The cuticle is an indispensable part of insects [1,2]. Its functions include resisting external environmental damage; forming, and maintaining the insect body shape; and acting as the major barrier to infection [[3], [4], [5]]. Aedes aegypti undergoes complete metamorphosis in its life cycle, with cuticle formation, differentiation, digestion, and reconstruction being parts of this process [[6], [7], [8], [9], [10]]. The insect cuticle is composed of three layers, the epicuticle, exocuticle, and endocuticle, as well as epidermal cells. Researchers have found that cuticular proteins (CPs) affect the structure and mechanical properties of the cuticle [[6], [7], [8], [9], [10]]. Upregulation of genes encoding some structural CPs can induce apoptosis by causing structural abnormalities in epidermal cells and endocuticles [11,12]. The largest CP family in insects is the CPR family, which contains the Rebers and Riddiford (R&R) consensus sequence [13,14]. RR-1 proteins participate in endocuticle (flexible cuticle) formation, while RR-2 proteins are involved in outer cuticle (hard cuticle) formation [10,[15], [16], [17], [18]].

The CPR family play important role in insects. RNAi of CYP4g1 (aldehyde oxidative decarbonylase P450) and NADPH-cytochrome P450 reductase (CPR) in Drosophila melanogaster has been found to block the biosynthesis of hydrocarbons in oenocytes [19]. Silencing a cuticular gene containing the RR-1 motif in locusts was found to cause thickening of the inner cuticular layer of larvae. Similarly, silencing an RR-1 motif-containing CPR in silkworms was found to reduce the extensibility and elasticity of the larval cuticle and thereby lead to a taut body and an inability to crawl, bend, or grip [20]. In Cryptopygus antarcticus and Onychiurus arcticus, the upregulation of CPR genes in cold regions showed that they play a role in the resistance of these insects to cold stress [21,22].

The serosa, which is the initial layer of cellular epithelial tissue located on the inner side of the eggshell, plays a vital role in protecting the egg from desiccation and other environmental factors [23,24]. The serosal cuticle has already formed 20 h after the egg has been laid and enables the egg to survive for several months in a dry environment [24]. The study of various insect serosal cuticles revealed that the serosal cuticle is a layered structure similar to the cuticle [25,26]. RNAi of chitin-synthase1 in Tribolium truncatum resulted in the inability of the serosal cuticle within the egg to form a layered structure [23]. Similarly, researchers, such as Arakane, observed that the egg production and hatching rate of females was affected upon interfering with chitin synthase in adult red rice beetles [27]. Additionally, it is speculated that cuticular structural proteins may participate in serosa construction. Transcriptomic data of wild type eggs and eggs without a serosa revealed that six genes encoding cuticular structural proteins were significantly downregulated in eggs without a serosa [28].

AaCPR100A is a cuticular protein found in Ae. aegypti and is an RR-1 motif-containing CPR protein. It was identified using the Ae. aegypti genome and RNA-seq data analysis [29]. Findings suggest that it may play vital roles in the soft cuticle region, especially that of the eggs and pupa. RNA interference of the gene encoding AaCPR100A resulted in mortality. It is of interest to know how AaCPR100A functions, especially in cuticle formation: whether it directly affects the formation of chitins or lipids in the cuticle or it requires partner proteins to enable their formation. In the present study, we investigated the AaCPR100A gene expression and protein localization, the interaction proteins of AaCPR100A, and phenotypes of AaCPR100A gene-silenced mosquitoes using RNAi-mediated gene knockdown. Our findings highlight AaCPR100A as a promising target for the development of innovative mosquito control techniques.