Summary Life inside ant colonies is orchestrated with diverse pheromones, but it is not clear how ants perceive these social signals. It has been proposed that pheromone perception in ants evolved via expansions in the numbers of odorant receptors (ORs) and antennal lobe glomeruli. Here we generate the first mutant lines in the clonal raider ant, Ooceraea biroi, by disrupting orco, a gene required for the function of all ORs. We find that orco mutants exhibit severe deficiencies in social behavior and fitness, suggesting they are unable to perceive pheromones. Surprisingly, unlike in Drosophila melanogaster, orco mutant ants also lack most of the approximately 500 antennal lobe glomeruli found in wild-type ants. These results illustrate that ORs are essential for ant social organization and raise the possibility that, similar to mammals, receptor function is required for the development and/or maintenance of the highly complex olfactory processing areas in the ant brain.

A major aim of sociogenomic research is to uncover common principles in the molecular evolution of sociality. This endeavor has been hampered by the small number of specific genes currently known to function in social behavior. Here we provide several lines of evidence suggesting that ants have evolved a large and novel clade of odorant receptor (OR) genes to perceive hydrocarbonbased pheromones, arguably the most important signals in ant communication. This genomic expansion is also mirrored in the ant brain via a corresponding expansion of a specific cluster of glomeruli in the antennal lobe. We show that in the clonal raider ant, hydrocarbon-sensitive basiconic sensilla are found only on the ventral surface of the female antennal club. Correspondingly, nearly all genes in a clade of 180 ORs within the 9-exon subfamily of ORs are expressed exclusively in females and are highly enriched in expression in the ventral half of the antennal club. Furthermore, we found that across species and sexes, the number of 9-exon ORs expressed in antennae is tightly correlated with the number of glomeruli in the antennal lobe region innervated by odorant receptor neurons from basiconic sensilla. Evolutionary analyses show that this clade underwent a striking gene expansion in the ancestors of all ants and slower but continued expansion in extant ant lineages. This evidence suggests that ants have evolved a large clade of genes to support pheromone perception and that gene duplications have played an important role in the molecular evolution of ant communication.sociogenomics | chemosensation | social evolution | antennal lobe | Formicidae T he field of sociogenomics seeks to understand the molecular basis of sociality, asking how social life evolved and how it is governed at the genomic level (1). A particularly pertinent question in the field is the role of novel genes vs. conserved genes in the evolution and regulation of social behaviors, including communication (2, 3). Communication is important to a broad range of organisms, yet relatively little is known about the genetic components of communication systems and how they evolve. A particularly interesting question in the field is how social signals are perceived and what sort of genetic and physiological specializations facilitate signal perception in highly social organisms (4, 5). In the most tractable sociogenomic models, the eusocial hymenopteran insects, communication is largely chemical, and the genetics and physiology of chemosensation in these species may hold the key to understanding their advanced communication (4,6, 74).Much is already known about the insect chemosensory system in general (8,9). Briefly, chemicals are detected by porous sensory hairs (sensilla) located on chemosensory organs such as the legs, wings, palps, and especially the antennae. The sensilla house the dendrites of olfactory and/or gustatory receptor neurons (ORNs/GRNs) with receptor proteins in the olfactory receptor (OR), the gustatory receptor (GR), or the ionotropic receptor (IR) gene ...

Summary Social insects are important models for social evolution and behavior. However, in many species experimental control over important factors that regulate division of labor, such as genotype and age, is limited [1, 2]. Furthermore, most species have fixed queen and worker castes, making it difficult to establish causality between the molecular mechanisms that underlie reproductive division of labor, the hallmark of insect societies [3]. Here we present the genome of the queenless clonal raider ant Cerapachys biroi, a powerful new study system that does not suffer from these constraints. Using cytology and RAD-Seq, we show that C. biroi reproduces via automixis with central fusion and that heterozygosity is lost extremely slowly. As a consequence, nestmates are almost clonally related (r=0.996). Workers in C. biroi colonies synchronously alternate between reproduction and brood care, and young workers eclose in synchronized cohorts. We show that genes associated with division of labor in other social insects are conserved in C. biroi and dynamically regulated during the colony cycle. With unparalleled experimental control over an individual’s genotype and age, and the ability to induce reproduction and brood care [4, 5], C. biroi has great potential to illuminate the molecular regulation of division of labor.

