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Simple Summary: At birth, wild boars and their hybrid offspring usually have camouflage coat colors with juvenile stripes (the black/dark longitudinal stripes separated by yellow/light ones) on their back and two side flanks, and the juvenile stripes disappear at 4–5 months old. To study the mechanism of juvenile stripe maintenance, piglets with juvenile stripes were produced by crossing a wild boar with three Duroc sows. Through picture recording and pigment extraction, we observed that the stripe tended to disappear within approximately 70 days. Furthermore, whole-transcriptome comparison between two adjacent stripes (dark and light) on the back revealed that ZIC4, ssc-miR-532-3p and ENSSSCG00000056225 might play a role in the formation of stripes. The present study laid the foundation for future research on the molecular mechanisms underlying juvenile stripes. Coloration is a crucial trait that allows species to adapt and survive in different environments. Wild boars exhibit alternating black (dark) and yellow (light) longitudinal stripes on their back during their infancy (juvenile stripes), and as adults, they transform into uniform wild-type coat color. Aiming to record the procedure of juvenile stripes disappearing, piglets (WD) with juvenile stripes were produced by crossing a wild boar with Duroc sows, and photos of their coat color were taken from 20 d to 220 d. The pigments in the hairs from the black and yellow stripes were determined. Furthermore, the differentially expressed genes between the black and yellow stripes were investigated in 5 WD with the age of 30 d using whole-transcriptome sequencing to explore the genetic mechanism of the juvenile stripes. The juvenile stripes started to disappear at about 70 d, and stripes were not distinguished with the naked eye at about 160 d; that is, the juvenile stripe completely disappeared. A hotspot of a differentially expressing (DE) region was found on chromosome 13, containing/covering 2 of 13 DE genes and 8 of 10 DE lncRNAs in this region. A network among ZIC4, ssc-miR-532-3p, and ENSSSCG00000056225 might regulate the formation of juvenile stripes. Altogether, this study provides new insights into spatiotemporal coat color pattern. [ABSTRACT FROM AUTHOR]

Introduction1: Sheep are the main source of wool and its fiber characteristics, such as diameter, length, and color, which are determined by genetics and environmental factors, are key features in the economic value of sheep wool. In sheep, white wool has the highest economic value due to its dyeability. Therefore, identifying the mechanisms responsible for coat color determination is very important from an economic perspective. In general, the coat color is determined based on the amounts and types of melanin produced and released by the melanocytes in the skin tissue (Ito et al., 2000). The genetic basis and genes involved in coat color are well understood in rodents, although many of these genes are incorporated in coat color regulation in other species; including Sheep also have a common role. In Iran, the Lori Bakhtiari sheep is one of the most important breeds of sheep in terms of the use of its wool in the textile and carpet industries. In his breed, the dominant coat of the wool is white, although sometimes a percentage of dark brown and pale brown is also observed (Saadat Nouri, M. & Siah Mansour, 1368). However, in this breed, some animals have black spots on their coat, which leads to a decrease in wool quality. Since MC1R, ASIP, KLF4 and MITF genes play an important role in controlling and determining coat color in mammals, the purpose of this study was to investigate the expression of these genes in two phenotypes of white and black spots in the skin tissue of Lori Bakhtiari sheep. Materials and Methods: Skin samples were obtained from both white and dark parts of 14 white-coated sheep with black spots and total RNA was extracted. The quality and quantity of extracted RNAs were evaluated by agarose gel electrophoresis and spectrophotometer. Extracted RNA samples were exposed to DNase1 enzyme digestion to remove the possible contamination of genomic DNA. Also, the quality of synthesized cDNA was evaluated using 1% agarose gel. In this research, in order to amplify a fragment of the studied genes, using the mRNA sequence of these genes in the GenBank database, appropriate primers were designed by Primer3plus software. To evaluate the relative expression of the target genes, β-actin and GAPDH genes were used as reference genes to normalize the data. Finally, BestKeeper and REST 2009 V2.0.13 software were used for the analysis of gene expression data. Results and Discussion: Based on the descriptive results of Ct values, MC1R and MITF genes revealed minimum and maximum expression stability among the target genes in skin samples with standard deviations of 1.34 and 3.62, respectively. In addition, the reference genes (β-actin and GAPDH) showed the highest stability among all the studied genes. No significant differences were observed in mRNA levels of MC1R, ASIP, KLF4, and MITF genes in the spotted skin tissue compared to the white part of the skin (p>0.05). However, the expression of the ASIP gene was more than 2 times in the spotted part compared to the white skin, but this difference was not significant (p=0.21). In addition, the MC1R gene showed minimum expression differences in black spots and white parts of the skin tissue. In addition, the MITF and MC1R genes showed the highest and lowest levels of expression in skin samples of Lori Bakhtiari breed sheep with average Ct of 25.86 and 30.42, respectively. However, among all the studied genes, the lowest mRNA level was observed for the GAPDH gene with an average Ct of 35.96. Conclusion: Mammalian coat color results from various factors such as the degree and distribution of melanin pigment and the interaction between genotype and environment (B. Li et al., 2018). In addition, melanogenesis is a complex process that includes melanocyte growth, melanosome formation, melanin synthesis, melanin transport, and melanosome release (Ito & Wakamatsu, 2011). According to the conducted studies, a large number of genes are involved in the mechanism of coat color determination, but two genes, MC1R and ASIP, play an essential role in the regulation and control of coat color (Searle, 1968). In our study, no significant difference was observed in the expression of MC1R, ASIP, KFL4, and MITF genes in the spotted compared to the white part of the skin tissue in Lori Bakhtiari sheep. These results showed that the development of skin spots is not under the control of the studied genes in Lori Bakhtiari sheep, and genes or other factors can play roles in the creation of dark spots in this breed. [ABSTRACT FROM AUTHOR]

