Pheomelanin Synthesis Research Articles

A review of therapies for hyperpigmentation modulating the synthesis of eumelanin to pheomelanin.

There are significant psychosocial burdens in patients with hyperpigmentation, which emphasizes the importance of treatment. Current gold standard for treatment is hydroquinone; however, alternatives have been developed given the concern for side effects of hydroquinone. Melanogenesis is responsible for the production of eumelanin and pheomelanin; there are many factors that will determine whether eumelanin or pheomelanin will be produced. Eumelanin is known for its photoprotective qualities, while pheomelanin is implicated in photocarcinogenesis and photoaging. Multiple treatment modalities for hyperpigmentation that shift eumelanin to pheomelanin synthesis exist. Cysteamine, glutathione, kojic acid, and methyl sulfonyl methane are four agents used to treat hyperpigmentation by shifting the production of eumelanin to pheomelanin. It is critical to discuss photoprotection with patients to help reduce the potential impact of increased pheomelanin production and to expand research in this area.

Oxidative Challenges Do Not Impact Pheomelanin-Dependent Coloration in Male Japanese Quails.

Colorful traits play an important role in animal communication. Melanin-based colorations are the most extended color traits in animals and are produced by two types of endogenous melanic pigments: eumelanins and pheomelanins, the last ones being the least studied in the context of communication. The production of pheomelanin requires a semi-essential amino acid, cysteine, which is also used for the synthesis of an important endogenous antioxidant, glutathione. Hence, it has been proposed that the synthesis of pheomelanin and glutathione may compete for the cysteine available in the organism. In that case, pheomelanic colorations are predicted to be less intense when the individual is facing an oxidative challenge, and therefore, they would provide information on the oxidative status of the bearer. Here, we experimentally evaluated this hypothesis using male Japanese quails (Coturnix japonica) as a model of study, a species with pheomelanin-based plumage in the breast and cheeks. During feather growth, individuals were exposed to one of three possible conditions: Control (saline), an endogenous oxidative challenge (Escherichia coli lipopolysaccharide injections), or an exogenous oxidative challenge (paraquat injections). Contrary to predictions, we found that: (1) Birds from the three groups exhibited less intense pheomelanic colorations in feathers after the experimental manipulation, and the magnitude of this change did not differ among groups. (2) There was no effect of the experimental treatments on the proportion reduced/oxidized glutathione, an index of oxidative status. (3) Lipid peroxidation was lower after the experimental manipulation, with birds exposed to the paraquat challenge experiencing a stronger decline than other groups. (4) Cysteine and total glutathione levels decreased after the experimental manipulation, with no differences per group in the magnitude of the decline. Taken together the results do not support the hypothesis that oxidative status plays a key role at determining the variation in the intensity of pheomelanic colorations.

The "Unconventional" Effect of Cysteine on the In Vitro Synthesis of Melanin.

This report details some of our observations regarding the impact of cysteine on the air-mediated oxidation of catecholamines, particularly epinephrine. The intent was to synthesize light-colored, pheomelanin-like materials. Pheomelanin is commonly described as a material generated from a mixture of catecholamines and cysteine. However, we observed that (1) the presence of cysteine resulted in a concentration-dependent delay in the onset of color formation and (2) the presence of cysteine resulted in darker, more eumelanin-like materials. These effects were particularly impactful in the case of epinephrine. More elaborate studies involving other amino acids or scaled-up reactions were conducted with epinephrine as the precursor. These studies show that other amino acids, e.g., methionine or serine, could lead to darker materials, but none were as impactful as cysteine. Although our results are in contrast to typical descriptions regarding the impact of cysteine on the synthesis of melanin, they may reflect crucial differences between the in vitrovsin vivo synthesis of pheomelanin.