Queens and workers of eusocial Hymenoptera are considered homologous to the reproductive and brood care phases of an ancestral subsocial life cycle. However, the molecular mechanisms underlying the evolution of reproductive division of labor remain obscure. Using a brain transcriptomics screen, we identified a single gene, (), which is always up-regulated in ant reproductives, likely because they are better nourished than their nonreproductive nestmates. In clonal raider ants (), larval signals inhibit adult reproduction by suppressing , thus producing a colony reproductive cycle reminiscent of ancestral subsociality. However, increasing ILP2 peptide levels overrides larval suppression, thereby breaking the colony cycle and inducing a stable division of labor. These findings suggest a simple model for the origin of ant eusociality via nutritionally determined reproductive asymmetries potentially amplified by larval signals.

BackgroundThe complex societies of ants and other social insects rely on sophisticated chemical communication. Two families of small soluble proteins, the odorant binding and chemosensory proteins (OBPs and CSPs), are believed to be important in insect chemosensation. To better understand the role of these proteins in ant olfaction, we examined their evolution and expression across the ants using phylogenetics and sex- and tissue-specific RNA-seq.ResultsWe find that subsets of both OBPs and CSPs are expressed in the antennae, contradicting the previous hypothesis that CSPs have replaced OBPs in ant olfaction. Both protein families have several highly conserved clades with a single ortholog in all eusocial hymenopterans, as well as clades with more dynamic evolution and many taxon-specific radiations. The dynamically evolving OBPs and CSPs have been hypothesized to function in chemical communication. Intriguingly, we find that seven members of the conserved clades are expressed specifically in the antennae of the clonal raider ant Cerapachys biroi, whereas only one dynamically evolving CSP is antenna specific. The orthologs of the conserved, antenna-specific C. biroi genes are also expressed in antennae of the ants Camponotus floridanus and Harpegnathos saltator, indicating that antenna-specific expression of these OBPs and CSPs is conserved across ants. Most members of the dynamically evolving clades in both protein families are expressed primarily in non-chemosensory tissues and thus likely do not fulfill chemosensory functions.ConclusionsOur results identify candidate OBPs and CSPs that are likely involved in conserved aspects of ant olfaction, and suggest that OBPs and CSPs may not rapidly evolve to recognize species-specific signals.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-718) contains supplementary material, which is available to authorized users.

The massive expansions of odorant receptor (OR) genes in ant genomes are notable examples of rapid genome evolution and adaptive gene duplication. However, the molecular mechanisms leading to gene family expansion remain poorly understood, partly because available ant genomes are fragmentary. Here, we present a highly contiguous, chromosome-level assembly of the clonal raider ant genome, revealing the largest known OR repertoire in an insect. While most ant ORs originate via local tandem duplication, we also observe several cases of dispersed duplication followed by tandem duplication in the most rapidly evolving OR clades. We found that areas of unusually high transposable element density (TE islands) were depauperate in ORs in the clonal raider ant, and found no evidence for retrotransposition of ORs. However, OR loci were enriched for transposons relative to the genome as a whole, potentially facilitating tandem duplication by unequal crossing over. We also found that ant OR genes are highly AT-rich compared to other genes. In contrast, in flies, OR genes are dispersed and largely isolated within the genome, and we find that fly ORs are not AT-rich. The genomic architecture and composition of ant ORs thus show convergence with the unrelated vertebrate ORs rather than the related fly ORs. This might be related to the greater gene numbers and/or potential similarities in gene regulation between ants and vertebrates as compared to flies.

Highlights d Ant odorant receptors show unique early expression and localization in pupae d Sensory neuron innervation is necessary for proper antennal lobe development d Sensory neuron ablation reveals latent antennal lobe structure in ants, but not flies d Insects with complex olfactory systems may employ a different developmental logic

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