Introduction: Sheep are the main source of wool and its fiber characteristics, such as diameter, length, and color, which are determined by genetics and environmental factors, are key features in the economic value of sheep wool. In sheep, white wool has the highest economic value due to its dyeability, thus, the identification of mechanisms responsible for coating color determination is very important from an economic point of view. In general, the coat color is determined based on the amounts and types of melanin produced and released by the melanocytes in the skin tissue (Ito et al., 2000). The genetic basis and genes involved in coat color are well understood in rodents, although many of these genes are incorporated in coat color regulation in other species; including Sheep also have a common role. In Iran, the Lori Bakhtiari sheep is one of the most important breeds of sheep in terms of the use of its wool in the textile and carpet industries. In his breed, the dominant coat of the wool is white, although sometimes a percentage of dark brown and pale brown is also observed (Saadat Nouri, M. & Siah Mansour, 1368). However, in this breed, some animals have black spots on their coat, which leads to a decrease in wool quality. Since MC1R, ASIP, KLF4 and MITF genes play an important role in controlling and determining coat color in mammals, the purpose of this study was to investigate the expression of these genes in two phenotypes of white and black spots in the skin tissue of Lori Bakhtiari sheep.Materials and Methods: Skin samples were obtained from both white and dark parts of 14 white-coated sheep with black spots and total RNA was extracted. The quality and quantity of extracted RNAs were evaluated by agarose gel electrophoresis and spectrophotometer. Extracted RNA samples were exposed to DNase1 enzyme digestion to remove the possible contamination of genomic DNA. Also, the quality of synthesized cDNA was evaluated using 1% agarose gel. In this research, in order to amplify a fragment of the studied genes, using the mRNA sequence of these genes in the GenBank database, appropriate primers were designed by Primer3plus software. To evaluate the relative expression of the target genes, β-actin and GAPDH genes were used as reference genes to normalize the data. Finally, BestKeeper and REST 2009 V2.0.13 software were used for the analysis of gene expression data.Results and Discussion: Based on the descriptive results of Ct values, MC1R and MITF genes revealed minimum and maximum expression stability among the target genes in skin samples with standard deviations of 1.34 and 3.62, respectively. In addition, the reference genes (β-actin and GAPDH) showed the highest stability among all the studied genes. No significant differences were observed in mRNA levels of MC1R, ASIP, KLF4, and MITF genes in the spotted skin tissue compared to the white part of the skin (p>0.05). However, the expression of the ASIP gene was more than 2 times in the spotted part compared to the white skin, but this difference was not significant (p=0.21). In addition, the MC1R gene showed minimum expression differences in black spots and white parts of the skin tissue. In addition, the MITF and MC1R genes showed the highest and lowest levels of expression in skin samples of Lori Bakhtiari breed sheep with average Ct of 25.86 and 30.42, respectively. However, among all the studied genes, the lowest mRNA level was observed for the GAPDH gene with an average Ct of 35.96.Conclusion: Mammalian coat color results from various factors such as the degree and distribution of melanin pigment and the interaction between genotype and environment (B. Li et al., 2018). In addition, melanogenesis is a complex process that includes melanocyte growth, melanosome formation, melanin synthesis, melanin transport, and melanosome release (Ito & Wakamatsu, 2011). According to the conducted studies, a large number of genes are involved in the mechanism of coat color determination, but two genes, MC1R and ASIP, play an essential role in the regulation and control of coat color (Searle, 1968). In our study, no significant difference was observed in the expression of MC1R, ASIP, KFL4, and MITF genes in the spotted compared to the white part of the skin tissue in Lori Bakhtiari sheep. These results showed that the development of skin spots is not under the control of the studied genes in Lori Bakhtiari sheep, and genes or other factors can play roles in the creation of dark spots in this breed.