Island volcanism predicts pheomelanin‐based plumage colouration in a cosmopolitan raptor

AbstractAimPheomelanin is a pigment responsible for yellowish‐to‐reddish colours of vertebrate teguments. Its biosynthesis is favoured under high concentration of intracellular thiols, which, in turn, can depend on the environmental exposure to sulphur. Thus, pheomelanin production should be more intense and frequent in environments characterized by high level of sulphur, such as volcanic regions. In this study, we aimed at addressing this hypothesis by investigating variation in plumage colour of insular populations of the cosmopolitan barn owl (Tyto alba species complex) according to the presence of soils of volcanic origin (i.e. andosols) and recent volcanic activity.LocationWorld.TaxonBarn owl species complex.MethodsWe measured plumage colouration of more than 2000 museum specimens from 50 islands and archipelagos worldwide. We then compared plumage colouration between populations living on volcanic (i.e. where andosols and/or recent volcanic activity are present) and non‐volcanic islands/archipelagos.ResultsConsistently with the prediction, plumage colouration is significantly darker (i.e. pheomelanic) on islands/archipelagos where andosols and/or recent volcanic activity are present than absent, although this environmental factor explains a small fraction of plumage colour variability across islands (<10%). Similar results were obtained when specimens' sex and climatic predictors were included in the analyses.Main ConclusionsBecause excessive intracellular levels of thiols can be toxic, pheomelanin synthesis may function as a mechanism keeping these compounds below the toxicity threshold and limiting their detrimental effects on physiology. Darker plumage may also be favoured because it promotes background matching against the dark environment typical of volcanic islands (dense vegetation cover and/or dark soil). Our results add to the little evidence that the environmental exposure of compounds that are involved in melanin biosynthesis can affect animal pigmentation and suggest that soil composition may be a factor that affects melanogenesis, possibly contributing to generate spatial variation in pheomelanin‐based traits in animals.

Transcription factor CgPOU3F4-like regulates expression of pheomelanin synthesis related gene CgB-aat1 in the Pacific oyster (Crassostrea gigas)

Previous study has found that b (0, +) -type amino acid transporter 1 (CgB-aat1) plays an essential role on mantle pigmentation in the Pacific oyster Crassostrea gigas. However, the molecular regulation of CgB-aat1 gene expression remains unclear. Herein, three POU domain family members, CgPOU2F1, CgPOU3F4-like and CgPOU4F3-X1 were characterized and they all had POUs and HOX domains, respectively, which were important in transcriptional regulation. CgPOU3F4-like gene expression was the highest in mantle edge. Subsequently, the dual-luciferase reporter result showed that the core regulatory region of CgB-aat1 gene was from −632 to −350 bp of promoter. In transient co-transfection assays, the strongest activity was activated only by CgPOU3F4-like, suggesting CgPOU3F4-like was a valid transcriptional activator of CgB-aat1 gene promoter. And the structural integrity of CgPOU3F4-like was essential for its activation function. In addition, site directed mutagenesis assay was applied to detect three key binding sites between CgPOU3F4-like and core region of CgB-aat1 gene promoter, and this interaction was verified by ChIP test. Furthermore, CgPOU3F4-like knockdown by RNA interference led to obvious decreases in CgB-aat1 and cystathionine beta-synthase (CgCbs) expressions at both mRNA and protein levels. Collectively, these results indicate that CgPOU3F4-like positively regulate CgB-aat1 gene expression and it may be a critical upstream transcriptional regulation factor in pheomelanin synthesis in C. gigas.

Synthesis and structural characteristics analysis of melanin pigments induced by blue light in Morchella sextelata.

Morchella sextelata, a highly sought-after edible mushroom worldwide, is evaluated based on its cap color as an essential commercial property indicator. In the present study, the effects of blue light on cap pigmentation in M. sextelata, as well as the synthesis and structural characteristics of melanin pigments within the cap were examined. The results showed that an increase in the proportion of blue light within the lighting environment promoted melanin synthesis and melanization of the cap. Transmission and scanning electron microscopy revealed the localization of melanin within the mycelium and its ultrastructural characteristics. The UV-visible analysis demonstrated that melanin exhibited a maximum absorption peak at 220 nm and possessed high alkaline solubility as well as acid precipitability. The structural characteristics of melanin were analyzed using FTIR, NMR, HPLC, and elemental analysis, which confirmed the presence of eumelanin, pheomelanin, and allomelanin in both brown and black caps. Furthermore, blue light can stimulate the synthesis of both eumelanin and pheomelanin. The obtained results can serve as the foundation for comprehending the mechanism by which light regulates color formation in mushrooms.

