Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-Associated Protein 9 Mediated Knockout Reveals Functions of the yellow-y Gene in Spodoptera litura

Liu, Xiaolong; Han, Wei‐Kang; Ze, Long-Ji; Peng, Yajun; Yang, Y.; Jin, Zhang; Yan, Qi; Dong, Shuang-Lin

doi:10.3389/fphys.2020.615391

Cited by 14 publications

(16 citation statements)

References 42 publications

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“…In particular, a functional importance of yellow-y ( Y-y ) in cuticle pigmentation/melanization appears to be highly conserved among insects. A reduction in function of Y-y caused lighter and yellowish cuticle, including species-specific body markings/spots in many insect species from several orders, such as Diptera, Lepidoptera, Coleoptera, Hymenoptera, Hemiptera, and Blattodea ( Wittkopp et al, 2002 ; Futahashi et al, 2008 ; Arakane et al, 2010 ; Shirataki et al, 2010 ; Riedel et al, 2011 ; Liu et al, 2016 ; Heinze et al, 2017 ; Chen et al, 2018 ; Paulo et al, 2019 ; Liu et al, 2020 ; Mun et al, 2020 ; Wang et al, 2020 ; Han et al, 2021 ; Nguyen et al, 2021 ; Nie et al, 2021 ; Shirai et al, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

Yellow-y Functions in Egg Melanization and Chorion Morphology of the Asian Tiger Mosquito, Aedes albopictus

Noh

Mun

Kramer

et al. 2021

Front. Cell Dev. Biol.

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The Asian tiger mosquito, Aedes albopictus, is one of the most serious public health pests, which can transmit various vector-borne diseases. Eggs from this mosquito species become dark black shortly after oviposition and exhibit high desiccation resistance. Some of the Yellow proteins that act as dopachrome conversion enzymes (DCEs) are involved in the tyrosine-mediated tanning (pigmentation and sclerotization) metabolic pathway that significantly accelerates melanization reactions in insects. In this research, we analyzed the function of one of the yellow genes, yellow-y (AalY-y), in eggshell/chorion melanization of Ae. albopictus eggs. Developmental and tissue-specific expression measured by real-time PCR showed that AalY-y transcripts were detected at all stages of development analyzed, with significantly higher levels in the ovaries from blood-fed adult females. Injection of double-stranded RNA for AalY-y (dsAalY-y) had no significant effect on fecundity. However, unlike dsEGFP-treated control eggs that become black by 2–3 h after oviposition (HAO), dsAalY-y eggs were yellow-brown at 2 HAO, and reddish-brown even at 48 HAO. dsEGFP eggs exhibited resistance to desiccation at 48 HAO, whereas approximately 50% of the dsAalY-y eggs collapsed when they were moved to a low humidity condition. In addition, TEM analysis revealed an abnormal morphology and ultrastructure of the outer-endochorion in the dsAalY-y eggs. These results support the hypothesis that AalY-y is involved in the tyrosine-induced melanin biosynthetic pathway, plays an important role in black melanization of the chorion and functions in conferring proper morphology of the outer-endochorion, a structure that is presumably required for egg desiccation resistance in Ae. albopictus.

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Section: Introductionmentioning

confidence: 99%

Yellow-y Functions in Egg Melanization and Chorion Morphology of the Asian Tiger Mosquito, Aedes albopictus

Noh

Mun

Kramer

et al. 2021

Front. Cell Dev. Biol.

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Section: Resultsmentioning

confidence: 99%

“…Further, unlike Bombyx, yellow-y knockouts in Agrotis are susceptible to dehydration (Chen et al . 2018) and mutants in Spodoptera exhibit defects in body development, copulation, oviposition, and hatchability (Liu et al . 2020, Han et al .…”

Section: Resultsmentioning

confidence: 99%

“…Comparative data from other species suggests that yellow-y functions differently among lepidopteran species. For example, while yellow-y loss-of-function is also found to be recessive in several other lepidopteran species (Liu et al 2020, Wang, Huang, et al 2020, Han et al 2021, Shirai et al 2021, it is dominant in the black cutworm, Agrotis ipsilon (Chen et al 2018). Further, unlike Bombyx, yellow-y knockouts in Agrotis are susceptible to dehydration (Chen et al 2018) and mutants in Spodoptera exhibit defects in body development, copulation, oviposition, and hatchability (Liu et al 2020, Han et al 2021, Shirai et al 2021.…”

Section: Resultsmentioning

confidence: 99%

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Allele-specific knockouts reveal a role for apontic-like in the evolutionary loss of larval pigmentation in the domesticated silkworm, Bombyx mori

