Pigment Mutants Research Articles

Quantification and Angler Opinion of Skin Pigment Mutations in Blue Catfish from Milford Reservoir, Kansas

Quantification and Angler Opinion of Skin Pigment Mutations in Blue Catfish from Milford Reservoir, Kansas

Deciphering putative protein profile of a photomorphogenic high pigment mutant of Solanum lycopersicum (hp-1) by high-throughput LC–MS/MS analysis

Deciphering putative protein profile of a photomorphogenic high pigment mutant of Solanum lycopersicum (hp-1) by high-throughput LC–MS/MS analysis

Identification of orange color-related gene, PhcpcC, in Pyropia haitanensis

Pigmentation-related mutations can be utilized to distinguish between differentially colored sectors of chimeric thalli, thereby facilitating the efficient breeding of economically valuable Pyropia/Porphyra seaweed species. However, the specific trait loci and alleles responsible for Pyropia/Porphyra coloration have yet to be identified, which limits the applicability of coloration mutants for breeding and genetic analyses. In this study, to preserve the genetic integrity of the population, only four-colored thalli were considered when constructing the doubled haploid (DH) Pyropia haitanensis population, which consisted of 480 homozygous offspring lines (representing the largest DH Pyropia/Porphyra population). The offspring lines in the DH population exhibited both wild-type colored and orange sectors, with a segregation ratio of approximately 1:1, indicating that the orange coloration was controlled by a single nuclear gene. Through BSA-seq analysis (99% confidence interval), a candidate region of 0.5 Mb was identified in the P. haitanensis genome. Additionally, a non-synonymous SNP [A/G] was detected at base-pair position 481 in the coding region of PhcpcC, which encodes a phycocyanin-associated rod linker protein. This SNP locus was verified in both DH and natural populations, with the wild-type colored lines having an A base and the orange lines having a G base at this locus. Therefore, PhcpcC may be the gene associated with the orange coloration of P. haitanensis. The molecular marker developed in this study can be employed to exploit pigmentation mutants for breeding and genetic analyses of Pyropia/Porphyra species.

ZnO nanoparticles: A key ingredient of sunscreen shows absence of adverse effects on Drosophila melanization pathway

The rapid development of zinc oxide nanoparticle (ZnO NP) based sunscreens, shampoos and other skin care products has raised a concern about their safety at the cellular level. The melanization pathway, being an important pathway of skin cells, is prone to being affected by ZnO NPs. Therefore it is necessary to understand the ZnO NP mediated effect on the melanin synthetic pathway. Our previous study on bulk zinc oxide has shown that large sized zinc oxide particles did not have any adverse effect on melanization in Drosophila, hence it would be interesting to study the behavior of ZnO when present in the nano-form by virtue of the changed size and thereby changed properties. The present study was undertaken to investigate ZnO NP mediated effects on key metabolites of the Drosophila melanin pathway. Spectrophotometric analysis of melanin was performed. followed by a study of the enzyme activity of phenol oxidase and HPLC quantification of dopa and tyrosine. FTIR spectroscopy was carried out to examine the interaction of the ZnO NPs and precursor amines of the melanin synthetic pathway. Pigmentation mutants of Drosophila were also investigated for validation of the results under a different genetic background. The ZnO NPs did not show any decrement in the levels of melanin after the treatment. Moreover there was no adverse effect on the phenol oxidase activity, Tyrosine and L-Dopa. The FTIR studies showed minimal interaction between the ZnO NPs and the metabolites of the melanin pathway. The results of the present study have clearly indicated the absence of any adverse effects on key metabolites of the melanin pathway after ZnO NP treatment in both wild-type and mutant Drosophila. This is the first report of a detailed analysis of ZnO NP mediated effects on key constituents of the Drosophila melanization pathway, providing information about its safety for the usage in cosmetics.