Are Plants Capable of Pheomelanin Synthesis? Gas Chromatography/Tandem Mass Spectrometry Characterization of Thermally Degraded Melanin Isolated from Echinacea purpurea

Echinacea is a widely used plant medicine, valued especially for its well-documented ability to stimulate the immune system. It has been suggested that melanin could be one of the bioactive factors responsible for the immunostimulatory properties of the plant. The biological functions of melanin pigments are closely related to their chemical composition and structural features. The aim of this study was to characterize the melanin from Echinacea purpurea based on the analysis of thermal degradation products of the well-purified pigment extracted from the dried herb. The melanin was pyrolyzed, and the resulting products were separated by gas chromatography and identified using a triple quadrupole mass spectrometer operating in full scan and multiple reaction monitoring modes. Three groups of marker products were detected in the melanin pyrolysate: polyphenol derivatives, nitrogen-containing heterocycles devoid of sulfur, and benzothiazines/benzothiazoles. This suggests that E. purpurea produces three structurally different melanin pigments: allomelanin, eumelanin, and pheomelanin, which in turn may affect the biological activity of the herb. Our results provide the first-ever evidence that plants are capable of synthesizing pheomelanin, which until now, has only been described for representatives of the animal and fungal kingdoms.

Involvement of B-aat1 and Cbs in regulating mantle pigmentation in the Pacific oyster (Crassostrea gigas).

Shell color formation is an important physiological process in bivalves, the molecular genetic basis has potential application in bivalve aquaculture, but there is still remaining unclear about this issue. The cystine/glutamate transporter (Slc7a11) and cystathionine beta-synthase (Cbs) are integral genes in pheomelanin synthesis pathway, which is vital to skin pigmentation. Here, the sequences of b (0, +) -type amino acid transporter 1 (B-aat1) and Cbs in Pacific oyster (Crassostrea gigas) (CgB-aat1, CgCbs) were characterized. Phylogenetically, the deduced amino acid sequences of CgB-aat1 and CgCbs both possessed conserved features. Genes were both ubiquitously expressed in six tested tissues with more abundant expression level in central mantle. Besides, the polyclonal antibodies of CgB-aat1, CgCbs, CgTyr, and CgTyrp2 were successfully prepared. Immunofluorescence analysis revealed that CgB-aat1 and CgCbs proteins were both expressed in gill rudiments of eyed-larvae and concentrated mainly in cytoplasm of epithelial cell and nerve axons in mantle. Additionally, after CgB-aat1 or CgCbs silencing, expressions at mRNA and protein levels of CgB-aat1 and CgCbs involved in pheomelanin synthesis were significantly suppressed, and CgTyr, CgTyrp1 and CgTyrp2 related to eumelanin synthesis were also down-regulated but no apparent differences, respectively. Moreover, micrographic examination found less brown-granules at mantle edge in CgB-aat1 interference group. These results implied that pheomelanin synthesis was possible induced by CgB-aat1-CgTyr-CgCbs axis, and it played an essential role on mantle pigmentation in the oysters. These findings provide the useful genetic knowledge and enrich the physiological information for the shell color formation in bivalve aquaculture.

Metabolic Basis and Clinical Evidence for Skin Lightening Effects of Thiol Compounds.

