, and provide a binding substrate for the parent ozone depleting chemicals (ODCs) such as CFCs (ClO + NO2 → ClONO2 ; ClONO2 + H2O → Cl2 + HNO3 ; Cl2 + hv → 2Cl.). When the sun begins to rise in the springtime, these ODCs photo dissociate forming chlorine free radicals that being to attach ozone in a series of chain reactions (Cl. + O3 → ClO. + O2 ;ClO. + ClO.→ ClOO. + Cl. ; ClOO. + M (N2) → Cl. + O2 + M (N2) ; 2Cl.+ 2O3 → 2ClO.+ 2O2 ; 2O3 → 3O2 (Net reaction)). The final reactions of Cl free radical destruction of ozone occur in a cascade with one Cl having the potential to attack thousands of O3 molecules.
Measurements of ozone depletion are quantified in Dobson units (DU), whereas one DU refers to a layer of ozone that would be 10 µm thick under standard temperature and pressure (i.e. 273.15 K (0 °C, 32 °F) and an absolute pressure of 100 kPa (14.504 psi, 0.986 atm). A semi-arbitrary marker of any value <220 DUs is set to designate an “ozone hole”. From the commencement of stratospheric monitoring in the 1950’s up until 1979, there were no values of <200 DU. Trends of historical and contemporary data indicate that magnitude of ozone depletion in the Antarctic region has been the greatest recorded. Between 1979–2008, 30 ozone “holes” were documented (22) during the Antarctic spring time, where ozone has been observed to undergo >90% decrease (23). The spring of 2000 is verified as the greatest loss of ozone recorded in the Antarctic region, measuring total column ozone of approximately 93.4 DU (15 day average ozone hole minimum) over a 29.8 million km2maximum area (22). Comparatively, 2008 was the 5th largest ozone depletion on record for the Antarctic region, measuring 99.9 DU over an expanse of 27.1km2 (22).
The persistence of ozone depletion in the stratosphere is apparent despite efforts of the Montreal Protocol (20). Continued ozone depletion “post Montreal Protocol” is not absolutely surprising given the extended lifespan of ozone depleting chemicals; in fact it is predicted that ozone depletion will continue throughout much of the 21st century (19). Moreover, climate change is considered a viable burden to ozone recovery (20), and it is noteworthy that 2011 marked the first time a true “ozone hole” was recorded in the Arctic stratosphere (2). During one week of the Arctic spring (March 2011), the ozone column of the Arctic was between 220–230 DU over a range of approximately 2 million km2 (2). For a period of 27 consecutive days, the 2011 Arctic ozone layer was > 250 DU. The Arctic vortex and area of ozone depletion is roughly the size of Germany, California, or 2x size of Ontario. Although the Arctic vortex is approximately 40% smaller than the Antarctic, the region of Arctic ozone depletion tends be displaced in its position relative to the center of the North Pole (2, 24, 25, 26) posing a viable threat of harmful UVR exposure for dense populations across 3 northern continents (i.e. North America, Europe, and Asia).
Polar stratospheric ozone depletion is a dynamic and complex phenomenon that is the subject of rigorous character analysis and predictive modeling which is beyond the scope of this review. Nevertheless, a few points are worth highlighting and from both functional and policy standpoints that suggest ozone depletion and climate change are not mutually exclusive phenomena. Because ODCs are also greenhouse gases, the Montreal Protocol also works toward the aim of the Kyoto Protocol (19, 27) (Figure 1), and the goals of Kyoto may help reduce ozone depletion. Furthermore, persistent ozone depletion also functions as a positive feedback for stratospheric cooling and subsequent ozone depletion (24), which can influence tropospheric conditions in both the Antarctic (28) and Arctic poles (29). Bidirectional coupling between the stratosphere and troposphere is paramount for climatic research focused on unraveling the relationships between ozone depletion and climate change (21, 28, 29).
Greenhouse gases (GHGs) have long been modeled as a causal factor in stratospheric cooling (26), and carbon dioxide emissions can exasperate the conditions that promote ozone depletion through several processes. Recent studies indicate GHGs cause an increased temperature differential between the troposphere and stratosphere which is linked to the acceleration of Brewer-Dobson circulation patterns (30). In this model of Arctic SPV enhancement, interannual variability of thermal gradients between sea and air is correlated with the GHGs influence on stratospheric forcing by Rossby waves (31). Additionally, modeling suggests that carbon dioxide may influence the tropospheric coupling with the SPV through tropical quasi-biennial oscillation and the Holten-Tan mechanism (32, 33, 34).