Random mutagenesis of Phaeodactylum tricornutum using ultraviolet, chemical, and X-radiation demonstrates the need for temporal analysis of phenotype stability

We investigated two non-ionising mutagens in the form of ultraviolet radiation (UV) and ethyl methanosulfonate (EMS) and an ionising mutagen (X-ray) as methods to increase fucoxanthin content in the model diatom Phaeodactylum tricornutum. We implemented an ultra-high throughput method using fluorescence-activated cell sorting (FACS) and live culture spectral deconvolution for isolation and screening of potential pigment mutants, and assessed phenotype stability by measuring pigment content over 6 months using high-performance liquid chromatography (HPLC) to investigate the viability of long-term mutants. Both UV and EMS resulted in significantly higher fucoxanthin within the 6 month period after treatment, likely as a result of phenotype instability. A maximum fucoxanthin content of 135 ± 10% wild-type found in the EMS strain, a 35% increase. We found mutants generated using all methods underwent reversion to the wild-type phenotype within a 6 month time period. X-ray treatments produced a consistently unstable phenotype even at the maximum treatment of 1000 Grays, while a UV mutant and an EMS mutant reverted to wild-type after 4 months and 6 months, respectively, despite showing previously higher fucoxanthin than wild-type. This work provides new insights into key areas of microalgal biotechnology, by (i) demonstrating the use of an ionising mutagen (X-ray) on a biotechnologically relevant microalga, and by (ii) introducing temporal analysis of mutants which has substantial implications for strain creation and utility for industrial applications.

The Adaptive Role of Carotenoids and Anthocyanins in Solanum lycopersicum Pigment Mutants under High Irradiance.

The effects of high-intensity light on the pigment content, photosynthetic rate, and fluorescence parameters of photosystem II in high-pigment tomato mutants (hp 3005) and low-pigment mutants (lp 3617) were investigated. This study also evaluated the dry weight percentage of low molecular weight antioxidant capacity, expression patterns of some photoreceptor-regulated genes, and structural aspects of leaf mesophyll cells. The 3005 mutant displayed increased levels of photosynthetic pigments and anthocyanins, whereas the 3617 mutant demonstrated a heightened content of ultraviolet-absorbing pigments. The photosynthetic rate, photosystem II activity, antioxidant capacity, and carotenoid content were most pronounced in the high-pigment mutant after 72 h exposure to intense light. This mutant also exhibited an increase in leaf thickness and water content when exposed to high-intensity light, suggesting superior physiological adaptability and reduced photoinhibition. Our findings indicate that the enhanced adaptability of the high-pigment mutant might be attributed to increased flavonoid and carotenoid contents, leading to augmented expression of key genes associated with pigment synthesis and light regulation.

Carotenoid and transcriptome profiles of a novel citrus cultivar ‘Jinlegan’ reveal mechanisms of yellowish fruit formation

Citrus fruit coloration is one of the vital quality traits that is determined primarily by the composition and content of carotenoids. Natural citrus fruit pigment mutants are available to study diverse and complex carotenoid metabolism. Here, ‘Jinlegan’ (MT) tangor is a spontaneous bud mutant derived from ‘Shiranuhi’ (WT) with distinctive bright yellow fruit. High performance liquid chromatography (HPLC) analysis revealed that the yellowish MT flavedo and pulp were primarily caused by the decrease in total carotenoid content. The total carotenoid content in MT flavedo was reduced by 75% (79.98 μg/g DW) compared with that in WT (318.40 μg/g DW), including approximately 84%, 80%, and 60% reductions in the contents of β-cryptoxanthin, violaxanthin and zeaxanthin, respectively. The total carotenoid content in MT pulp was 60% lower (10.09 μg/g DW) than that in WT pulp (26.61 μg/g DW), which was mainly due to a 70% and 30% decrease in the contents of β-cryptoxanthin and zeaxanthin, respectively. To explore the molecular mechanism underlying carotenoid variation in MT, RNA-seq analyses were performed on the flavedo and pulp of WT and MT at five developmental stages. The reduced expression of phytoene synthase (CrPSY) and β-carotenehydroxylase 1 (CrBCH1) in the flavedo and pulp of MT at the breaker stage might be the major cause of the reduction in carotenoids. Weighted gene co-expression network analysis (WGCNA) further identified 23 key transcription factors that are closely associated with carotenoid accumulation. This study demonstrated a comprehensive picture of the metabolic and transcriptional alterations of a unique yellowish citrus fruit mutant, which provides new insights into the molecular regulation of carotenoid accumulation in citrus fruit.

Generation of a transparent killifish line through multiplex CRISPR/Cas9mediated gene inactivation.