Objective: The goal of this study was to characterize and investigate the effect of single nucleotide polymorphisms (SNPs) or InDels in tyrosinase related protein 1 (TYRP1), dopachrome tautomerase (DCT), and Ras-related protein 38 (RAB38) genes on coat color in South American camelids. TYRP1 is among one of the major regulators of pigmentation in mammals. DCT is a pigment cell-specific gene that plays an important role in the melanin synthesis pathway and the expression of skin color. RAB38 is involved in the transport of tyrosinase (TYR) and TYRP1, which are key enzymes in melanin production. Methods: Five fleece color tones from 327 alpacas (white, black, brown, gray and light fawn) were investigated to characterize and identify polymorphisms that play a role in coat color. Among the SNPs and Indels identified in the coding region, statistical analysis was performed to determine the association between the polymorphisms and fleece color in camelids, as well as the determination of allelic frequencies. Results: Twenty two polymorphisms in the coding region, seven of them were nonsynonymous, eleven were synonymous, three located in the 5’UTR region and one deletion in the 3’UTR region were identified in the TYRP1 gene; in the DCT gene were identified twenty two polymorphisms, where five of them were non-synonymous, twelve were synonymous, one polymorphism in the 5’UTR region and four in the 3’UTR region; in adition, in the RAB38 gene six polymorphisms were identified, one was nonsynonymous, two were synonymous, in addition to three polymorphisms in the 3’UTR region were found in the RAB38 gene. There were low to moderate polymorphisms in the populations. Significant differences (P<0.001) were found with color fiber in the alpaca populations studied for TYRP1 and DCT genes. [ABSTRACT FROM AUTHOR]

The transcriptome data of skin cells from domestic cats with brown, orange, and white coats were analyzed using a public database to investigate the possible relationship between coat color‐related gene expression and squamous cell carcinoma risk, as well as the mechanism of deafness in white cats. We found that the ratio of the expression level of genes suppressing squamous cell carcinoma to that of genes promoting squamous cell carcinoma might be considerably lower than the theoretical estimation in skin cells with orange and white coats in white‐spotted cat. We also found the possibility of the frequent production of KIT lacking the first exon (d1KIT) in skin cells with white coats, and d1KIT production exhibited a substantial negative correlation with the expression of SOX10, which is essential for melanocyte formation and adjustment of hearing function. Additionally, the production of d1KIT was expected to be due to the insulating activity of the feline endogenous retrovirus 1 (FERV1) LTR in the first intron of KIT by its CTCF binding sequence repeat. These results contribute to basic veterinary research to understand the relationship between cat skin coat and disease risk, as well as the underlying mechanism. [ABSTRACT FROM AUTHOR]

Objective: Pigment production and distribution are controlled through multiple proteins, resulting in different coat color phenotypes of sheep. Methods: The expression distribution of vimentin (VIM) and transthyretin (TTR) in white and black sheep skins was detected by liquid chromatography-electrospray ionization tandem MS (LC-ESI-MS/MS), gene ontology (GO) statistics, immunohistochemistry, Western blot, and quantitative real time polymerase chain reaction (qRT-PCR) to evaluate their role in the coat color formation of sheep. Results: LC-ESI-MS/MS results showed VIM and TTR proteins in white and black skin tissues of sheep. Meanwhile, GO functional annotation analysis suggested that VIM and TTR proteins were mainly concentrated in cellular components and biological process, respectively. Further research confirmed that VIM and TTR proteins were expressed at significantly higher levels in black sheep skins than in white sheep skins by Western blot, respectively. Immunohistochemistry notably detected VIM and TTR in hair follicle, dermal papilla, and outer root sheath of white and black sheep skins. qRT-PCR results also revealed that the expression of VIM and TTR mRNAs was higher in black sheep skins than in white sheep skins. Conclusion: The expression of VIM and TTR were higher in black sheep skins than in white sheep skins and the transcription and translation were unanimous in this study. VIM and TTR proteins were expressed in hair follicles of white and black sheep skins. These results suggested that VIM and TTR were involved in the coat color formation of sheep. [ABSTRACT FROM AUTHOR]