Surface climates in the polar regions illustrate a time shifted summer warming trend since the 1970’s, correlated with more contracted and intensified SPVs and increased wintertime stratospheric cooling (21, 35). In both hemispheres, the stratospheric influences on surface conditions are largely mediated through polar Annular Modes (21, 28, 29), while in the Arctic region, the Annual Mode shows significantly larger interannual variability (21, 36). In addition to various factors of interannual climatic variability (21), fluxes in the Annular Mode appear to be contributing to the shape irregularities and displacement of the vortex from the pole (24, 26, 36). Overall, differential cooling and patterns of interannual variability between the Arctic and Antarctic regions are suspected factors of their divergent patterns of ozone depletion (23, 24, 37). In the Arctic, atypical stratospheric cooling in the presence of ozone depletion is occurring in both the winter and spring months (24), versus just during the spring in Antarctica, which is naturally colder in the winter.
In both the northern and southern hemisphere, the SPV and interannual variations in the Annual Modes are important drivers for predictive modeling of surface climatic trends (28, 29). Nevertheless, the complexity and dynamics associated with ozone depletion and global climate change challenge consensus. Interestingly, 2011 summer ice sheet melts were the second greatest on record for the Arctic region (most ice sheet melt in 2007) (37, 38), while in Antarctica, sea ice has expanded in some areas and receded in others (37, 39). An undefined combination of factors drives these differential patterns, though the variations in the Annular Modes are likely contributors (37, 39, 40). If greenhouse gas emissions are not hindered, Antarctic ice advances are predicted to retreat later in the 21st century as ozone depletion begins to be minimized (37).
Achieving the goals of the Montreal and Kyoto Protocols are not likely to be met concurrently, and the dynamics between ozone depletion and climate change remain elusive and controversial. Additionally, it remains to be seen what types of indirect benefits may arise from long-term storage of GHGs in geological sequestrations, or the development of renewable green energy, including biofuel (41, 42, 43).
In the context of this review, further dialog is encouraged in an attempt to incorporate and model the interrelationships between ozone depletion driven UVR exposures and GHG mediated climate change that may further intensify modulations of environmental health endpoints (44). Physiological dysregulations induced by UVR exposures will be in the context of other physiological stresses sourced from climate change, including fluxes of short and long wave radiations, and warmer temperatures (45, 46). Physiological stress associated with global warming induces the expression of antioxidant enzymes that are elevated in response to UVR (47, 48), further suggesting that UVR mediated physiological responses should be considered in the context of global climate change.
Periodicity and timing are critical to the life history strategy of virtually all sexually reproducing species. The primary timing strategy is driven by photoperiod, which works as a zeitgeber “time giver”, scheduling life history strategies according to predictable environmental climatic patterns. The life histories of plants and animals have evolved in the context of photoperiod as the primary ecological timing signal triggering hormonal regulation of an organisms’ physiology and coordinating downstream endocrine pathways. This signaling system provides environmental context for the timing of development, growth, feeding, migration and reproduction, translating into seasonal patterns of blood plasma levels for several hormone systems, receptors, and metabolizing enzymes (49, 50, 51, 52, 53, 54, 55).
Recent data indicates a molecular basis for photoperiod based timing (56, 57) among both plant and animal lineages, in which the light responsive pathways are tryptophan dependent, utilizing functionally similar tryptophan monooxygenase enzymes. The reliance of tryptophan dependent pathways mediating the initial photoperiod driven cues among plants and animals is noteworthy considering its ratio of occurrence of tryptophan within the genetic code, where it is the least abundant amino acid along, and along with methionine, it the only amino acid that is coded by a single codon. From an evolutionary standpoint, eukaryotic plants and eukaryotic animals are not sister taxa (meaning they are not considered to share the same common ancestor). These observations suggest a profoundly specific functional role of tryptophan in light sensitive biochemistry (58), herein emphasizing the dependence of tryptophan within specific light driven pathways that meet the demands to be in “ecological time”. Among plants, photoperiod is the initial signal for the synthesis of auxin, the primary plant development hormone (59, 60, 61), while melatonin synthesis in response to light cycles for invertebrates and vertebrates acts as the initial cue of the neuroendocrine system, via tryptophan dependent pathways (62, 63, 64).
