The role of the epidermis enhancer element in positive and negative transcriptional regulation of ebony in Drosophila melanogaster

The spatiotemporal regulation of gene expression is essential to ensure robust phenotypic outcomes. Pigmentation patterns in Drosophila are determined by pigments biosynthesized in the developing epidermis and the cis-regulatory elements (CREs) of the genes involved in this process are well-characterized. Here we report that the known primary epidermal enhancer (priEE) is dispensable for the transcriptional activation of ebony (involved in light-colored pigment synthesis) in the developing epidermis of D. melanogaster. The evidence was obtained by introducing an approximately 1 kbp deletion at the priEE by genome editing. The effect of the priEE deletion on pigmentation and on the endogenous expression pattern of a mCherry-fused ebony allele was examined in the abdomen. The expression levels of the mCherry-fused ebony in the priEE-deleted strains were slightly higher than that of the control strain, indicating that the sequences outside the priEE have an ability to drive an expression of this gene in the epidermis. Interestingly, the priEE deletion resulted in a derepression of this gene in the dorsal midline of the abdominal tergites, where dark pigmentation is present in the wild-type individuals. This indicated that the priEE fragment contains a silencer. Furthermore, the endogenous expression pattern of ebony in the two additional strains with partially deleted priEE revealed that the silencer resides within a 351-bp fragment in the 5' portion of the priEE. These results demonstrated that deletion assays combined with reporter assays are highly effective in detecting the presence of positively and negatively regulating sequences within and outside the focal CREs.

The effects of high-monosaccharide diets on development and biochemical composition of white-eyed mutant strain of house cricket (Acheta domesticus)

Tryptophan (TRP) is one of the essential amino acids in the animal body. Its exogenicity and low concentrations mean that it can be regarded as one of the key regulatory molecules at the cellular as well as physiological level. It has been shown to have a number of essential functions, such as in the production of other biologically active molecules. The main objective of this project was to investigate the effects of a high monosaccharide diet (HMD) on a hemimetabolic insect—house cricket (Acheta domesticus) and a mutant strain with impaired visual pigment synthesis (closely related to the tryptophan and kynurenine (KYN) metabolic pathway)—white eye. This study was aimed at determining the effects of glucose and fructose on cricket development and biochemical composition. A parallel goal was to compare the response of both cricket strains to HMD. ELISA assays indicated dysfunction of the TRP-KYN pathway in white strain insects and an elevated KYN/TRP ratio. Biochemical analyses demonstrated the effects of HMD mainly on fat and glycogen content. A decrease in food intake was also observed in the groups on HMD. However, no changes in imago body weight and water content were observed. The results of the study indicate a stronger response of the white strain to HMD compared to the wild-type strain. At the same time, a stronger detrimental effect of fructose than of glucose was apparent. Sex was found to be a modulating factor in the response to HMD.

Genome-Wide Identification and Expression Analysis of R2R3-MYB Family Genes Associated with Petal Pigment Synthesis in Liriodendron

The MYB transcription factor family is one of the largest families in plants, and its members have various biological functions. R2R3-MYB genes are involved in the synthesis of pigments that yield petal colors. Liriodendron plants are widely cultivated as ornamental trees owing to their peculiar leaves, tulip-like flowers, and colorful petals. However, the mechanism underlying petal coloring in this species is unknown, and minimal information about MYB genes in Liriodendron is available. Herein, this study aimed to discern gene(s) involved in petal coloration in Liriodendron via genome-wide identification, HPLC, and RT-qPCR assays. In total, 204 LcMYB superfamily genes were identified in the Liriodendron chinense genome, and 85 R2R3-MYB genes were mapped onto 19 chromosomes. Chromosome 4 contained the most (10) R2R3-MYB genes, and chromosomes 14 and 16 contained the fewest (only one). MEME analysis showed that R2R3-MYB proteins in L. chinense were highly conserved and that their exon-intron structures varied. The HPLC results showed that three major carotenoids were uniformly distributed in the petals of L. chinense, while lycopene and β-carotene were concentrated in the orange band region in the petals of Liriodendron tulipifera. Furthermore, the expression profiles via RT-qPCR assays revealed that four R2R3-MYB genes were expressed at the highest levels at the S3P/S4P stage in L. tulipifera. This result combined with the HPLC results showed that these four R2R3-MYB genes might participate in carotenoid synthesis in the petals of L. tulipifera. This work laid a cornerstone for further functional characterization of R2R3-MYB genes in Liriodendron plants.