Body pigmentation is a limitation for in vivo imaging and thus for the performance of longitudinal studies in biomedicine. A possibility to circumvent this obstacle is the employment of pigmentation mutants, which are used in fish species like zebrafish and medaka. To address the basis of aging, the short-lived African killifish Nothobranchius furzeri has recently been established as a model organism. Despite its short lifespan, N. furzeri shows typical signs of mammalian aging including telomere shortening, accumulation of senescent cells, and loss of regenerative capacity. Here, we report the generation of a transparent N. furzeri line by the simultaneous inactivation of three key loci responsible for pigmentation. We demonstrate that this stable line, named klara, can serve as a tool for different applications including behavioral experiments and the establishment of a senescence reporter by integration of a fluorophore into the cdkn1a (p21) locus and in vivo microscopy of the resulting line.

MicroRNA regulation of skin pigmentation in golden-back mutant of crucian carp from a rice-fish integrated farming system

BackgroundMicroRNAs (miRNAs) are endogenous small non-coding RNAs (21–25 nucleotides) that act as essential components of several biological processes. Golden-back crucian carp (GBCrC, Carassius auratus) is a naturally mutant species of carp that has two distinct body skin color types (golden and greenish-grey), making it an excellent model for research on the genetic basis of pigmentation. Here, we performed small RNA (sRNA) analysis on the two different skin colors via Illumina sequencing.ResultsA total of 679 known miRNAs and 254 novel miRNAs were identified, of which 32 were detected as miRNAs with significant differential expression (DEMs). 23,577 genes were projected to be the targets of 32 DEMs, primarily those involved in melanogenesis, adrenergic signaling in cardiomyocytes, MAPK signaling pathway and wnt signaling pathway by functional enrichment. Furthermore, we built an interaction module of mRNAs, proteins and miRNAs based on 10 up-regulated and 13 down-regulated miRNAs in golden skin. In addition to transcriptional destabilization and translational suppression, we discovered that miRNAs and their target genes were expressed in the same trend at both the transcriptional and translational levels. Finally, we discovered that miR-196d could be indirectly implicated in regulating melanocyte synthesis and motility in the skin by targeting to myh7 (myosin-7) gene through the luciferase reporter assay, antagomir silencing in vivo and qRT-PCR techniques.ConclusionsOur study gives a systematic examination of the miRNA profiles expressed in the skin of GBCrC, assisting in the comprehension of the intricate molecular regulation of body color polymorphism and providing insights for C. auratus breeding research.

Genetic Underpinnings of Carotenogenesis and Light-Induced Transcriptome Remodeling in the Opportunistic Pathogen Mycobacterium kansasii.

Mycobacterium kansasii (Mk) causes opportunistic pulmonary infections with tuberculosis-like features. The bacterium is well known for its photochromogenicity, i.e., the production of carotenoid pigments in response to light. The genetics defining the photochromogenic phenotype of Mk has not been investigated and defined pigmentation mutants to facilitate studies on the role of carotenes in the bacterium's biology are not available thus far. In this study, we set out to identify genetic determinants involved in Mk photochromogenicity. We screened a library of ~150,000 transposon mutants for colonies with pigmentation abnormalities. The screen rendered a collection of ~200 mutants. Each of these mutants could be assigned to one of four distinct phenotypic groups. The insertion sites in the mutant collection clustered in three chromosomal regions. A combination of phenotypic analysis, sequence bioinformatics, and gene expression studies linked these regions to carotene biosynthesis, carotene degradation, and monounsaturated fatty acid biosynthesis. Furthermore, introduction of the identified carotenoid biosynthetic gene cluster into non-pigmented Mycobacterium smegmatis endowed the bacterium with photochromogenicity. The studies also led to identification of MarR-type and TetR/AcrR-type regulators controlling photochromogenicity and carotenoid breakdown, respectively. Lastly, the work presented also provides a first insight into the Mk transcriptome changes in response to light.

Colony growth and biofilm formation of Aspergillus niger under simulated microgravity.