MiRNAs as a series of small noncoding RNAs that play a crucial part in regulating coat color and hair follicle development. In the previous Solexa sequencing experiments, there were many miRNAs expressed differentially in alpacas with different coat color, including miR-193b.But the mechanism of miR-193b in mammalian pigmentation is still unknown. In this study, bioinformatics analysis showed that WNT10A and GNAI2 might be the target genes of miR-193b. qRT-PCR showed the expression of miR-193b in white Cashmere goats' skins was obviously lower than that in browns, and the expression of WNT10A and GNAI2 were similar with miR-193b. The protein levels of WNT10A and GNAI2 indicated the same findings. Furthermore, the expression of WNT10A and GNAI2 in keratinocytes were analyzed from mRNA and protein levels, the results manifested that the group of overexpression of miR-193b in HaCaT cells increased the expressions of target genes, and miR-193b inhibition group reduced expressions. Luciferase report assays confirmed that the targeting relationship between miR-193b and target genes (WNT10A and GNAI2), the results showed that miR-193b was positively correlated with target genes. These experimental data showed that miR-193b might participate in adjustment of coat color in skin tissue of Cashmere goat by targeting WNT10A and GNAI2. [ABSTRACT FROM AUTHOR]

In North American gray wolves, black coat color is dominantly inherited via a 3 base pair coding deletion in the canine beta defensin 3 (CBD103) gene. This 3 base pair deletion, called the K B allele, was introduced through hybridization with dogs and subsequently underwent a selective sweep that increased its frequency in wild wolves. Despite apparent positive selection, K BB wolves have lower fitness than wolves with the K yB genotype, even though the 2 genotypes show no observable differences in black coat color. Thus, the K B allele is thought to have pleiotropic effects on as-yet unknown phenotypes. Given the role of skin-expressed CBD103 in innate immunity, we hypothesized that the K B allele influences the keratinocyte gene expression response to TLR3 pathway stimulation and/or infection by canine distemper virus (CDV). To test this hypothesis, we developed a panel of primary epidermal keratinocyte cell cultures from 24 wild North American gray wolves of both K yy and K yB genotypes. In addition, we generated an immortalized K yy line and used CRISPR/Cas9 editing to produce a K yB line on the same genetic background. We assessed the transcriptome-wide responses of wolf keratinocytes to the TLR3 agonist polyinosinic:polycytidylic acid (polyI:C), and to live CDV. K locus genotype did not predict the transcriptional response to either challenge, suggesting that variation in the gene expression response does not explain pleiotropic effects of the K B allele on fitness. This study supports the feasibility of using cell culture methods to investigate the phenotypic effects of naturally occurring genetic variation in wild mammals. [ABSTRACT FROM AUTHOR]

c-KIT, a type III receptor protein tyrosine kinase, plays an essential role in melanocyte development, migration, and survival. Mutations within the c-KIT gene are previously shown to cause the white coat color phenotypes in pigs, mice, goats, and humans. However, up so far, the splicing isoform(s), genomic architecture of c-KIT have not been characterized well in merino sheep. Reverse transcriptase (RT)-PCR analysis with molecular prediction identified two basic splice variants: Transcript Variant-1, 2 for 12 bp insertion coding sequences (CDS) corresponding to the four amino acids 'GNSK', respectively. Using 5′ RACE, here we report for the first time a novel c-KIT 'Transcript Variant-3' from the skin of merino sheep by comparative genome analyses at exon(1)-intron(1)-exon(2) boundaries. In contrast, a single product of 795 bp was characterized by 3' RACE. We also demonstrated that the c-KIT gene expression at the transcript level is not mediated via an intron-9 splicing event. Overall, beyond what was observed in other mammals, our data provide novel insights into the molecular structure of the c-KIT gene in sheep. [ABSTRACT FROM AUTHOR]

Summary: The llama (Lama glama) is a fiber‐producing species that presents a wide range of coat colors, among which white is one of the most important for the textile industry. However, there is little information about the molecular mechanisms that control the white phenotype in this species. In domestic mammals, a white coat is usually produced by mutations in the KIT proto‐oncogene receptor tyrosine kinase (KIT) and microphthalmia‐associated transcription factor (MITF) genes. In this work we have sequenced and described the coding regions of KIT and MITF‐M, the melanocyte‐specific isoform, and the two transcriptional variants MITF‐M(−) and MITF‐M(+). Moreover, we studied the expression of these genes in the skin of white and colored llamas. Although no variants were revealed to be associated with white coat color, significant differences between phenotypes were observed in the expression levels of KIT and MITF‐M. Interestingly, white llamas expressed less MITF‐M(+) than did colored ones, which is consistent with a consequent reduction in the synthesis of melanin. Even though our results indicate that downregulation of KIT and MITF‐M expression is involved in white phenotype production in llamas, the causative gene of white coat color remains unknown. [ABSTRACT FROM AUTHOR]

