RNAi-Mediated Manipulation of Cuticle Coloration Genes in Lygus hesperus Knight (Hemiptera: Miridae)

Abstract

Cuticle coloration in insects is a consequence of the accumulation of pigments in a species-specific pattern. Numerous genes are involved in regulating the underlying processes of melanization and sclerotization, and their manipulation can be used to create externally visible markers of successful gene editing. To clarify the roles for many of these genes and examine their suitability as phenotypic markers in Lygus hesperus Knight (western tarnished plant bug), transcriptomic data were screened for sequences exhibiting homology with the Drosophila melanogaster proteins. Complete open reading frames encoding putative homologs for six genes (aaNAT, black, ebony, pale, tan, and yellow) were identified, with two variants for black. Sequence and phylogenetic analyses supported preliminary annotations as cuticle pigmentation genes. In accord with observable difference in color patterning, expression varied for each gene by developmental stage, adult age, body part, and sex. Knockdown by injection of dsRNA for each gene produced varied effects in adults, ranging from the non-detectable (black 1, yellow), to moderate decreases (pale, tan) and increases (black 2, ebony) in darkness, to extreme melanization (aaNAT). Based solely on its expression profile and highly visible phenotype, aaNAT appears to be the best marker for tracking transgenic L. hesperus.

Efficient nanoparticle‐based CRISPR‐Cas13d induced mRNA disruption of an eye pigmentation gene in the white‐backed planthopper, Sogatella furcifera

“…Similar knockdown of the Tenebrio molitor gene eliminated eye color (Oppert et al, 2019) and CRISPR-Cas9 knockout of TO genes in Spodoptera frugiperda (Zhu et al, 2020a), Helicoverpa zea (Perera et al, 2018), and Tribolium castaneum (Adrianos et al, 2018) yielded atypical eye pigmentation phenotypes. In the hemipteran Lygus hesperus, knockdown of vermillion/TO transcripts resulted in reddening around the eye margins (Brent et al, 2022) . This degree of conservation suggests that the TO gene could function as a useful visual marker for gene editing.…”

“…, 2018) yielded atypical eye pigmentation phenotypes. In the hemipteran Lygus hesperus , knockdown of vermillion/TO transcripts resulted in reddening around the eye margins (Brent et al ., 2022) . This degree of conservation suggests that the TO gene could function as a useful visual marker for gene editing.…”