Comprehensive microarray-based analysis for stage-specific larval camouflage pattern-associated genes in the swallowtail butterfly, Papilio xuthus

<p>Abstract</p> <p>Background</p> <p>Body coloration is an ecologically important trait that is often involved in prey-predator interactions through mimicry and crypsis. Although this subject has attracted the interest of biologists and the general public, our scientific knowledge on the subject remains fragmentary. In the caterpillar of the swallowtail butterfly <it>Papilio xuthus</it>, spectacular changes in the color pattern are observed; the insect mimics bird droppings (mimetic pattern) as a young larva, and switches to a green camouflage coloration (cryptic pattern) in the final instar. Despite the wide variety and significance of larval color patterns, few studies have been conducted at a molecular level compared with the number of studies on adult butterfly wing patterns.</p> <p>Results</p> <p>To obtain a catalog of genes involved in larval mimetic and cryptic pattern formation, we constructed expressed sequence tag (EST) libraries of larval epidermis for <it>P. xuthus</it>, and <it>P. polytes </it>that contained 20,736 and 5,376 clones, respectively, representing one of the largest collections available in butterflies. A comparison with silkworm epidermal EST information revealed the high expression of putative blue and yellow pigment-binding proteins in <it>Papilio </it>species. We also designed a microarray from the EST dataset information, analyzed more than five stages each for six markings, and confirmed spatial expression patterns by whole-mount <it>in situ </it>hybridization. Hence, we succeeded in elucidating many novel marking-specific genes for mimetic and cryptic pattern formation, including pigment-binding protein genes, the melanin-associated gene <it>yellow-h3</it>, the ecdysteroid synthesis enzyme gene <it>3-dehydroecdysone 3b-reductase</it>, and <it>Papilio</it>-specific genes. We also found many cuticular protein genes with marking specificity that may be associated with the unique surface nanostructure of the markings. Furthermore, we identified two transcription factors, <it>spalt </it>and ecdysteroid signal-related <it>E75</it>, as genes expressed in larval eyespot markings. This finding suggests that <it>E75 </it>is a strong candidate mediator of the hormone-dependent coordination of larval pattern formation.</p> <p>Conclusions</p> <p>This study is one of the most comprehensive molecular analyses of complicated morphological features, and it will serve as a new resource for studying insect mimetic and cryptic pattern formation in general. The wide variety of marking-associated genes (both regulatory and structural genes) identified by our screening indicates that a similar strategy will be effective for understanding other complex traits.</p

Catalogue of epidermal genes: Genes expressed in the epidermis during larval molt of the silkworm Bombyx mori

<p>Abstract</p> <p>Background</p> <p>The insect cuticle is composed of various proteins and formed during the molt under hormonal regulation, although its precise composition and formation mechanism are largely unknown. The exhaustive catalogue of genes expressed in epidermis at the molt constitutes a massive amount of information from which to draw a complete picture of the molt and cuticle formation in insects. Therefore, we have catalogued a library of full-length cDNAs (designated epM) from epidermal cells during the last larval molt of <it>Bombyx mori</it>.</p> <p>Results</p> <p>Of the 10,368 sequences in the library, we isolated 6,653 usable expressed sequence tags (ESTs), which were categorized into 1,451 nonredundant gene clusters. Seventy-one clusters were considered to be isoforms or premature forms of other clusters. Therefore, we have identified 1,380 putative genes. Of the 6,653 expressed sequences, 48% were derived from 92 cuticular protein genes (RR-1, 24; RR-2, 17; glycine-rich, 29; other classes, 22). A comparison of epM with another epidermal EST data set, epV3 (feeding stage: fifth instar, day 3), showed marked differences in cuticular protein gene. Various types of cuticular proteins are expressed in epM but virtually only RR-1 proteins were expressed in epV3. Cuticular protein genes expressed specifically in epidermis, with several types of expression patterns during the molt, suggest different types of responses to the ecdysteroid pulse. Compared with other <it>Bombyx </it>EST libraries, 13 genes were preferentially included in epM data set. We isolated 290 genes for proteins other than cuticular proteins, whose amino acid sequences retain putative signal peptides, suggesting that they play some role in cuticle formation or in other molting events. Several gene groups were also included in this data set: hormone metabolism, P450, modifier of cuticular protein structure, small-ligand-binding protein, transcription factor, and pigmentation genes.</p> <p>Conclusion</p> <p>We have identified 1,380 genes in epM data set and 13 preferentially expressed genes in epidermis at the molt. The comparison of the epM and other EST libraries clarified the totally different gene expression patterns in epidermis between the molting and feeding stages and many novel tissue- and stage-specifically expressed epidermal genes. These data should further our understanding of cuticle formation and the insect molt.</p

Repression of tyrosine hydroxylase is responsible for the sex-linked chocolate mutation of the silkworm, \u3cem\u3eBombyx mori\u3c/em\u3e