The biotechnology- and medicine-relevant fungus Aspergillus niger is a common colonizer of indoor habitats such as the International Space Station (ISS). Being able to colonize and biodegrade a wide range of surfaces, A. niger can ultimately impact human health and habitat safety. Surface contamination relies on two key-features of the fungal colony: the fungal spores, and the vegetative mycelium, also known as biofilm. Aboard the ISS, microorganisms and astronauts are shielded from extreme temperatures and radiation, but are inevitably affected by spaceflight microgravity. Knowing how microgravity affects A. niger colony growth, in particular regarding the vegetative mycelium (biofilm) and spore production, will help prevent and control fungal contaminations in indoor habitats on Earth and in space. Because fungal colonies grown on agar can be considered analogs for surface contamination, we investigated A. niger colony growth on agar in normal gravity (Ground) and simulated microgravity (SMG) conditions by fast-clinorotation. Three strains were included: a wild-type strain, a pigmentation mutant (ΔfwnA), and a hyperbranching mutant (ΔracA). Our study presents never before seen scanning electron microscopy (SEM) images of A. niger colonies that reveal a complex ultrastructure and biofilm architecture, and provide insights into fungal colony development, both on ground and in simulated microgravity. Results show that simulated microgravity affects colony growth in a strain-dependent manner, leading to thicker biofilms (vegetative mycelium) and increased spore production. We suggest that the Rho GTPase RacA might play a role in A. niger’s adaptation to simulated microgravity, as deletion of ΔracA leads to changes in biofilm thickness, spore production and total biomass. We also propose that FwnA-mediated melanin production plays a role in A. niger’s microgravity response, as ΔfwnA mutant colonies grown under SMG conditions showed increased colony area and spore production. Taken together, our study shows that simulated microgravity does not inhibit A. niger growth, but rather indicates a potential increase in surface-colonization. Further studies addressing fungal growth and surface contaminations in spaceflight should be conducted, not only to reduce the risk of negatively impacting human health and spacecraft material safety, but also to positively utilize fungal-based biotechnology to acquire needed resources in situ.

Elevated carbon assimilation and metabolic reprogramming in tomato high pigment mutants support the increased production of pigments.

High pigment mutants in tomato (Solanum lycopersicum L.), a loss of function in the control of photomorphogenesis, with greater pigment production, show altered growth, greater photosynthesis, and a metabolic reprogramming. High pigment mutations cause plants to be extremely responsive to light and produce excessive pigmentation as well as fruits with high levels of health-beneficial nutrients. However, the association of these traits with changes in the physiology and metabolism of leaves remains poorly understood. Here, we performed a detailed morphophysiological and metabolic characterization of high pigment 1 (hp1) and high pigment 2 (hp2) mutants in tomato (Solanum lycopersicum L. 'Micro-Tom') plants under different sunlight conditions (natural light, 50% shading, and 80% shading). These mutants occur in the DDB1 (hp1) and DET1 (hp2) genes, which are related to the regulation of photomorphogenesis and chloroplast development. Our results demonstrate that these mutations delay plant growth and height, by affecting physiological and metabolic parameters at all stages of plant development. Although the mutants were characterized by higher net CO2 assimilation, lower stomatal limitation, and higher carboxylation rates, with anatomical changes that favour photosynthesis, we found that carbohydrate levels did not increase, indicating a change in the energy flow. Shading minimized the differences between mutants and the wild type or fully reversed them in the phenotype at the metabolic level. Our results indicate that the high levels of pigments in hp1 and hp2 mutants represent an additional energy cost for these plants and that extensive physiological and metabolic reprogramming occurs to support increased pigment biosynthesis.

RNAi-induced knockdown of white gene in the southern green stink bug (Nezara viridula L.)

The southern green stink bug (SGSB) Nezara viridula L. is one of the most common stink bug species in the United States and can cause significant yield loss in a variety of crops. A suitable marker for the assessment of gene-editing tools in SGSB has yet to be characterized. The white gene, first documented in Drosophila, has been a useful target to assess the efficiency of introduced mutations in many species as it controls pigmentation processes and mutants display readily identifiable phenotypes. In this study we used the RNAi technique to investigate functions and phenotypes associated with the white ortholog in the SGSB and to validate white as a marker for genetic transformation in this species. This study revealed that white may be a suitable marker for germline transformation in the SGSB as white transcript knockdown was not lethal, did not impair embryo development and provided a distinguishable phenotype. Our results demonstrated that the white ortholog in SGSB is involved in the pathway for ommochrome synthesis and suggested additional functions of this gene such as in the integument composition, management of hemolymph compounds and riboflavin mobilization.