Melanin pigment is a major factor in determining the color of the skin, and its abnormal increase or decrease can cause serious pigmentation disorders. The melanin pigment of the skin is divided into light pheomelanin and dark eumelanin, and a big difference between them is whether they contain sulfur. Melanin synthesis starts from a common reaction in which tyrosine or dihydroxyphenylalanine (DOPA) is oxidized by tyrosinase (TYR) to produce dopaquinone (DQ). DQ is spontaneously converted to leukodopachrome and then oxidized to dopachrome, which enters the eumelanin synthesis pathway. When DQ reacts with cysteine, cysteinyl dopa is generated, which is oxidized to cysteinyl DQ and enters the pheomelanin synthesis pathway. Therefore, thiol compounds can influence the relative synthesis of eumelanin and pheomelanin. In addition, thiol compounds can inhibit enzymatic activity by binding to copper ions at the active site of TYR, and act as an antioxidant scavenging reactive oxygen species and free radicals or as a modulator of redox balance, thereby inhibiting overall melanin synthesis. This review will cover the metabolic aspects of thiol compounds, the role of thiol compounds in melanin synthesis, comparison of the antimelanogenic effects of various thiol compounds, and clinical trials on the skin lightening efficacy of thiol compounds. We hope that this review will help identify the advantages and disadvantages of various thiol compounds as modulators of skin pigmentation and contribute to the development of safer and more effective strategies for the treatment of pigmentation disorders.

Oxidative stress in the skin: Impact and related protection.

Skin, our first interface to the external environment, is subjected to oxidative stress caused by a variety of factors such as solar ultraviolet, infrared and visible light, environmental pollution, including ozone and particulate matters, and psychological stress. Excessive reactive species, including reactive oxygen species and reactive nitrogen species, exacerbate skin pigmentation and aging, which further lead to skin tone unevenness, pigmentary disorder, skin roughness and wrinkles. Besides these, skin microbiota are also a very important factor ensuring the proper functions of skin. While environmental factors such as UV and pollutants impact skin microbiota compositions, skin dysbiosis results in various skin conditions. In this review, we summarize the generation of oxidative stress from exogenous and endogenous sources. We further introduce current knowledge on the possible roles of oxidative stress in skin pigmentation and aging, specifically with emphasis on oxidative stress and skin pigmentation. Meanwhile, we summarize the science and rationale of using three well-known antioxidants, namely vitamin C, resveratrol and ferulic acid, in the treatment of hyperpigmentation. Finally, we discuss the strategy for preventing oxidative stress-induced skin pigmentation and aging.

Chemical and biochemical control of skin pigmentation with special emphasis on mixed melanogenesis.

Melanins are widely distributed in animals and plants; in vertebrates, most melanins are present on the body surface. The diversity of pigmentation in vertebrates is mainly attributed to the quantity and ratio of eumelanin and pheomelanin synthesis. Most natural melanin pigments in animals consist of both eumelanin and pheomelanin in varying ratios, and thus, their combined synthesis is called "mixed melanogenesis." Gene expression is an established mechanism for controlling melanin synthesis; however, there are multiple factors that affect melanin synthesis besides gene expression. Due to the differential sensitivity of the eumelanin and pheomelanin synthetic pathways to pH, melanosomal pH likely plays a major role in mixed melanogenesis. Here, we focused on various factors affecting mixed melanogenesis including (1) chemical regulation of melanin synthesis, (2) melanosomal pH regulation during normal melanogenesis and effect on mixed melanogenesis, and (3) mechanisms of melanosomal pH control (proton pumps, channels, transporters, and signaling pathways).

Slc7a11 downregulation is rapidly reversed after cessation of competitive social stress in zebra finches.

Gene expression can be modulated by epigenetic modifications, which may lead to a rapid adaptation to environmental stress. After stress cessation, changes in gene expression could be reversed, which would allow organisms to maintain their phenotype under transient environments, but this mechanism is poorly understood. Social stress downregulates a gene directly involved in pheomelanin synthesis (Slc7a11) by changing DNA m5C levels, avoiding cellular damage caused by stress. We thus investigated if Slc7a11 expression is reversed in melanocytes of growing flank feathers to avoid changes in the pigmentation phenotype. We measured the expression level of Slc7a11 at three time points: before stress exposure, immediately after stress exposure and six weeks after stress cessation in 37 male zebra finches (Taeniopygia guttata). No differences in Slc7a11 expression were detected between birds exposed to stress and controls six weeks after stress elimination, indicating that stress removal led to a cessation of Slc7a11 downregulation. Reversibility in Slc7a11 expression, probably mediated by reversible changes in DNA methylation, may thus avoid altering the pigmentation phenotype during transient stressful conditions. This is one of the few studies in vertebrates supporting the idea that reversible gene expression responses allow organisms adapting to changing environmental conditions.

