Romain Libbrecht

Interplay between insulin signaling, juvenile hormone, and vitellogenin regulates maternal effects on polyphenism in ants

Polyphenism is the phenomenon in which alternative phenotypes are produced by a single genotype in response to environmental cues. An extreme case is found in social insects, in which reproductive queens and sterile workers that greatly differ in morphology and behavior can arise from a single genotype. Experimental evidence for maternal effects on caste determination, the differential larval development toward the queen or worker caste, was recently documented in Pogonomyrmex seed harvester ants, in which only colonies with a hibernated queen produce new queens. However, the proximate mechanisms behind these intergenerational effects have remained elusive. We used a combination of artificial hibernation, hormonal treatments, gene expression analyses, hormone measurements, and vitellogenin quantification to investigate how the combined effect of environmental cues and hormonal signaling affects the process of caste determination in Pogonomyrmex rugosus. The results show that the interplay between insulin signaling, juvenile hormone, and vitellogenin regulates maternal effects on the production of alternative phenotypes and set vitellogenin as a likely key player in the intergenerational transmission of information. This study reveals how hibernation triggers the production of new queens in Pogonomyrmex ant colonies. More generally, it provides important information on maternal effects by showing how environmental cues experienced by one generation can translate into phenotypic variation in the next generation.

Robust DNA Methylation in the Clonal Raider Ant Brain

Social insects are promising model systems for epigenetics due to their immense morphological and behavioral plasticity. Reports that DNA methylation differs between the queen and worker castes in social insects [1-4] have implied a role for DNA methylation in regulating division of labor. To better understand the function of DNA methylation in social insects, we performed whole-genome bisulfite sequencing on brains of the clonal raider ant Cerapachys biroi, whose colonies alternate between reproductive (queen-like) and brood care (worker-like) phases [5]. Many cytosines were methylated in all replicates (on average 29.5% of the methylated cytosines in a given replicate), indicating that a large proportion of the C. biroi brain methylome is robust. Robust DNA methylation occurred preferentially in exonic CpGs of highly and stably expressed genes involved in core functions. Our analyses did not detect any differences in DNA methylation between the queen-like and worker-like phases, suggesting that DNA methylation is not associated with changes in reproduction and behavior in C. biroi. Finally, many cytosines were methylated in one sample only, due to either biological or experimental variation. By applying the statistical methods used in previous studies [1-4, 6] to our data, we show that such sample-specific DNA methylation may underlie the previous findings of queen- and worker-specific methylation. We argue that there is currently no evidence that genome-wide variation in DNA methylation is associated with the queen and worker castes in social insects, and we call for a more careful interpretation of the available data.

Vitellogenin Underwent Subfunctionalization to Acquire Caste and Behavioral Specific Expression in the Harvester Ant Pogonomyrmex barbatus

The reproductive ground plan hypothesis (RGPH) proposes that the physiological pathways regulating reproduction were co-opted to regulate worker division of labor. Support for this hypothesis in honeybees is provided by studies demonstrating that the reproductive potential of workers, assessed by the levels of vitellogenin (Vg), is linked to task performance. Interestingly, contrary to honeybees that have a single Vg ortholog and potentially fertile nurses, the genome of the harvester ant Pogonomyrmex barbatus harbors two Vg genes (Pb_Vg1 and Pb_Vg2) and nurses produce infertile trophic eggs. P. barbatus, thus, provides a unique model to investigate whether Vg duplication in ants was followed by subfunctionalization to acquire reproductive and non-reproductive functions and whether Vg reproductive function was co-opted to regulate behavior in sterile workers. To investigate these questions, we compared the expression patterns of P. barbatus Vg genes and analyzed the phylogenetic relationships and molecular evolution of Vg genes in ants. qRT-PCRs revealed that Pb_Vg1 is more highly expressed in queens compared to workers and in nurses compared to foragers. By contrast, the level of expression of Pb_Vg2 was higher in foragers than in nurses and queens. Phylogenetic analyses show that a first duplication of the ancestral Vg gene occurred after the divergence between the poneroid and formicoid clades and subsequent duplications occurred in the lineages leading to Solenopsis invicta, Linepithema humile and Acromyrmex echinatior. The initial duplication resulted in two Vg gene subfamilies preferentially expressed in queens and nurses (subfamily A) or in foraging workers (subfamily B). Finally, molecular evolution analyses show that the subfamily A experienced positive selection, while the subfamily B showed overall relaxation of purifying selection. Our results suggest that in P. barbatus the Vg gene underwent subfunctionalization after duplication to acquire caste- and behavior- specific expression associated with reproductive and non-reproductive functions, supporting the validity of the RGPH in ants.

Aphidius ervi Preferentially Attacks the Green Morph of the Pea Aphid, Acyrthosiphon pisum

The pea aphid, Acyrthosiphon pisum Harris (Hemiptera: Aphididae) is found in red and green color morphs. Previous work has suggested that the aphidiine parasitoid Aphidius ervi Haliday preferentially attacks green pea aphids in the field. It is not clear whether these results reflect a real preference, or some unknown clonal difference, such as in immunity, between the aphids used in the previous studies. We used three susceptibility-matched pairs of red and green morph pea aphid clones to test for preferences. In a no-choice situation, the parasitoids attacked equal proportions of each color morph. When provided with a choice, A. ervi was significantly more likely to oviposit into colonies formed from green morphs when the neighboring colony was formed from red morph aphids. In contrast, red morphs were less likely to be attacked when their neighboring colony was of the green morph. By preferentially attacking green colonies, A. ervi may reduce the likelihood of intraguild predation, as it is suggested that visually foraging predators preferentially attack red aphid colonies. Furthermore, if this host choice behavior is replicated in the field, we speculate that color morphs of the pea aphid may interact indirectly through their shared natural enemies, leading to intraspecific apparent competition.