Coat color is a key economic trait in sheep. Some candidate genes associated with animal's coat color were found. Particularly, v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) and microphthalmia-associated transcription factor (MITF) play a key role in the modulation of hair pigmentation in mammals. This study investigated those two candidate genes' mutations and expressions associated with wool color in Tibetan sheep. First, the gene polymorphisms of those two genes were analyzed, and then, relative mRNA expression levels of those two genes in skin tissue with different coat colors were compared. Thirdly, KIT and MITF protein expression levels were detected through Western blot and immune-histochemical. Allele C was predominant allele in the white coat color Tibetan sheep population of the MITF coding region g. 1548 C/T loci. The relative MITF mRNA expression in black coat skin tissue was significantly higher than white (P0.05), while the level of KIT protein expression in skin tissues of white and black coats was also roughly equivalent. Our study observed that, the level of MITF protein expression in black coat skin tissue was significantly higher than that in white coat skin tissue, and positive staining for MITF protein expression was detected mainly in the epidermis and the dermal papilla, bulb, and outer root sheath of hair follicles. We conclude that the black coat of Tibetan sheep is related to high MITF expression in the hair follicles, and MITF may be important for coat color formation of Tibetan sheep.

The aim of the present study was to identify coat color associated genes that are differentially expressed in mature Korean brindle cattle (KBC) with different coat colors and in Hanwoo cows. KBC calves, before and after coat color appearance, were included. Total cellular RNA was isolated from the tail hair cells and used for microarray. The number of expressed coat color associated genes/probes was 5813 in mature KBC and Hanwoo cows. Among the expressed coat color associated genes/probes, 167 genes were the coat color associated genes listed in the Gene card database and 125 genes were the pigment and melanocyte genes listed in the Gene ontology_bovine database. There were 23 genes/probes commonly listed in both databases and their expressions were further studied. Out of the 23 genes/probes, MLPH, PMEL, TYR and TYRP1 genes were expressed at least two fold higher (p

In this study, we used high-throughput technology to provide the first transcriptome dataset for differentially expressed miRNA in mixed pools of dermis tissue from black- and white-coated yak to research the possible molecular mechanisms of yak coat pigmentation. In this study, 92,636,002 and 95,917,842 clear reads were generated through Illumina paired-end sequencing. A total of 78 differentially expressed miRNAs (DEMs) were identified, including 59 upregulated and 19 downregulated miRNAs in the mixed pools of white-coated yak compared with the mixed pools of black-coated yak. In addition, 3634 genes were predicted as putative targets of DEMs. These DEGs related to 59 GO categories and were enriched in 216 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, including melanogenesis and the Wnt signaling pathway. The results of the current study indicated that the coat color of the yak involved the transcriptional regulation process of miRNAs. These results provide helpful data to understand the molecular mechanisms of yak coat pigmentation. [ABSTRACT FROM AUTHOR]

Background: Studies on the molecular genetics of horse skin pigmentation have typically focused on very few genes and proteins. In this study, we used Illumina sequencing to determine the global gene expression profiles in horses with white-colored coats and those with black-colored coats, with the goal of identifying novel genes that could regulate horse coat color. Results: Genes encoding ribosomal-associated proteins were highly expressed in horse skin. We found a total of 231 unigenes that were differentially expressed between horses with white coats and horses with black coats; 119 were down-regulated, and 112 were up-regulated. Many of the up-regulated genes in black horses, such as genes related to tyrosine metabolism, may directly regulate dark coat color. Keratin genes, MIA family genes, fatty acidrelated genes, and melanoma-associated genes were also differentially regulated, which suggests that they may play important roles in coat color formation. Conclusions: These findings show that the transcription profiles from white and black horse skin provide useful information to understand the genetics underlying the control of skin melanin synthesis in horses, which may enhance our knowledge of human skin diseases, such as melanoma and albinism. [ABSTRACT FROM AUTHOR]