Effects of short‐wavelength ultraviolet ray irradiation on the early development and induction of pigmentation mutants of conchospore germlings in Pyropia haitanensis (Rhodophyta)

Ultraviolet mutagenesis is a safe and efficient method to induce mutations for breeding new germplasm of laver. In this study, high-frequency short-wave ultraviolet (UV-C) irradiation (λ = 254 nm) with different doses (50, 100, 200, 300, 400 and 500 J m−2) was used to induce colour mutants during the germination of conchospores in Pyropia haitanensis. The early development of conchospore germination and the morphogenesis of tetrad germlings were observed, which revealed that low-dose irradiation (50 J m−2) promoted the germination and early development of conchospores, while irradiation doses above 100 J m−2 inhibited the germination of conchospores and growth of tetrad germlings. In the dose range of 50–500 J m−2, the survival rate and germination rate of conchospores gradually decreased, while the mutation rate of colour mutants increased first and then decreased with increasing doses. The highest mutation rate was observed with the dose of 300 J m−2 or with irradiation time 72 s. These results indicated that the optimal UV-C irradiation dose was 300 J m−2 for the mutagenesis of P. haitanensis conchospores. In addition, UV-C irradiation also had a significant effect on the early development of conchospores and phenotypes of pigmentation mutant arranged in tetrad germlings, which was mainly manifested in the large number of colour-sectored blades produced by irradiated conchospores and in the decrease in their aspect ratios. This study provides new insights in carrying out UV-C mutagenesis for the breeding of P. haitanensis.

Contingent stimulus delivery assay for zebrafish reveals a role for CCSER1 in alcohol preference.

Alcohol use disorders are complex, multifactorial phenomena with a large footprint within the global burden of diseases. Here, we report the development of an accessible, two-choice self-administration zebrafish assay (SAZA) to study the neurobiology of addiction. Using this assay, we first demonstrated that, although zebrafish avoid higher concentrations of alcohol, they are attracted to low concentrations. Pre-exposure to alcohol did not change this relative preference, but acute exposure to an alcohol deterrent approved for human use decreased alcohol self-administration. A pigment mutant used in whole-brain imaging studies displayed a similar relative alcohol preference profile; however, mutants in CCSER1, a gene associated with alcohol dependence in human genetic studies, showed a reversal in relative preference. The presence of a biphasic response (hormesis) in zebrafish validated a key aspect of vertebrate responses to alcohol. SAZA adds a new dimension for discovering novel alcohol deterrents and studying the neurogenetics of addiction using the zebrafish.

Complete Genome Sequence of the Red-Pigmented Strain Serratia marcescens SCQ1 and Its Four Spontaneous Pigment Mutants.

Serratia marcescens SCQ1 is a red-pigmented bacterium isolated from silkworm larva with septicemia. Pigment-deficient spontaneous mutants arise when S. marcescens SCQ1 is incubated under relatively stable laboratory conditions for a long time. Here, we present the complete genome sequence of SCQ1 and the resequenced genomes of four spontaneous pigment mutants.

Development of a Model Mutagenesis System for Snapdragon

Snapdragon (Antirrhinum majus) has long been a very popular perennial in the United States due to its unique flower shape with a range of striking colors (Huo and Chen, 2018). Based on their height, snapdragons are typically classified into three categories: dwarf (6-15 inches), medium (1-2 feet), and tall (6-15 feet) . The dwarf variety has a dense, bushy growth pattern, producing numerous flower spikes. They grow on average 6 to 15 inches tall and are ideal plants for use as low borders or in containers. Mid-sized varieties grow 1-2 feet tall and are typically used in borders (either alone or with other bedding plants) and sometimes as cut flowers. Tall varieties range anywhere from 2 to 3 feet in height (Gilman et al. 2018). The magnificent flowers with a wide range of petal colors atop the long green spikes make the tall variety a desirable cut flower for container, bouquets, or gardens. In 2015, fresh-cut snapdragon sales increased 51.7% from 2010 and reached $12.93 million, making it a top ten fresh cut flower in United States（USDA, 2015).
 With all of their aesthetic attributes and versatility, snapdragons are also an important model system for genetics and molecular studies of various plant processes. For example, snapdragon pigmentation mutants produced by transposon (a type of mobile DNAs) mutagenesis have provided researchers a good way to study anthocyanin biosynthesis and subsequently aid plant breeders in developing new varieties with novel flower colors (Jackson et al. 1992). Furthermore, snapdragon has a mechanism by which transposable mutations can be regulated into active and inactive states through temperature control (Hashida et al., 2006). Advantages of this elegant transposon mutagenesis system and how it relates to plant breeding are described in this paper.