Estimating the copy number of the agouti signaling protein (ASIP) gene in goat breeds with different color patterns

The agouti signaling protein (ASIP) gene has a crucial role in pigmentation by encoding a protein that binds the melanocortin 1 receptor and stimulates the synthesis of pheomelanin rather than eumelanin. Several studies have suggested that an increased copy number of the ASIP gene might explain the white pigmentation of certain goat breeds, as previously demonstrated in sheep. In the current work, we have identified the segregation of the ASIP CNV in Murciano-Granadina (black or brown coat), Malagueña (brown, blond or white coat) and Saanen (white coat) goats with available Illumina Goat SNP50 BeadChip (Illumina Inc., San Diego, CA) data by using the PennCNV v1.0.5 and QuantiSNP v2 tools. This result shows that the ASIP CNV segregates in non-white breeds. To gain new insights into this issue, we have estimated the copy number of the ASIP gene in 83 goats from 8 breeds with different coloration patterns using a real-time quantitative PCR approach. Our results showed an increased ASIP copy number not only in Saanen (3.50 ± 0.23 copies relative to the calibrator) and white Malagueña (3.51 ± 0.51 copies) goats, but also in the Murciano-Granadina breed (3.33 ± 0.58 copies) as well as in blond/brown individuals from the Malagueña (3.58 ± 0.73 copies) breed. The number of ASIP copies was not significantly different in these four caprine populations (P > 0.05). Moreover, we did not observe a trend towards increased ASIP copy number in breeds with predominantly white colors, such as Maltese (2.85 ± 0.28 copies), Jonica (2.82 ± 0.39 copies) and Blanca de Rasquera (2.37 ± 0.33 copies). Our results, combined with recent findings demonstrating the high structural complexity of the ASIP locus, indicate that additional functional and expression studies should be performed in order to fully understand the role of ASIP structural variation in goat pigmentation.

Measurement of Melanin Metabolism in Live Cells by [U-13C]-L-Tyrosine Fate Tracing Using Liquid Chromatography-Mass Spectrometry

Melanin synthesis occurs within a specialized organelle called the melanosome. Traditional methods for measuring melanin levels rely on the detection of chemical degradation products of melanin by high-performance liquid chromatography (HPLC). Although these methods are robust, they are unable to distinguish between melanin synthesis and degradation, and are best suited to measure melanin changes over long periods of time. We developed a method that actively measures both eumelanin and pheomelanin synthesis by fate tracing [U-13C] tyrosine using liquid chromatography-mass spectrometry (LC-MS). Using this method, we confirmed previous reports of melanin synthesis differences between melanocytes derived from individuals with different skin color or MC1R genotype, and uncovered new information regarding the differential de novo synthesis of eumelanin and pheomelanin, also called mixed melanogenesis. We also revealed that distinct mechanisms that alter melanosomal pH differentially induce new eumelanin and pheomelanin synthesis. Finally, we revealed that the synthesis of L-DOPA, an important metabolite of tyrosine, is differentially controlled by multiple factors. Because tyrosine fate tracing is compatible with untargeted LC-MS based metabolomics, this approach enables the broad measurement of cellular metabolism in combination with melanin metabolism, and we anticipate that this approach will shed new light on multiple mechanisms of melanogenesis.

MFSD12 mediates the import of cysteine into melanosomes and lysosomes.