Among extant vertebrates, melatonin is produced in the pineal body and the hypothalamus is the primary interface between the central nervous system and physiological activities throughout the body (65). Historical significance for photoperiod environmental signaling is further evident in the fossil record of some Paleozoic fishes (66). The Placoderms and Ostracoderms fossils possess a “third eye” on their dorsum, which has been hypothesized to be a light sensing organ, homologous to the external pineal body of extant lampreys. Among lampreys, the pineal body is an external version of the pineal gland that is generally located within the cranial cavity of most vertebrates.
The neuroendocrine system of vertebrates is very complex and interacts with downstream pathways through processes of positive and negative feedback. The light dependent hypothalamus maintains proper function of the central clock within the suprachiasmatic nucleus (SCN). The biological timing strategy synchronizes the central clock and circadian rhythmic function of normal homeostatic physiology. Melatonin levels that vary according to daily light cycles entrains the central clock of invertebrate and vertebrates, by synchronizing peripheral circadian oscillators within various organs through a negative feedback loop of genetic transcription (Figure 2) (67, 68). The circadian clock drives daily rhythms of gene expression for nuclear receptors, metabolizing enzymes, and hormones associated with reproduction, blood glucose levels, and energy metabolism (67, 68, 69, 70, 71).
Circadian rhythms share a common adaptive value across major taxonomic lineages (72, 73), whereas selective pressure acting over the course of geologic time led to UVR sensitive processes being scheduled and coordinated according to light/day cycles (74). Central clock mediated gene expression translates into circadian patterns of molecular signaling pathways that maximize physiological efficiency (75, 76), minimize DNA damage (77), oxidative stress (78) and metabolism (71, 79, 80). Variations in the overall oscillator feedback pathways suggest independent evolutionary origins of specific circadian mechanisms among algae, plants, and animals (74, 81), however the occurrence of the basic Helix-Loop-Helix (bHLH) - Per-ARNT-SIM (PAS) related family of transcription factors supports a hypothesis of homology among select circadian genes across multiple kingdoms (72, 74).
Homologous clusters of bHLH-PAS circadian genes including CLOCK, cycle, and Period (Per) have been found throughout the Bilateria (82). The mammalian PAS domain shares structural homology to the prokaryotic environmental sensor protein, photoactive yellow proteins (PYP) (83, 84, 85). The PAS domain is primarily limited to proteins directly involved in signal transduction pathways (84) and among prokaryotes, environmental cues including light (86) and oxygen (84) stimulate PYP, suggesting ancestral origins for light as a signal for PAS mediated genetic transcription. Among fungi (Neurospora), the white collar (wc-1, wc-2) genes are circadian regulators with bHLH-PAS domains, and in plants (Arabidopsis) the PIF-3 phytochrome is a PAS containing regulator of flowering time (74, 87).
Oxidative stress generated by the production of ROS, including free radicals and oxidants such as hydrogen peroxides, are common physiological effects of UVR exposure among plants and animals (88, 89). The biotransformation of ROS and xenobiotics through phase I and phase II reactions is a ubiquitous regulatory mechanism among organisms that functions in part to maintain relative homeostasis in response to adverse environmental exposures, including UVR. In preparation for phase II translocation from cells, phase I reactions produce metabolites that are more water soluble than the binding toxin, while phase II reactions produce conjugate bases (90, 91). The aryl hydrocarbon receptor (AhR) and the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) are transcription factors that are emerging as significant targets of UVR mediated dysregulations leading to potentially adverse health effects (92, 93). The AhR and Nrf2 transcription factors provide a primary means of response to UVR induced stress, and their modulations are linked to various UVR skin diseases and effects of photoaging caused by ROS (94).
In addition to seasonal and circadian patterns of expression among metabolic enzymes (95, 96), the activities of phase I and phase II enzymes are often synchronized via response elements of target genes, including those expressed by the AhR and Nrf2 transcription factors (97, 98). Phase I and phase II enzymes are induced by AhR in association with the xenobiotic responsive element (XRE) promotor, while phase II enzymes and antioxidant enzymes are more commonly regulated through Nrf2 in association with the antioxidant response element (ARE) promotor (91, 98). The phase II induction by AhR pathways occurs for enzymes such as glutathione S-transferase (GST) and NAD(P)H quinone oxidase (NQO), which are identified to possess both the XRE and the ARE promotor (99, 100). Furthermore, bi-directional crosstalk between AhR and Nrf2 dependent gene transcription has been verified by the identification of XRE promoters on Nrf2 gene and ARE promoters on AhR genes (101, 102, 103). Accordingly, the coordination of phase I and phase II enzymes are regulated by ligands of Ahr, oxidative stressors including phase I metabolites that regulate AhR and or Nrf2 mediated phase I and phase II activities and transcriptional crosstalk between AhR and Nrf2 (91, 98, 104).
