Identification of Genes Displaying Sex- and Tissue-biased Expression Patterns in the Monarch Butterfly (Danaus plexippus) with an Emphasis on Chemosensory Perception

Monarch butterflies (Danaus plexippus) rely on milkweeds as larval host plants. Host plant seeking and verification by female butterflies may be mediated by gustatory (GRs) and olfactory receptors (ORs). Here we employed RNA-Seq, bioinformatics and RT-qPCR techniques to identify sex- and tissue-specific gene expression. We focused on chemosensation related genes and pathways, including putative ORs, GRs, ionotropic receptors (IRs), odorant-binding proteins, chemosensory proteins, and steroid hormone mediated signaling in specific chemosensory tissues (i.e., antennae, legs and proboscis). Twelve butterflies evenly split between males and females were caught and used for RNA extraction. Tissue-specific sequencing libraries were prepared and sequenced using Illumina NovaSeq 6000 or HiSeq 3000, generating 2 billion 150-bp, paired-end reads. Many more genes and gene sets were differentially expressed between tissue types than between sexes. A total of 148 chemosensation-related genes exhibited sex- and/or tissue-biased expression. RT-qPCR of a small set of genes confirmed their differential expression between tissue types or between males and females. These findings laid a solid foundation for further investigations into the biological roles of these identified genes underlying chemosensation-mediated behaviors such as foraging and reproduction. Overall design: We first generated a large RNA-Seq dataset for antennae, leg and proboscis tissues of both male and female monarch butterflies, then applied various bioinformatic tools to infer differentially expressed chemosensory receptor (CR) genes and pathways relevant to chemosensation between either tissue types or sexes, and finally validated the differential expression of select CR genes