This study performed a comprehensive expression profiling of genes expressed in the skin of goats with three different coat colors by Illumina Sequencing. A total of 91 significantly expressed genes were detected when comparing gray skin to white skin library and these included 74 up-regulated and 17 down-regulated genes in gray skin. There were 67 differentially expressed genes between brown skin and white skin libraries, 23 of which were up-regulated and 44 were down-regulated in brown skin. When we compared brown and gray libraries, 154 differentially expressed genes were found, of which 33 showed higher expression and 121 showed lower expression in brown skin. To our surprise, MC1R, MITF, TYR and KIT showed no significant difference in expression between the goats with three skin colors, whereas ASIP was detected in white skin but not in dark skins. In this study, PMEL, TRPM1, TYRP1 and DCT were significantly up-regulated in brown goat skin compare with gray and white skins. PMEL showed higher expression in gray goat skin compared with white goat skin, whereas there were no significant differences in the expression of TYRP1, TRPM1 and DCT between gray and white skin samples. In addition, ELOVL3 showed higher expression in gray goat skin than in brown and white skins, whereas there was no significant differences in the expression of ELOVL3 between brown and white skin samples. These results expand our understanding of the complex molecular mechanisms of skin physiology and melanogenesis in goat and provide a foundation for future studies. [ABSTRACT FROM AUTHOR]

ASIP has been described to regulate lipid metabolism and acts as a paracrine factor in the regulation of adiposity. The RTqPCR and Western Blotting was used to investigate the expression level of Agouti mRNA and Agouti signaling protein (ASIP) in viscera (heart, liver, spleen, lung and kidney) of goat with white, brown with gray and black coat color. The main trend of Agouti mRNA abundance represented white goat and liver tissue obviously. The protein was detected in all viscera tissues and it expressed white goat and kidney mainly. However, ASIP differential expresses in different viscera for goats weren't consistent with the mRNA levels. The differences might be the result from the transcription regulation, translation regulation, protein modified and protein interaction and so on. [ABSTRACT FROM AUTHOR]

Introduction. To investigate whether the membrane-associated transporter protein SLC45A2 is differentially expressed in the skin of sheep with different coat colors and to determine its correlation with coat color establishment in sheep. Material and methods. The expression of SLC45A2 in sheep skin samples with different coat colors was qualitatively and quantitatively analyzed by PCR amplification, RT-PCR, immunohistochemical staining and Western blotting. Results. A 193-bp SLC45A2 CDS sequence was successfully amplified from sheep skin samples with diverse coat colors. RT-PCR analysis revealed that SLC45A2 mRNA was expressed in all sheep skin samples tested, with relative expression levels of 512.74 ± 121.51 in black skin, 143.38 ± 119.31 and 1.36 ± 0.09 in black dots and white dots of piebald skin, respectively, and 1.02 ± 0.23 in white skin (p < 0.01**). Positive SLC45A2 protein bands were also detected in all skin samples by Western blot analysis, with relative expression levels of 0.85 ± ± 0.17** in black skin, 0.60 ± 0.05** and 0.34 ± 0.07 in black dots and white dots of piebald skin, respectively, and 0.20 ± 0.05 in white skin (p < 0.01**). Immunohistochemical assays revealed that SLC45A2 was expressed in the hair follicle matrix, the inner and outer root sheath, and the dermal papilla in the skin tissues with different coat colors. These patterns were quantified by optical density (OD) analysis, which yielded relative expression levels of 0.23 ± 0.11 in black skin, 0.19 ± 0.09 and 0.10 ± 0.03 in black dots and white dots of piebald skin, respectively, and 0.08 ± 0.01 in white skin (p < 0.05*). Conclusion. SLC45A2 is detectably expressed in sheep skin of all coat colors, though at significantly different levels. SLC45A2 may participate in the establishment of coat color by regulating the synthesis and trafficking of melanin.   [ABSTRACT FROM AUTHOR]