Phylogenetic studies indicate that the vertebrate AhR gene is basal to a pair of reciprocally monophyletic AhR clades; denoted as AhR1 and AhR 2 (105). The AhR protein is a member of the bHLH–PAS family of transcription factors (83) that occurs throughout the metazoa (Figure 3). When unbound by a ligand, the AhR transcription factor is suppressed in the cytoplasm by association with heat shock protein 90 (hsp90), the co-chaperone protein (p23), and the X-associated protein (XAP 2) that is also known as AhR-interacting protein (AIP) (106, 107, 108). Lipid soluble ligands that can pass through the cell membrane and bind with the AhR transcription factor while positioned in the cytosol, in association with the chaperone complex (109). The ligand binding domain (LBD) of AhR is located within the PAS core of the PAS-B domain.
The Ah receptor is also known as the “dioxin receptor”, as the exogenous halogenated aromatic hydrocarbon (HAH) 2’,3’,7’,8’-Tetrachlorodibenzo-p-dioxin (TCDD; or dioxin) (Figure 4) has historically demonstrated the strongest binging affinity for AhR (109, 110). Other xenobiotics, including polycyclic aromatic hydrocarbons (PAHs) also induce AhR mediated gene expression. The continued quest for endogenous ligands for AhR most recently implicates tryptophan photoproduct, 6-Formylindolo [3,2-b] carbazole (FICZ) (Figure 4) as the most probable natural ligand in vivo (111, 112, 113). Indeed, endogenous FICZ shows the strongest affinity for AhR, exceeding the binding affinity of exogenous TCDD (111, 112). Numerous genes are expressed via the AhR pathway, with the cytochrome p450 metabolizing enzyme CYP1A1 being the most thoroughly characterized. Recent findings of strong affinity of FICZ to AhR support earlier in vitro studies that showed UVR regulates AhR dependent gene expression of CYP1A1 in mammalian keratinocytes and lymphocytes through tryptophan photoproducts (114), and substantiates a role for AhR in skin related environmental response.
The AhR is a highly likely target for UVR mediated alterations of gene expression in the context of ozone depletion. Mice studies indicate that UVB exposed AhR knock out mice failed to express CYP1A1 in keratinocytes, as opposed to AhR+ mice (115) and AhR dependent cytochrome genes failed to be expressed in AhR knockout chicken embryos treated with tryptophan metabolites produced by exposing tryptophan to natural sunlight (116). Similarly, zebrafish embryos demonstrate CYP1 gene expression in an AhR dependent manner when exposed to UVB (117) or FICZ (118), indicating that a positive relationship between tryptophan photoproducts and AhR mediated gene expression occurs in a taxonomically diverse array of vertebrates.
The propensity for skin diseases such as melanomas and psoriasis, as a result of UVR modulations of AhR signaling pathways is gaining increased attention within the environmental health sciences (9, 119, 120) including AhR dependent photoresponsive immune responses (121, 122, 123), speculatively linked to psoriasis. The activation of AhR by FICZ promotes the differentiation of CD4+ cells into Th17 lymphocytes and downstream secretion of IL-22 cytokines (121, 122, 124), both of which are major inflammatory components of psoriasis (125, 126). Some evidence suggests that TCDD treatments differentially induce the CD4+ into Treg cells (123). TCDD treatments of human keratinocytes demonstrate a suppression of this pathway, possibly through cross-talk between transforming growth factor (TGF-β) and AhR (123).
Among humans, there are polymorphic variants of AhR dependent phase I and II enzymes that either provide protection against or increase the risk of psoriasis (127). UVR signaling of AhR induces multiple CYP genes within the epidermis, and their expression exhibits cross-sectional stratification (128). In human keratinocytes exposed to UVB, CYP1A1 was expressed basally to CYP1B1 (128). AhR dependent enzyme expression in response to UVR appears varied across tissue boundaries and the complete role of AhR mediated skin disorders is not fully characterized.