Speck, First Identified in Drosophila melanogaster in 1910, Is Encoded by the Arylalkalamine N-Acetyltransferase (AANAT1) Gene.

The pigmentation mutation speck is a commonly used recombination marker characterized by a darkly pigmented region at the wing hinge. Identified in 1910 by Thomas Hunt Morgan, speck was characterized by Sturtevant as the most “workable” mutant in the rightmost region of the second chromosome and eventually localized to 2-107.0 and 60C1-2. Though the first speck mutation was isolated over 110 years ago, speck is still not associated with any gene. Here, as part of an undergraduate-led research effort, we show that speck is encoded by the Arylalkylamine N-acetyltransferase 1 (AANAT1) gene. Both alleles from the Morgan lab contain a retrotransposon in exon 1 of the RB transcript of the AANAT1 gene. We have also identified a new insertion allele and generated multiple deletion alleles in AANAT1 that all give a strong speck phenotype. In addition, expression of AANAT1 RNAi constructs either ubiquitously or in the dorsal portion of the developing wing generates a similar speck phenotype. We find that speck alleles have additional phenotypes, including ectopic pigmentation in the posterior pupal case, leg joints, cuticular sutures and overall body color. We propose that the acetylated dopamine generated by AANAT1 decreases the dopamine pool available for melanin production. When AANAT1 function is decreased, the excess dopamine enters the melanin pathway to generate the speck phenotype.

A quantitative modelling approach to zebrafish pigment pattern formation.

Pattern formation is a key aspect of development. Adult zebrafish exhibit a striking striped pattern generated through the self-organisation of three different chromatophores. Numerous investigations have revealed a multitude of individual cell-cell interactions important for this self-organisation, but it has remained unclear whether these known biological rules were sufficient to explain pattern formation. To test this, we present an individual-based mathematical model incorporating all the important cell-types and known interactions. The model qualitatively and quantitatively reproduces wild type and mutant pigment pattern development. We use it to resolve a number of outstanding biological uncertainties, including the roles of domain growth and the initial iridophore stripe, and to generate hypotheses about the functions of leopard. We conclude that our rule-set is sufficient to recapitulate wild-type and mutant patterns. Our work now leads the way for further in silico exploration of the developmental and evolutionary implications of this pigment patterning system.

Identification, growth, and pigment content of a spontaneous green mutant of Pyropia kinositae (Bangiales, Rhodophyta)

A green-colored gametophytic blade of wild Pyropia was found in a natural population in the estuary area of the Naruse River, northeast Japan. To identify the isolated green mutant strain K-G1 and the wild-type strain K-1 derived from the same population, we carried out morphological observations and molecular analysis of the RUBISCO spacer region and the rbcL gene. As a result, the two strains were identified as Pyropia kinositae, which is a species closely related to the marine crop Pyropia yezoensis. Under the same laboratory culture, the growth and pigment contents of the green mutant K-G1 were compared with those of the wild-type K-1. The growth of the conchocelis and gametophytic blade tended to be slower in the green mutant than in the wild type, although differences in their growth were not apparent in some of the temperature conditions examined. Maturation of the conchocelis and blade in the two strains was fastest at 20 °C among the temperature conditions examined (5–25 °C). The phycoerythrin content was significantly lower in the green mutant than in the wild type. Additionally, the color phenotype of the green mutant was genetically stable in both the sporophytic conchocelis and gametophytic phases. Because the color of a pigmentation mutant facilitates cross-confirmation as a genetic marker in monoecious Pyropia species, these results suggest that the green mutant of P. kinositae will contribute as new breeding material for cross-breeding and polyploidy breeding in Pyropia.