The purpose of this study was to evaluate the association among the polymorphism of the tyrosinase-related protein 1 (TYRP1) gene, differential expression in the skin, and coat color in Mongolian horses. The polymorphisms of TYRP1 in five groups of Mongolian horse, Sanhe horse and thoroughbreds were genotyped using PCR-SSCP. The relative gene expression level of TYRP1 in the skin tissues among different coat color Mongolian horses was determined using quantitative real-time PCR. The association of the TYRP1 polymorphisms with coat color was analyzed. A missense mutation (C189T) in exon 2 of TYRP1 was detected in the Mongolian horse, resulting in two genotypes: CC and CT. The TT genotype was not observed in these Mongolian horses. Allele C was found to be the predominant allele in black, bay, and chestnut horses. The results showed the TYRP1 gene polymorphism to be closely associated with dark coat color (P=2.3 x 10-6). The expression level of the TYRP1 gene was the highest in the chestnut skin tissues and the lowest in the gray skin tissues of Mongolian horses. These results suggest that the allele and expression of the TYRP1 gene are associated with coat color and colored coat formation in Mongolian horses. This is the first report of the association of the TYRP1 gene polymorphism with coat color in the Mongolian horse. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) play a key role in many biological processes by regulating gene expression at the post-transcriptional level. A number of miRNAs have been identified from livestock species. However, compared with other animals, such as pigs and cows, the number of miRNAs identified in goats is quite low, particularly in hair follicles. In this study, to investigate the functional roles of miRNAs in goat hair follicles of goats with different coat colors, we sequenced miRNAs from two hair follicles samples (white and black) using Solexa sequencing. A total of 35,604,016 reads were obtained, which included 30,878,637 clean reads (86.73%). MiRDeep2 software identified 214 miRNAs. Among them, 205 were conserved among species and nine were novel miRNAs. Furthermore, DESeq software identified six differentially expressed miRNAs. Quantitative PCR confirmed differential expression of two miRNAs, miR-10b and miR-211. KEGG pathways were analyzed using the DAVID website for the predicted target genes of the differentially expressed miRNAs. Several signaling pathways including Notch and MAPK pathways may affect the process of coat color formation. Our study showed that the identified miRNAs might play an essential role in black and white follicle formation in goats. [ABSTRACT FROM AUTHOR]

Light-dark coat color variation is a common aspect of color diversity within and across mammalian taxa. This variation in pelage brightness is associated with aspects of evolutionary ecology, particularly for primates, but little is known about the genetic mechanisms underlying light-dark differences in pelage pigmentation. Previous work, focusing particularly on macaques (Genus Macaca), has found no clear relationship between color variation and coding sequences of key pigmentation genes. This suggests that other loci and/or gene regulatory differences underlie this variation and raises the question of how patterns of gene expression differ in light verses dark hair follicles. Here, we examine relative expression levels of pigmentation genes in hair follicles from free-ranging rhesus macaques ( Macaca mulatta) showing stark light-dark coat color variation. We quantified the brightness (reflectance) of plucked hair tufts using a spectrophotometer. We extracted RNA from the follicles and used quantitative RT-PCR to measure the relative amounts of gene product (mRNA) for seven candidate pigmentation genes ( MITF, MC1R, MGRN1, ATRN, SLC24A5, TYRP1, and DCT). Expression values were normalized with the house-keeping gene ACTB. All candidate genes were expressed at similar levels in dark, intermediate, and light hair, and thus, light-dark variation in macaque coat color is unlikely to be due to differences in the expression of these key pigmentation genes. This study represents the first examination of gene expression and natural color variation in a non-human primate population. Our results indicate that even in a system, like pigmentation, where a candidate-gene approach is promising, identifying important intra-specific gene regulatory differences remains challenging. [ABSTRACT FROM AUTHOR]

The objective of this study was to identify the Melanocortin 1 Receptor (MC1R) gene in sorrel-, black- and white-Bali cows originated from Kupang-East Nusa Tenggara. Blood samples were taken from 46 cows consisted of of 17, 18 and 8 of sorrel, black and 8 heads, respectively, and 3 heads of bull. The PCR-RFLP method was performed to identify the gene. Amplification of PCR gene MC1R of Kupang's Bali Cattle (KBC) was 296 bp. The digestion by using MspI enzyme showed that there were EE genotype (169, 136 bp) in Bali bull, sorrel and white cows. Two 2 genotypes, those were EE (169, 136 bp) and Ee (296, 169, 136 bp) were found in black Bali cow. The frequency of E alele of KBC was 0.99 (99%) and e alele was 0.01 (1%). MC1R gene in KBC was monomorphic and the it can be used as the marker of the coat color in the population. [ABSTRACT FROM AUTHOR]

Background: MicroRNAs (miRNAs) are small, non-coding 21–25 nt RNA molecules that play an important role in regulating gene expression. Little is known about the expression profiles and functions of miRNAs in skin and their role in pigmentation. Alpacas have more than 22 natural coat colors, more than any other fiber producing species. To better understand the role of miRNAs in control of coat color we performed a comprehensive analysis of miRNA expression profiles in skin of white versus brown alpacas. Results: Two small RNA libraries from white alpaca (WA) and brown alpaca (BA) skin were sequenced with the aid of Illumina sequencing technology. 272 and 267 conserved miRNAs were obtained from the WA and BA skin libraries, respectively. Of these conserved miRNAs, 35 and 13 were more abundant in WA and BA skin, respectively. The targets of these miRNAs were predicted and grouped based on Gene Ontology and KEGG pathway analysis. Many predicted target genes for these miRNAs are involved in the melanogenesis pathway controlling pigmentation. In addition to the conserved miRNAs, we also obtained 22 potentially novel miRNAs from the WA and BA skin libraries. Conclusion: This study represents the first comprehensive survey of miRNAs expressed in skin of animals of different coat colors by deep sequencing analysis. We discovered a collection of miRNAs that are differentially expressed in WA and BA skin. The results suggest important potential functions of miRNAs in coat color regulation. [ABSTRACT FROM AUTHOR]

