Antennal transcriptome of Manduca sexta

Abstract

In recent years, considerable progress has been made in understanding the molecular mechanisms underlying olfaction in insects. Because of the diverse nature of the gene families involved, this process has largely relied on genomic data. As a consequence, studies have focused on a small subset of species with extensive genomic information. For Lepidoptera, a large order historically crucial to olfactory research, this circumstance has mostly limited advances to the domesticated species Bombyx mori, with some progress in the noctuid Heliothis virescens based on a nonpublic partial genome database. Because of the limited behavioral repertoire and nonexistent ecological importance of Bombyx, molecular data on the tobacco hornworm Manduca sexta are of utmost importance, especially with regards to its position as a classical olfactory model and its complex natural behavior. Here we present the use of transcriptomic and microarray data to identify members of the main olfactory gene families of Manduca. To assess the quality of our data, we correlate information on expressed receptor genes with detailed morphological data on the antennal lobe. Finally, we compare the expression of the near-complete transcript sets in male and female antennae.

Fig. 1.

GO analyses of Manduca antennal transcriptome data. GO analysis of Manduca sequences corresponding to 7,131 contigs, as predicted for their involvement in molecular functions (A) and biological processes (B), is shown. Data are presented as level 3 GO categorization for molecular function and level 2 GO categorization for biological process. Classified gene objects are depicted as percentages (in brackets) of the total number of gene objects with GO assignments.

Fig. 2.

Aligned putative OR protein sequences of Manduca (red), Bombyx (blue), and Heliothis (green). The identified receptor candidates of Manduca are spread evenly inside the family. Two candidates belonging to the subgroup of male pheromone receptors (orange) have been identified. Additionally, female-specific ORs of Bombyx have been marked (blue). Two female-specific receptor groups also include receptor candidates of Manduca. Additional instances with clear homologs in all three species have been indicated by name. Coreceptors (MsexOR-2, BmorOR-2, and HR2) have been excluded.

Fig. 3.

Example of the right hemisphere of the Manduca brain with focus on the AL and its innervation by antennal nerve fibers (AN). (A) Confocal micrographs extracting one single optical orthogonal slice from the 3D dataset of one male (A1) and one female AL (A2). (B) 3D reconstructions of the AL of both sexes depicting a ventral view of reconstructed glomeruli (B1: males, n = 68; B2: females, n = 70 ± 1) corresponding to optical slices in A. ALs display typical moth glomerular architecture: the striking sex-specific glomeruli, the male-specific MGC, and the medial large (mLFG) and lateral large female glomeruli (lLFG) are situated at the entrance of the antennal nerve and are illustrated in red. Brain outlines of adjacent neuropil areas serve as orientation guidelines. CB, central body; PC, protocerebrum; OL, optic lobe; AN, antennal nerve; glom, glomerulus. (Scale bars: 100 μm.)