Dozens of genes contribute to the vast variation in human pigmentation. Many of these encode proteins that localize to the melanosome, the lysosome-related organelle that synthesizes pigment, but have unclear functions 1,2. Here, we describe the MelanoIP method for rapidly isolating melanosomes and profiling their labile metabolite contents. We use it to study MFSD12, a transmembrane protein of unknown molecular function that when suppressed causes darker pigmentation in mice and humans 3,4. We find that MFSD12 is required to maintain normal levels of cystine, the oxidized dimer of cysteine, in melanosomes, and to produce cysteinyldopas, the precursors of pheomelanin synthesis made in melanosomes via cysteine oxidation 5,6. Tracing and biochemical analyses show that MFSD12 is necessary for the import of cysteine into melanosomes, and, in non-pigmented cells, lysosomes. Indeed, loss of MFSD12 reduced the accumulation of cystine in lysosomes of fibroblasts from patients with cystinosis, a lysosomal storage disease caused by inactivation of the lysosomal cystine exporter CTNS (Cystinosin)7–9. Thus, MFSD12 is an essential component of the long-sought cysteine importer for melanosomes and lysosomes.

Transcriptome analysis of feather follicles reveals candidate genes and pathways associated with pheomelanin pigmentation in chickens

Yellow plumage is common in chickens, especially in breeds such as the Huiyang Bearded chicken, which is indigenous to China. We evaluated plumage colour distribution in F1, F2, and F3 populations of an Huiyang Bearded chicken × White Leghorn chicken cross, the heredity of the yellow plumage trait was distinguished from that of the gold plumage and other known plumage colours. Microscopic analysis of the feather follicles indicated that pheomelanin particles were formed in yellow but not in white feathers. To screen genes related to formation of the pheomelanin particles, we generated transcriptome data from yellow and white feather follicles from 7- and 11-week-old F3 chickens using RNA-seq. We identified 27 differentially expressed genes (DEGs) when comparing the yellow and white feather follicles. These DEGs were enriched in the Gene Ontology classes ‘melanosome’ and ‘melanosome organization’ related to the pigmentation process. Down-regulation of TYRP1, DCT, PMEL, MLANA, and HPGDS, verified using quantitative reverse transcription PCR, may lead to reduced eumelanin and increased pheomelanin synthesis in yellow plumage. Owing to the presence of the Dominant white locus, both white and yellow plumage lack eumelanin, and white feathers showed no pigments. Our results provide an understanding of yellow plumage formation in chickens.

Differential influence of Slc7a11 expression and body condition on pheomelanin‐based pigmentation in two Eurasian nuthatch Sitta europaea populations with different predation risk

The expression of the gene Slc7a11 promotes the antioxidant capacity of cells by providing them with cysteine that can be used for the synthesis of glutathione (GSH), the most important intracellular antioxidant. In melanocytes, intracellular cysteine can also enter melanosomes and get incorporated in the pheomelanin pigment synthesis pathway, thus decreasing cysteine availability for GSH synthesis and potentially creating chronic oxidative stress. We thus hypothesized that a mechanism limiting the use of intramelanocytic cysteine for pheomelanin synthesis in environmental conditions generating oxidative stress may be physiologically advantageous and favored by natural selection. Here we searched for evidence of such a mechanism by comparing the influence of melanocytic Slc7a11 expression on pheomelanin‐based pigmentation in developing Eurasian nuthatch Sitta europaea nestlings from two populations differing in predation risk, a natural source of oxidative stress. Pheomelanin synthesis and pigmentation tended to increase with Slc7a11 expression in the low‐risk population as expected from the activity of this gene, but decreased with Slc7a11 expression in the high‐risk population. The same was not observed in the expression of five other genes influencing pheomelanin synthesis without affecting cysteine availability in melanocytes. The influence of body condition on the intensity of pheomelanin‐based pigmentation also differed between populations, being positive in the low‐risk population and negative in the high‐risk population. The resulting pigmentation of birds was more intense in the high‐risk population. These findings suggest that birds perceiving high predation risk may limit the use of cysteine for pheomelanin synthesis, which becomes independent of Slc7a11 expression. Some birds may have thus evolved the ability to adjust their pigmentation phenotype to environmental stress.

