Vitellogenesis is one of the most well-studied physiological processes in mosquitoes. Expression of mosquito vitellogenin genes is classically described as being restricted to female adult reproduction. We report premature vitellogenin transcript expression in three vector mosquitoes: Culex tarsalis, Aedes aegypti and Anopheles gambiae. Vitellogenins expressed during non-reproductive stages are alternatively spliced to retain their first intron and encode premature termination codons. We show that intron retention results in transcript degradation by translation-dependent nonsense-mediated mRNA decay. This is probably an example of regulated unproductive splicing and translation (RUST), a mechanism known to regulate gene expression in numerous organisms but which has never been described in mosquitoes. We demonstrate that the hormone 20-hydroxyecdysone (20E) is responsible for regulating post-transcriptional splicing of vitellogenin. After exposure of previtellogenic fat bodies to 20E, vitellogenin expression switches from a non-productive intron-retaining transcript to a spliced protein-coding transcript. This effect is independent of factors classically known to influence transcription, such as juvenile hormone-mediated competence and amino acid signalling through the target of rapamycin pathway. Non-canonical regulation of vitellogenesis through RUST is a novel role for the multifunctional hormone 20E, and may have important implications for general patterns of gene regulation in mosquitoes.