Abstract: To evaluate if loci responsible for coat color phenotypes contribute to behavioral characteristics, we specified novel gene loci associated with social exploratory behavior and examined the effects of the frequency of each allele at distinct loci on behavioral expression. We used the F2 generation, which arose from the mating of F1 mice obtained by interbreeding DBA/2 and ICR mice. Phenotypic analysis indicated that the agouti and albino loci affect behavioral traits. A genotype-based analysis revealed that novel exploratory activity was suppressed in a manner dependent on the frequency of the dominant wild-type allele at the agouti, but not albino, locus. The allele-dependent suppression was restricted to colored mice and was not seen in albino mice. The present results suggest that the agouti locus contributes to a particular behavioral trait in the presence of a wild-type allele at the albino locus, which encodes a structural gene for tyrosinase. [Copyright &y& Elsevier]

Abstract: Although the influence of endocrine factors is well established, the molecular and cellular mechanisms controlling coat color are not completely understood. A major mechanism for post-transcriptional regulation of gene expression is through the action of microRNAs (miRNAs), which anneal to the 3’ untranslated region of mRNAs in a sequence-specific fashion and either block translation or promote transcript degradation. In this study, we investigated the expression of miRNAs in the skin of alpacas with brown vs white coat color using a microarray screen; identified potential mRNA targets for identified miRNAs among coat color genes; and subsequently determined the ability of a specific, differentially expressed miRNA (miR-25) to suppress expression of micropthalmia-associated transcription factor (MITF), a predicted miR-25 target gene that regulates genes linked to coat color. Expression of 10 different miRNA species in the skin of alpacas with brown vs white coat color was identified from microarray screens. Of the 10 alpaca skin miRNAs identified, predicted binding sites in the 3′ untranslated region of RNAs encoding for known genes linked to coat color were primarily for miR-25, but sites were also identified for miR-129 and miR-377. Potential miR-25 binding sites were present in transcripts encoding for 11 coat color genes, including MITF. An inverse relationship between transcript abundance for MITF and miR-25 was observed in skin samples collected from alpacas with white vs brown coat color. Furthermore, overexpression of miR-25 in cultured melanocytes reduced MITF mRNA and protein abundance and corresponding mRNA abundance for the MITF-regulated enzymes tyrosinase and tyrosinase-related protein 1. Results support a novel functional role for miRNA-25 in the regulation of gene expression linked to coat color. [Copyright &y& Elsevier]

• Seven ASIP transcripts with alternative 5′UTR were isolated. • A fusion transcript between ASIP and NCOA6 gene was found. • The novel non-coding exon 1A" was identified within the 5́UTR regions. • Multiple start sites and alternative splicing create diversity of ASIP transcripts. • ASIP expression patterns differed among color phenotypes. The Agouti gene (ASIP) is one of the most important genes for coat color determination in mammals. It has a complex structure with several promoters and alternative non-coding first exons that are transcribed into mRNAs with different 5'UTR. These mRNA isoforms regulate the temporal and spatial expression of the gene, producing diverse pigmentation patterns. Here, we studied ASIP transcriptional variants and their expression in the skin of llamas with different coat color phenotypes. We also described the ASIP locus, including promoter usage and the splicing events that originate each transcript variant. Using 5'RACE-PCR we isolated seven ASIP transcripts with alternative 5′UTR, where exons 1A, 1A', 1C, 1D, and a novel non-coding exon 1A" were identified. Additionally, new alternative spliced forms were found. The diversity of ASIP 5′UTRs is originated by a complex pattern of alternative promoter usage, multiple transcription start sites and splicing events that include exon skipping and alternative 3′ splicing site selection. We found that ASIP was highly expressed in llamas with white and brown phenotypes while black animals presented very low expression. The main responsible for this difference was a fusion transcript between ASIP and NCOA6 genes, which was present in the skin of white and brown llamas but not in the black ones. The rest of ASIP transcripts presented very low expression in the skin, indicating that the main regulation point for ASIP gene expression is at the transcriptional level. Nevertheless, the characteristics of the 5′UTRs sequences suggest that alternative transcripts could be regulated differently at the protein synthesis level. [ABSTRACT FROM AUTHOR]