Impairment of mixed melanin-based pigmentation in parrots.

Parrots and allies (Order Psittaciformes) have evolved an exclusive capacity to synthesize polyene pigments called psittacofulvins at feather follicles, which allows them to produce a striking diversity of pigmentation phenotypes. Melanins are polymers constituting the most abundant pigments in animals, and the sulphurated form (pheomelanin) produces colors that are similar to those produced by psittacofulvins. However, the differential contribution of these pigments to psittaciform phenotypic diversity has not been investigated. Given the color redundancy, and physiological limitations associated to pheomelanin synthesis, we hypothesized that the latter would be avoided by psittaciform birds. Here we test this by using Raman spectroscopy to identify pigments in feathers exhibiting colors suspicious of being produced by pheomelanin (i.e., dull red, yellow and grey- and green-brownish) in 26 species from the three main lineages of Psittaciformes. We detected the non-sulphurated melanin form (eumelanin) in black, grey and brown plumage patches, and psittacofulvins in red, yellow and green patches, but no evidence of pheomelanin. As natural melanins are assumed to be composed of eumelanin and pheomelanin in varying ratios, our results represent the first report of impairment of mixed melanin-based pigmentation in animals. Given that psittaciforms also avoid the uptake of circulating carotenoid pigments, these birds seem to have evolved a capacity to avoid functional redundancy between pigments, likely by regulating follicular gene expression. Ours study provides the first vibrational characterization of different psittacofulvin-based colors and thus helps to determine the relative polyene chain length in these pigments, which is related to their antireductant protection activity.

A source of exogenous oxidative stress improves oxidative status and favors pheomelanin synthesis in zebra finches

Some organisms can modulate gene expression to trigger physiological responses that help adapt to environmental stress. The synthesis of the pigment pheomelanin in melanocytes seems to be one of these responses, as it may contribute to cellular homeostasis. We experimentally induced environmental oxidative stress in male zebra finches Taeniopygia guttata by the administration of the herbicide diquat dibromide during feather growth to test if the expression of genes involved in pheomelanin synthesis shows epigenetic lability. As pheomelanin synthesis implies decreasing the availability of the main cellular antioxidant (glutathione), it is expected to cause oxidative stress unless a protective mechanism limits pheomelanin synthesis and thus favors the antioxidant capacity. However, diquat exposure did not only improve the antioxidant capacity of birds, but also upregulated the expression of a gene (AGRP) that promotes pheomelanin synthesis in feather melanocytes, leading to the development of darker plumage coloration. No changes in the expression of other genes involved in pheomelanin synthesis (Slc7a11, Slc45a2, MC1R, ASIP and CTNS) were detected. DNA methylation levels only changed in MC1R, suggesting that epigenetic modifications other than changes in methylation may regulate AGRP expression lability. Our results suggest that exogenous oxidative stress induced a hormetic response that enhanced the oxidative status of birds and, consequently, promoted pheomelanin-based pigmentation, supporting the idea that birds adjust pheomelanin synthesis to their oxidative stress conditions.

Juvenile pheomelanin‐based plumage coloration has evolved more frequently in carnivorous species

Distinctive pheomelanin‐based plumage coloration in juvenile birds has been proposed as a signal of immaturity to avoid aggression by older conspecifics, but recent findings suggest a detoxifying strategy. Pheomelanin synthesis implies the consumption of cysteine, a semi‐essential amino acid that is necessary for the synthesis of the antioxidant glutathione (GSH) but that may be toxic if in excess in the diet. As the nestling stage probably represents a low‐stress period with limited requirement for GSH protection, the synthesis of pheomelanin in developing birds may help to maintain cysteine homeostasis, particularly in species with a high content of protein in the diet (i.e. carnivores). Here we confirm this hypothesis showing that, among 53 species of Western Palaearctic birds, juvenile pheomelanin‐based coloration has evolved more frequently in strictly carnivorous species than in species with other diets.