Identification and Functional Insights of Knickkopf Genes in the Larval Cuticle of Leptinotarsa decemlineata.

Simple Summary: The Colorado potato beetle, Leptinotarsa decemlineata, poses a significant threat to potato crops worldwide, causing substantial agricultural damage. To better understand how these pests develop and potentially find ways to control them, we studied a group of genes called Knickkopf (Knk) genes in the larvae of the beetle. These genes are crucial for forming and maintaining the insect's outer shell, known as the cuticle, which protects them from the environment. We identified four LdKnk-family genes in the beetle and analyzed when and where these genes are expressed during the insect's growth stages. By using RNA interference (RNAi) to silence these genes, we observed that larvae became weaker, with damaged cuticles and higher death rates. These findings indicate that LdKnk-family genes are vital for the beetle's survival and development. Targeting LdKnk-family genes could provide a new strategy for pest control, potentially weakening the beetles' defenses and making them more susceptible to environmental factors and insecticides, ultimately helping to protect potato crops. The Colorado potato beetle (Leptinotarsa decemlineata) is a major pest of potato crops. While Knickkopf (Knk) genes are essential for insect cuticle formation, their roles in pests like L. decemlineata remain unclear. This study aims to identify and characterize Knk genes in L. decemlineata and explore their functions in larval development and cuticle integrity. We used genomic and transcriptomic databases to identify LdKnk-family genes, validated through RT-PCR and RACE. Gene expression was analyzed at various developmental stages and tissues using qRT-PCR. RNA interference (RNAi) and Transmission electron microscopy (TEM) were applied to determine the functional roles of these genes. Four LdKnk-family genes were identified. Spatio-temporal expression analysis indicated significant gene expression during larval molting and pupal stages, especially in the epidermis. RNAi experiments showed that silencing LdKnk and LdKnk3-5′ led to reduced larval weight, cuticle thinning, and increased mortality, while LdKnk3-FL knockdown caused abnormal cuticle thickening and molting disruptions. LdKnk2 knockdown increased epicuticle and endocuticle thickness without visible phenotypic changes. The study highlights the essential roles of LdKnk-family genes in maintaining cuticle structure and integrity, suggesting their potential as targets for RNAi-based pest control. [ABSTRACT FROM AUTHOR]