The AhR receptor clearly plays a role in melanogenic pathways of tanning among mammals. Both FICZ and UVB exposures to murine keratinocyte cell lines have identified a facultative AhR–CYP1A1 pathway of melanin synthesis that causes an upregulation of tyrosinase, which is known as the rate limiting step of melanogenesis (92). The AhR pathway of melanogenesis appears restricted to melanocytes and has been verified in human cells in vitro, with TCDD induction of AhR (129). Furthermore, studies on human HaCaT keratinocytes exposed to UVR and FICZ have been forwarded to suggest that UVR mediated expression of CYP1A1 and CYP1B1 through AhR may prime skin tissue to PAH exposures, thereby increasing the risk factor for skin cancers exposed to PAHs (130). Alternatively, UVR mediated activation of AhR may provide resistance to PAH driven malignancies (130).
The NF-E2-related factor 2 (Nrf2) transcription factor is a primary target of response to oxidative stress and metabolic action aimed at minimizing carcinogenic metabolites (88). Nrf2 is a phylogenetically ubiquitous gene of the basic leucine zipper (bZIP) family, that has invertebrate homologs known as SKN-1 (Caenorhabditis sp.) and CncC 9 (Drosphila sp.) (88, 131, 132). Among vertebrates, the Nrf2 system appears to be highly conserved comparing teleost fishes to mammals. In the presence of thiol reactive ROS and xenobiotic inducers, Nrf2 dissociates from the Kealp-1 suppressor protein in association with the endoplasmic reticulum, and translocates to the nucleus to form a heterodimer with a small Maf protein that then binds with the antioxidant responsive element (ARE) (88, 132). Numerous proteins and enzymes that target ROS and exogenous molecules are transcriptionally regulated by Nrf2-ARE binding including, glutathione S-transferase (GST), NAD(P)H quinone oxidase (NQO), super oxide dismutase (SOD), heme oxigenase 1 (HO1), and UDP-glucuronosytransferase (UGT) (131, 133).
Recent research is steadily characterizing the effects of UVR induced ROS on Nrf2 dependent phase II enzymes among various types of mammalian skin cells. The expression of several Nrf2 dependent phase II enzymes are altered in response to UVR, and among human keratinocytes and melanocytes, both UVA and UVB have been shown to upregulate the expression of NQO and HO-1 in vitro, though the expression of HO-1 was only observed in melanocytes (134). Nrf2 dependent induction of HO-1 has also been observed in an in vitro melanocyte cell line treated with hydrogen peroxide (135). Additionally, in vitro studies of human fibroblasts have positively identified Nrf2 mediated gene expression of HO-1 in the presence of UVA (136). Skin culture studies indicate that exposure to chronic levels of ambient UVA and UVB can cause ROS photoaging in both keratinocytes and fibroblasts, and verify differential expression of Nrf2 dependent phase II enzymes among cell types (94). In this study (94) keratinocytes demonstrated modulation in NQO expression, whereas fibroblasts showed upregulation in multiple Nrf2 regulated genes, including NQO, GST, and HO-1. These studies indicate that Nrf2 activity is most significant in deeper layers of skin tissue.
The differential expression of Nrf2 dependent antioxidant enzymes among layered tissues of the skin is coupled with varying degrees of Nrf2 mediated resistance to apoptosis (137, 138). In addition, the end point of enzyme induction versus cellular apoptosis depends on the type of UVR exposure (i.e. UVA, or UVB). Within fibroblast tissue, studies using Nrf2 knockout mice demonstrate that UVA more often induces the activation of Nrf2 mediated antioxidant enzymes, while exposure to UVB increases the propensity for apoptosis (138). Moreover, using transgenic mice containing constitutively active Nrf2 within keratinocytes (137), a gradient of Nrf2 mediated enzyme induction and apoptosis was noted across cross sections of epidermal tissue exposed to UVB. Essentially, in the deeper regions of the epidermis, UVB is more likely to induce apoptosis, whereas, the reliance of Nrf2 dependent antioxidants occurs more superficially (137).
The significance of differential activation of Nrf2 dependent antioxidant enzymes and apoptosis is relevant in the functional context of skin layering. In addition to the pronounced photoaging via the production of ROS by UVA and UVB (94, 139), UVB has been shown to be genotoxic to the epidermis, as observed by the accumulation of oxidative DNA products (140). While malignant cells can occur in either layer of the skin, the relative reduction of Nrf2 dependent antioxidant genes in epidermal keratinocytes (94, 137) is accounted for by the presence of pigment and the frequent renewal of new cells. Indeed, some protection from malignancy is required in the epidermis, as is evident in the gradient of apoptotic induction within the epidermis (137); however, the dermis receives greater amounts of harmful UVB and requires more robust antioxidant protection and safeguards to fight malignancy.