Pigmentation patterning has long interested biologists, integrating topics in ecology, development, genetics, and physiology. Wild-type neonatal larvae of the silkworm, Bombyx mori, are completely black. By contrast, the epidermis and head of larvae of the homozygous recessive sex-linked chocolate (sch) mutant are reddish brown. When incubated at 30 °C, mutants with the sch allele fail to hatch; moreover, homozygous mutants carrying the allele sch lethal (schl) do not hatch even at room temperature (25 °C). By positional cloning, we narrowed a region containing sch to 239,622 bp on chromosome 1 using 4,501 backcross (BC1) individuals. Based on expression analyses, the best sch candidate gene was shown to be tyrosine hydroxylase (BmTh). BmTh coding sequences were identical among sch, schl, and wild-type. However, in sch the ∼70-kb sequence was replaced with ∼4.6 kb of a Tc1-mariner type transposon located ∼6 kb upstream of BmTh, and in schl, a large fragment of an L1Bm retrotransposon was inserted just in front of the transcription start site of BmTh. In both cases, we observed a drastic reduction of BmTh expression. Use of RNAi with BmTh prevented pigmentation and hatching, and feeding of a tyrosine hydroxylase inhibitor also suppressed larval pigmentation in the wild-type strain, pnd+ and in a pS (black-striped) heterozygote. Feeding L-dopa to sch neonate larvae rescued the mutant phenotype from chocolate to black. Our results indicate the BmTh gene is responsible for the sch mutation, which plays an important role in melanin synthesis producing neonatal larval color

RNA interference in Lepidoptera: an overview of successful and unsuccessful studies and implications for experimental design

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Color vision and color formation in dragonflies

Dragonflies including damselflies are colorful and large-eyed insects, which show remarkable sexual dimorphism, color transition, and color polymorphism. Recent comprehensive visual transcriptomics has unveiled an extraordinary diversity of opsin genes within the lineage of dragonflies. These opsin genes are differentially expressed between aquatic larvae and terrestrial adults, as well as between dorsal and ventral regions of adult compound eyes. Recent topics of color formation in dragonflies are also outlined. Non-iridescent blue color is caused by coherent light scattering from the quasiordered nanostructures, whereas iridescent color is produced by multilayer structures. Wrinkles or wax crystals sometimes enhances multilayer structural colors. Sex-specific and stage-specific color differences in red dragonflies is attributed to redox states of ommochrome pigments

Two new subspecies of Hemicordulia tenera Lieftinck, 1930 (Corduliidae) from Cambodia and Thailand

Hemicordulia tenera donnellyi ssp. nov. (holotype ♂: Chieng Mai Prov., Kunklang: highway 1009, Restaurant; 16°32.0’ N 98°31.3’ E, 1000 m, 22 v 1996, FSCA) and H. t. vikhrevi ssp. nov. (holotype ♂: Cambodia, Koh Kong Province, ~13 km ENE of Koh Kong, ‘Hemicordulia brook’, 11°39’55’’ N, 103°05’34’’ E, 315 m, 04 xii 2010, RMNH) are described from North Thailand and South-West Cambodia, respectively. The nominotypical H. tenera tenera Lieftinck, 1930 is distributed in the Malay Peninsula, Borneo, Java and Sumatra. Although these three subspecies are genetically very close, they are distinguishable by the relative length and shape of the caudal appendages

The genus Planaeschna McLachlan, 1895 and its subgroupings in Vietnam, with descriptions of three new species (Odonata: Aeshnidae)

Kompier, Tom, Karube, Haruki, Futahashi, Ryo, Phan, Quoc Toan (2021): The genus Planaeschna McLachlan, 1895 and its subgroupings in Vietnam, with descriptions of three new species (Odonata: Aeshnidae). Zootaxa 5027 (1): 1-35, DOI: https://doi.org/10.11646/zootaxa.5027.1.

Butterfly wing color made of pigmented liquid

Summary: A previously undescribed mechanism underlying butterfly wing coloration patterns was discovered in two distantly related butterfly species, Siproeta stelenes and Philaethria diatonica. These butterflies have bright green wings, but the color pattern is not derived from solid pigments or nanostructures of the scales or from the color of the cuticular membrane but rather from a liquid retained in the wing membrane. Wing structure differs between the green and non-green areas. In the non-green region, the upper and lower cuticular membranes are attached to each other, whereas in the green region, we observed a space of 5–10 μm where green liquid is held and living cells are present. A pigment analysis and tracer experiment revealed that the color of the liquid is derived from hemolymph components, bilin and carotenoid pigments. This discovery broadens our understanding of the diverse ways in which butterfly wings obtain their coloration and patterns

PsDefensin1 GNBPRNAi and Ec

Relative expression level to L32 (fig. 5a)