ASIP promoter variants predict sesame coat color in Shiba Inu dogs

Different patterns of coat color pigmentation in dogs are produced by a sophisticated interaction of several genes. Understanding the mechanisms underlying the diversity of coat colors and their inheritance is important for professional breeders because it helps to predict the phenotypes of the progeny. Although genetics of the main coat colors in dogs is extensively studied, there are types of coat pigmentation that are not explained yet. Recently a new model connected the variants in ASIP gene promoters with different coat colors in dogs. Here we used this model as a framework to investigate the genetics of the rare sesame coat color in Shiba Inu dogs. We determined the combination of two alleles of ASIP gene that determine sesame coat color. This finding can be used by the breeders to produce the dogs with this rare coat color pattern. We also demonstrate the incomplete dominance between the ASIP alleles involved in sesame coat formation. These results are in good agreement with the new model explaining how different levels of ASIP gene expression affect the regulation of pigment synthesis in melanocytes.

The vitamin a transporter STRA6 adjusts the stoichiometry of chromophore and opsins in visual pigment synthesis and recycling

The retinal pigment epithelium of the vertebrate eyes acquires vitamin A from circulating retinol binding protein for chromophore biosynthesis. The chromophore covalently links with an opsin protein in the adjacent photoreceptors of the retina to form the bipartite visual pigment complexes. We here analyzed visual pigment biosynthesis in mice deficient for the retinol binding protein receptor STRA6. We observed that chromophore content was decreased throughout the life cycle of these animals, indicating that lipoprotein-dependent delivery pathways for the vitamin cannot substitute for STRA6. Changes in the expression of photoreceptor marker genes, including a down-regulation of the genes encoding rod and cone opsins, paralleled the decrease in ocular retinoid concentration in STRA6-deficient mice. Despite this adaptation, cone photoreceptors displayed absent or mislocalized opsins at all ages examined. Rod photoreceptors entrapped the available chromophore but exhibited significant amounts of chromophore-free opsins in the dark-adapted stage. Treatment of mice with pharmacological doses of vitamin A ameliorated the rod phenotype but did not restore visual pigment synthesis in cone photoreceptors of STRA6-deficient mice. The imbalance between chromophore and opsin concentrations of rod and cone photoreceptors was associated with an unfavorable retinal physiology, including diminished electrical responses of photoreceptors to light, and retinal degeneration during aging. Together, our study demonstrates that STRA6 is critical to adjust the stoichiometry of chromophore and opsins in rod cone photoreceptors and to prevent pathologies associated with ocular vitamin A deprivation.

Identification and Characterization of Verticillium nonalfalfae-Responsive MicroRNAs in the Roots of Resistant and Susceptible Hop Cultivars

MicroRNAs are 21- to 24-nucleotide-long, non-coding RNA molecules that regulate gene expression at the post-transcriptional level. They can modulate various biological processes, including plant response and resistance to fungal pathogens. Hops are grown for use in the brewing industry and, recently, also for the pharmaceutical industry. Severe Verticillium wilt caused by the phytopathogenic fungus Verticillium nonalfalfae, is the main factor in yield loss in many crops, including hops (Humulus lupulus L.). In our study, we identified 56 known and 43 novel miRNAs and their expression patterns in the roots of susceptible and resistant hop cultivars after inoculation with V. nonalfalfae. In response to inoculation with V. nonalfalfae, we found five known and two novel miRNAs that are differentially expressed in the susceptible cultivar and six known miRNAs in the resistant cultivar. Differentially expressed miRNAs target 49 transcripts involved in protein localization and pigment synthesis in the susceptible cultivar, whereas they are involved in transcription factor regulation and hormone signalling in the resistant cultivar. The results of our study suggest that the susceptible and resistant hop cultivars respond differently to V. nonalfalfae inoculation at the miRNA level and that miRNAs may contribute to the successful defence of the resistant cultivar.