The emerging patterns of UVR mediated Nrf2 activity in mammalian skin is just one part of a complex and interactive response mechanism. Recent reports using in vitro human keratinocytes have implicated AhR –Nrf2 crosstalk as part of an anti-inflammatory response mechanism that can be activated by ROS induction of AhR, followed by subsequent downregulation of pro-inflammatory cytokines and Nrf2 dependent expression of NQO (141). Conversely, studies of Nrf2 knockout mice (142) have demonstrated that Nrf2 functions as part of a pro-inflammatory response in keratinocytes during wound healing. In this model, Nrf2 activity increased in the presence of ROS and growth factors, and induced the expression of HO-1 and GST. Also, the presence of Nrf2 provided positive feedback for the maintenance of pro-inflammatory cytokines, purportedly through a transcription complex with the tumor necrosis factors-α promotor (TNF). The occurrence of positive feedback to pro-inflammatory cytokines via Nrf2 is contrary to notions of Nrf2 as a transcription factor for antioxidants and anti-inflammatory response (141, 143); however, recent studies have demonstrated a positive relationship in the expression of Nrf2 and pro-inflammatory cytokines (144), and a transcription complex between Nrf2 and TNF (145). Although the AhR transcription factor was not investigated in the earlier Nrf2 knockout study (142), the potential for crosstalk exists, given the ROS signaling of AhR in keratinocytes (141).
UVR induced modulation lead to altered carcinogenicity of metabolites, increase oxidative stress, and the disruption of lipid metabolism (9, 104). UVA produced lipid peroxides have been also been shown to signal Nrf2 dependent antioxidants in keratinocytes and fibroblasts (93). Additionally, low density lipoproteins which are sources of oxidized lipids, are verified inducers of AhR in vascular tissue (146). This lipid based pathway of AhR activation is hypothesized to contribute to fatty liver disease and atherosclerosis (146, 147). Modulations of adipogenic processes have been demonstrated in mammalian fibroblasts through bidirectional crosstalk between Nrf2 and AhR (148). This study demonstrated that the Nrf2 pathway in mouse fibroblasts was shown to inhibit adipocyte differentiation in an AhR dependent manner. It is suspected that UVR activation of these two transcription factors may exasperate the photoaging process that reduces the lipid content of skin and contributes to adipogenic disease (i.e. dyslipidemia) (9).
The relatively extended timeframe of seasonal cycles challenge the assessment of UVR mediated physiological dysregulations across seasons. Nevertheless, given the threats of stratospheric ozone depletion are greatest during the spring season, short term excessive UVR exposures can potentially drive melatonin based hormonal shifts during critical life history stages associated with the spring season (49). Homeostatic shifts from nominal cycles of reproduction, growth and metabolism can potentially alter individual fitness. Furthermore, seasonally correlated effects of UVR exposure are likely to be co-linked with physiological responses to climate change, including abiotic shifts such as temperature and biotic shifts in ecosystem productivity (149, 150). Climate change may have deleterious modulations of immunity and increase the propensity for diseases that can impact physiological condition independent of UVR (150, 151, 152). Moreover, photoperiod plays a normal role in seasonal cycling of immune response genes (153) and may be modulated by UVR exposure through AhR and Nrf2.
Melatonin dependent signaling originating in the central nervous system (CNS) links the immune system across seasons and circadian cycles, suggesting that UVR mediated shifts in circadian homeostasis are likely, over and above seasonal dysregulations (153, 154). Environmental factors including UVR may cause rhythmic modulations of the inflammatory system that naturally cycles diurnally according to melatonin and cortisol levels, with subsequent regulation of pro and anti-inflammatory cytokine levels (154). Conversely, environmental stress factors can modulate expression of circadian genes such as Per (155, 156), potentiating the risk of tumorgeneis (155). Indeed, the propensity for cancer development and immunological modulations resulting from melatonin and circadian rhythm disruption are active areas of basic and clinical research (155, 157, 158). Circadian linked tumorogenesis is complex and not fully characterized; however, alterations of melatonin dependent cytokine expression (154, 158) and glucocorticoid dependent synchronization of peripheral clocks are emerging pathways of circadian linked cancers (157).