Tomihara

Andolfatto

Kiuchi

2022

Preprint

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The domesticated silkworm, Bombyx mori, and its wild progenitor, B. mandarina, are extensively studied as a model case of the evolutionary process of domestication. A conspicuous difference between these species is the dramatic reduction in pigmentation in both larval and adult B. mori. Here we evaluate the efficiency of CRISPR/Cas9-targeted knockouts of pigment-related genes as a tool to understand their potential contributions to domestication-associated pigmentation loss in B. mori. To demonstrate the efficacy of targeted knockouts in B. mandarina, we generated a homozygous CRISPR/Cas9-targeted knockout of yellow-y. In yellow-y knockout mutants, black body color became lighter throughout the larval, pupal and adult stages, confirming a role for this gene in pigment formation. Further, we performed allele-specific CRISPR/Cas9-targeted knockouts of the pigment-related transcription factor, apontic-like (apt-like) in B. mori × B. mandarina F1 hybrid individuals. Knockout of the B. mandarina allele of apt-like in F1 embryos results in depigmented patches on the dorsal integument of larvae, whereas corresponding knockouts of the B. mori allele consistently exhibit normal F1 larval pigmentation. These results demonstrate a contribution of apt-like to the evolution of reduced pigmentation in B. mori. Together, our results demonstrate the feasibility of CRISPR/Cas9-targeted knockouts as a tool for understanding the genetic basis of traits associated with B. mori domestication.Brief abstractBombyx mori and its wild progenitor are an important model for the study of phenotypic evolution associated with domestication. As proof-of-principle, we used CRISPR/Cas9 to generate targeted knockouts of two pigmentation-related genes. By generating a homozygous knockout of yellow-y in B. mandarina, we confirmed this gene”s role in pigment formation. Further, by generating allele-specific knockouts of apontic-like (apt-like) in B. mori × B. mandarina F1 hybrids, we establish that evolution of apt-like contributed to reduced pigmentation during B. mori domestication.Graphical TOC/Abstract

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“…In insects, yellow s are a rapidly evolving gene family generating functionally diverse paralogs (Ferguson et al, 2011; Massey et al, 2019). One of the physiological roles of insect Yellows is cuticle pigmentation/melanization during ecdysis (Futahashi & Osanai‐Futahashi, 2021; Liu et al, 2020). Biochemically, the pigmentation/melanization of insect cuticle derives from a combination of melanins (black dopa‐melanin and brown dopamine‐melanin) and sclerotins (yellow NBAD [β‐alanyl dopamine]‐sclerotin and colorless NADA [N‐acetyl dopamine]‐sclerotin) (Matsuoka & Monteiro, 2018; Mun et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

Yellow‐b, ‐c, ‐e and ‐h are required for normal body coloration in Henosepilachna vigintioctopunctata

Wang

Jin

et al. 2021

Arch Insect Biochem Physiol

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The involvement of yellow genes y-b, y-c, y-e, and y-h in cuticle tanning has poorly been clarified. In the present paper, six putative yellow (y-y, y-b, y-c, y-e y-f, and y-h) genes were identified in Henosepilachna vigintioctopunctata.Hvy-b, Hvy-c, Hvy-e, and Hvy-h were abundantly transcribed at early larval and late pupal stages, especially in the epidermis. Accordingly, RNA interference (RNAi) experiments were performed by an injection of dsy-b, dsy-c, dsye, or dsy-h into the second instar larvae and 1-day-old pupae. The head capsule, scoli and strumae, and legs in the fourth-instar larvae became blacker; the blackish spots in the pupae were darkened and widened after RNAi of Hvy-b, compared with those of dsegfp-treated controls.Depletion of Hvy-b at the 1-day-old pupal stage expanded two pair of black markings on the sternum of the metathorax, and darkened the black patched on the sterna of the abdomen segments I-VI in the resultant adults.Depletion of Hvy-e caused darker pigmented adult body and elytral cuticles than those of dsegfp-introduced controls. However, there was no obvious difference in pigmentation of the black markings. Hvy-h-deficient larvae displayed dark yellow body color, whereas the body color of the dsegfp-injected control was pale yellow. There was no obvious difference in coloration of larval specific-black markings or pupal cuticle between dsHvy-h-and dsegfptreated animals. Moreover, silence of Hvy-c at the second

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Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-Associated Protein 9 Mediated Knockout Reveals Functions of the yellow-y Gene in Spodoptera litura

Cited by 14 publications

References 42 publications

Yellow-y Functions in Egg Melanization and Chorion Morphology of the Asian Tiger Mosquito, Aedes albopictus

Yellow-y Functions in Egg Melanization and Chorion Morphology of the Asian Tiger Mosquito, Aedes albopictus

Allele-specific knockouts reveal a role for apontic-like in the evolutionary loss of larval pigmentation in the domesticated silkworm, Bombyx mori

Yellow‐b, ‐c, ‐e and ‐h are required for normal body coloration in Henosepilachna vigintioctopunctata

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