Identification and Characterization of Verticillium Nonalfalfae-responsive Micrornas in the Roots of Resistant and Susceptible Hop Cultivars

Micro RNAs are 21- to 24-nucleotide-long, non-coding RNA molecules that regulate gene expression at the post-transcriptional level. They can modulate various biological processes, including plant response and resistance to fungal pathogens. Hops are grown for use in the brewing industry and recently also for pharmaceutical industry. Severe Verticillium wilt caused by the phytopathogenic fungus Verticillium nonalfalfae, is the main factor in yield loss in many crops, including hops (Humulus lupulus L.). In our study, we identified 56 known and 43 novel miRNAs and their expression patterns in the roots of susceptible and resistant hop cultivars after inoculation with V. nonalfalfae. In response to inoculation with V. nonalfalfae, we found five known and two novel miRNAs that are differentially expressed in the susceptible cultivar and six known miRNAs in the resistant cultivar. Differentially expressed miRNAs target 49 transcripts involved in protein localization and pigment synthesis in the susceptible cultivar, whereas they are involved in transcription factor regulation and hormone signalling in the resistant cultivar. The results of our study suggest that the susceptible and resistant hop cultivars respond differently to V. nonalfalfae inoculation at the miRNA level and that miRNAs may contribute to the successful defence of the resistant cultivar.

Bioherbicides: An Eco-Friendly Tool for Sustainable Weed Management

Weed management is an arduous undertaking in crop production. Integrated weed management, inclusive of the application of bioherbicides, is an emerging weed control strategy toward sustainable agriculture. In general, bioherbicides are derived either from plants containing phytotoxic allelochemicals or certain disease-carrying microbes that can suppress weed populations. While bioherbicides have exhibited great promise in deterring weed seed germination and growth, only a few in vitro studies have been conducted on the physiological responses they evoke in weeds. This review discusses bioherbicide products that are currently available on the market, bioherbicide impact on weed physiology, and potential factors influencing bioherbicide efficacy. A new promising bioherbicide product is introduced at the end of this paper. When absorbed, phytotoxic plant extracts or metabolites disrupt cell membrane integrity and important biochemical processes in weeds. The phytotoxic impact on weed growth is reflected in low levels of root cell division, nutrient absorption, and growth hormone and pigment synthesis, as well as in the development of reactive oxygen species (ROS), stress-related hormones, and abnormal antioxidant activity. The inconsistency of bioherbicide efficacy is a primary factor restricting their widespread use, which is influenced by factors such as bioactive compound content, weed control spectrum, formulation, and application method.

optix is involved in eyespot development via a possible positional information mechanism

optix, a gene essential and sufficient for eye development in Drosophila melanogaster, also plays important roles in the development of both the structure and pigmentation of butterfly wing scales. In particular, optix regulates wing scale lower lamina thickness and ommochrome pigment synthesis. Here we explore the role of optix in wing pattern development of Bicyclus anynana butterflies by examining its expression using immunostainings and testing its function via CRISPR-Cas9. We found Optix to be expressed in multiple domains, most prominently in the orange ring of the eyespots and in other scattered orange scales, and to regulate the pigmentation and the development of the upper lamina of the orange scales. We further explored the interaction of Optix with Spalt, a protein involved in the development of black scales in the eyespots, and expressed adjacent to the Optix domain. CRISPR knockouts of optix or spalt, followed by immunostainings, showed that Spalt represses optix expression in cells of the central black region of the eyespot. This regulatory interaction mimics that found in the anterior compartment of the wing disc where both genes respond to Decapentaplegic (Dpp) signaling and play a role in venation patterning. Using in situ hybridizations we show that dpp is expressed in the center of the eyespots and propose that this same circuit might have been recruited for eyespot development where Decapentaplegic acts as a central morphogen, activating optix and spalt at different concentration thresholds, and where spalt cross-regulates optix resulting in the formation of a sharp boundary between the two eyespot color rings.