In mammals, cancers linked to circadian dysregulations may be attributed to the decoupling of circadian entrainment of cell cycles (157) that are purportedly coordinated for peripheral clock optimization (159). UVR induced modulations of this coordination system may target the linkage between circadian (Per, Timeless (Tim), cryptochrome and cell cycling proteins (Checkpoint proteins; Chk) (160, 161, 162). Additionally, in mammals, UVB can accelerate cellular proliferation by altering the phosphorylation status of cyclin-dependent kinase/cyclin complexes (cdk-4- cyclin D1, E, A) (163). Interestingly, studies indicate that in the absence of exogenous ligands, AhR dependent CYP1A1 maintains an integral proliferative role in the cell cycling of various tissues including keratinocytes (164), in an association with cyclins and retinoblastoma proteins (RB) (164, 165, 166), while TCDD activation of AhR induces cell cycle arrest in the G1 phase and inhibition of cellular apoptosis (165, 166). A tissue specific role of AhR in cell proliferation is apparent such as in the human lung, activated AhR promotes progression from G1 to the S phase following the PAS domain heterodimer AhR/Arnt complex interaction with the E2F1 regulatory domain (167), whereas no interaction was observed in liver cells. Additionally, AhR activation in human keratinocytes exposed to UVB causes cell proliferation by activation of EGFR and TNF (123, 168).
The mammalian expression of AhR dependent CYP (1A1, 1B1) genes in response to FICZ in vivo was first reported by Mukai and Tischkau (113), with concurrent data to suggest an endogenous role of AhR in the regulation of circadian rhythms, and possible pathway leading to an increased risk factor for cancer (113). These initial studies also reported a time dependent repression of CYP1A1, BMAL and Per, and Cry in suprachiasmatic nucleus (SCN) cells, in vitro (113) with the treatment of tryptophan photoproducts. Together these findings further validate light as a cue in the SCN and highlight the coactivation of AhR and circadian PAS genes including CLOCK, BMAL, and Per. Follow-up studies confirming interactions between circadian disruptions and AhR have primarily utilized treatments of TCDD, further implicating atypical time activation of PAS genes as a decoupling mechanism between master and peripheral clocks via modulations of Per gene expression (169, 170). Treatment of Hepa-1c1c7 cells in vitro and extracted mice liver cells treated in vivo with TCDD collectively demonstrate that AhR activation causes a downstream repression of Per1 through competitive heterodimerization of AhR/BMAL (versus CLOCK/BMAL) (170). The AhR/BMAL heterodimer inhibits binding to the Per Ebox promotor, thereby downregulating Per expression (170). Essentially the same observation was noted with TCDD treated mouse ovaries treated in vivo, where AhR interactions with BMAL caused a modulation of Per2 expression (169). Similarly, the circadian driven maturation of blood cells from stem cells was modulated by disruption of Per1 and Per2 after treatment with TCDD (171). Collectively, these studies reveal a pattern of circadian clock disruption, including a consistent repression of Period genes that may lead to various cancers.
Circadian signaling is tightly coupled to the redox state of cells, and studies suggest there is the likelihood for UVR mediated oxidative stress to modulate circadian rhythms and metabolic homeostasis through circadian genes (172, 173, 174) or redox sensitive metabolizing enzymes (68, 71). Mechanically, circadian genes including BMAL, CLOCK, and NPAS2 (homologue of CLOCK, with brain specific expression in mammals), function as redox sensors that respond to the redox state of nicotinamide adenine dinucleotide (NADH, reduced; NAD+, oxidized) (174). The reduced NADH induces the heterodimer complex of Clock or NPAS2 with BMAL, that subsequently upregulates expression of circadian Per and cryptochromes (71, 173). This circadian redox sensoring system is responsive to external inputs such as diet as a signal that providing feedback for circadian oscillators and the entrainment of peripheral clocks that translates to cycling of cellular metabolism (173). Repression of this metabolic/circadian system occurs through the oxidized NAD (173) or through negative feedback via Per and Cry (74). Interestingly, polymorphisms in NPAS2 have been linked to “Seasonal Affective Disorder” (172).
The occurrence of AhR dependent CYP expression among various signaling pathways that cycle in a circadian pattern suggests that UVR mediated oxidative stress may result in oxidative modulations of circadian patterns of fatty acid metabolism (175), cross talk with classical estrogen receptors (176), and development among both invertebrates (177) and vertebrate lineages (178). Lastly, melatonin is a strong antioxidant with potential interactions with Nrf2 (179), over and above its role in photoperiod signaling. Melatonin is UVR responsive in keratinocytes (180), yet the interplay of UVR, oxidative stress, melatonin and Nrf2 are poorly understood.
While food intake secondarily signals peripheral circadian oscillators that entrain circadian expression of metabolic enzyme expression, the metabolism of dietary intake is regulated by circadian rhythms (67, 68, 71, 79). This bidirectional feedback between circadian clocks and metabolism acts on multiple systems, including Nrf2 (131). In addition to the upregulation of the Nrf2 transcription factor in the presence of UVR mediated intracellular ROS and oxidative stress (88), Nrf2 is a major regulator of dietary metabolism (131), including fatty acid metabolism that can be modulated by oxidative stress (104). In fact, Nrf2 functions in an intestinal regulatory capacity during development that is hypothesized as the precursor to the detoxifying action of Nrf2 in response to oxidative stress (181). During the intestinal development of invertebrates, the Nrf2 (Cnc) homologs regulate stem cell proliferation and mesendodermal tissue differentiation through balancing cellular redox potential (181, 182). In larva teleost fishes, the expression of dietary enzymes has been shown to be modulated by exposure to UVB (183), suggesting that the normal circadian expression of Nrf2 may be modulated by altered UVB exposures (68), translating to genomic alterations of normal digestive patterns.
Finally, as an extension of the coupling between cyclic rhythms, metabolism, and their functional roles in the maintenance of metabolic homeostasis, suspected consequences on lifespan are emerging across broad taxonomic scales. The role of invertebrate Nrf2 homologs in regulating oxidative stress has been coupled with the capacity to determine lifespan (182, 184), and similar Nrf2 roles have been identified for vertebrate species (104, 131, 185). In addition to nutritional feedback and entrainment, the coupling between circadian cycles and metabolism are also correlated with feedback loops that rely on glucocorticoid signaling and nuclear receptors including, ROR (activator) and REV-ERB (repressor), which can reset circadian clocks, alter lipid metabolism, and exasperate the inflammatory system (67, 71, 186). Interestingly among humans, polymorphisms in the circadian Per gene system are linked to alterations in glucose metabolism (172), while aging in Drosophila is correlated with stress dependent accumulation of Per mutations (187). The relationships of circadian redox sensors, metabolism and aging are all the more interesting when considering the role of accumulated free radicals in the aging process. It is perhaps fruitful to speculate how UVR exposure may modulate these complex set of conceptual interrelationships.
While the Montreal Protocol has been a great achievement in Environmental Policy a real threat of detrimental exposure to UVR persists. Long term trends and implications of persistent ozone depletion must be considered in the context of global climate change as recent research has shown interactions between these two phenomena (23, 27). Naturally biochemical and hormonal signaling is driven by the sun, and in this context we have highlighted the potential for excessive UVR exposure to induce dysregulations of biological rhythmicity at the scale of seasons and circadian cycles. Current research in signal transduction has revealed newly discovered physiological targets of UVR exposure, including the AhR and Nrf2 transcription factors, which have been discussed herein as UVR targets of dysregulation. This review has attempted to demonstrate the convergence of the current hot topics of ozone depletion, climate change, and biological rhythmicity, with the strong potential for negative consequences for ecological integrity and human health. It is hopeful that further research in any one of these individual topics of study will consider results in a manner that integrates the interaction of different fields of study (i.e. ozone depletion, climate change, ecosystems in context of the former, organismal and human health in the context of the former).
Indeed, biological timing that is driven by solar cues is a ubiquitous trait across the taxonomic landscape and the consequences for anthropogenic mediated alterations in solar radiance reaching the earths are equally broad in scope. The coupling of physiological homeostasis and biological rhythmicity should not be underemphasized. UVR mediated imbalances can translate to changes in overall ecosystem landscape, loss of individual fitness (a marker of natural selection) due to temporal life history shifts, or health consequences such as cancer. Moreover, the AhR and Nrf2 transcription factors highlighted in this review are widely conserved across taxonomic groups, providing further evidence that UVR induced modulation of AhR/Nrf2 signaling may be targeted pathways of dysregulation throughout large portions of the taxonomic tree of life. For humans in particular, the AhR and Nrf2 pathways are ever emerging as routes of UVR-induced skin cancer, which remains a major health concern in this modern era of climate change and increased UVR exposure. In conclusion, these are worthy avenues of continued pursuit in an era of climate change and increased UVR exposure.
This project was supported by NOAA-Environmental Cooperative Science Center Grant # NA11SEC4810001-Sub Contract # 003499; NIH RCMI-Center for Environmental Health Grant # 5G12RR013459-15, and NIH NIMHD Grant # 8G12MD007581-15, at Jackson State University.