Ant genomics sheds light on the molecular regulation of social organization.

Ants are powerful model systems for the study of cooperation and sociality. In this review, we discuss how recent advances in ant genomics have contributed to our understanding of the evolution and organization of insect societies at the molecular level.

Molecular mechanisms of phenotypic plasticity in social insects

Polyphenism in insects, whereby a single genome expresses different phenotypes in response to environmental cues, is a fascinating biological phenomenon. Social insects are especially intriguing examples of phenotypic plasticity because division of labor results in the development of extreme morphological phenotypes, such as the queen and worker castes. Although sociality evolved independently in ants, bees, wasps and termites, similar genetic pathways regulate phenotypic plasticity in these different groups of social insects. The insulin/insulin-like growth signaling (IIS) plays a key role in this process. Recent research reveals that IIS interacts with other pathways including target of rapamycin (TOR), epidermal growth factor receptor (Egfr), juvenile hormone (JH) and vitellogenin (Vg) to regulate caste differentiation.

Genetic components to caste allocation in a multiple-queen ant species.

Reproductive division of labor and the coexistence of distinct castes are hallmarks of insect societies. In social insect species with multiple queens per colony, the fitness of nestmate queens directly depends on the process of caste allocation (i.e., the relative investment in queen, sterile worker and male production). The aim of this study is to investigate the genetic components to the process of caste allocation in a multiple‐queen ant species. We conducted controlled crosses in the Argentine ant Linepithema humile and established single‐queen colonies to identify maternal and paternal family effects on the relative production of new queens, workers, and males. There were significant effects of parental genetic backgrounds on various aspects of caste allocation: the paternal lineage affected the proportion of queens and workers produced whereas the proportions of queens and males, and females and males were influenced by the interaction between parental lineages. In addition to revealing nonadditive genetic effects on female caste determination in a multiple‐queen ant species, this study reveals strong genetic compatibility effects between parental genomes on caste allocation components.

GENETIC COMPATIBILITY AFFECTS DIVISION OF LABOR IN THE ARGENTINE ANT LINEPITHEMA HUMILE

Division of labor is central to the organization of insect societies. Within‐colony comparisons between subfamilies of workers (patrilines or matrilines) revealed genetic effects on division of labor in many social insect species. Although this has been taken as evidence for additive genetic effects on division of labor, it has never been experimentally tested. To determine the relative roles of additive and nonadditive genetic effects (e.g., genetic compatibility, epistasis, and parent‐of‐origin imprinting effects) on worker behavior, we performed controlled crosses using the Argentine ant Linepithema humile. Three of the measured behaviors (the efficiency to collect pupae, the foraging propensity, and the distance between non‐brood‐tenders and brood) were affected by the maternal genetic background and the two others (the efficiency to feed larvae and the distance between brood‐tenders and brood) by the paternal genetic background. Moreover, there were significant interactions between the maternal and paternal genetic backgrounds for three of the five behaviors. These results are most consistent with parent‐of‐origin and genetic compatibility effects on division of labor. The finding of nonadditive genetic effects is in strong contrast with the current view and has important consequences for our understanding of division of labor in insect societies.

Genetics and Evolution of Social Behavior in Insects

The study of insect social behavior has offered tremendous insight into the molecular mechanisms mediating behavioral and phenotypic plasticity. Genomic applications to the study of eusocial insect species, in particular, have led to several hypotheses for the processes underlying the molecular evolution of behavior. Advances in understanding the genetic control of social organization have also been made, suggesting an important role for supergenes in the evolution of divergent behavioral phenotypes. Intensive study of social phenotypes across species has revealed that behavior and caste are controlled by an interaction between genetic and environmentally mediated effects and, further, that gene expression and regulation mediate plastic responses to environmental signals. However, several key methodological flaws that are hindering progress in the study of insect social behavior remain. After reviewing the current state of knowledge, we outline ongoing challenges in experimental design that remain to be overcome in order to advance the field.

Ant larvae regulate worker foraging behavior and ovarian activity in a dose-dependent manner.

Division of labor in insect societies relies on simple behavioral rules, whereby individual colony members respond to dynamic signals indicating the need for certain tasks to be performed. This in turn gives rise to colony-level phenotypes. However, empirical studies quantifying colony-level signal-response dynamics are lacking. Here, we make use of the unusual biology and experimental amenability of the queenless clonal raider ant Cerapachys biroi to jointly quantify the behavioral and physiological responses of workers to a social signal emitted by larvae. Using automated behavioral quantification and oocyte size measurements in colonies of different sizes and with different worker-to-larvae ratios, we show that the workers in a colony respond to larvae by increasing foraging activity and inhibiting ovarian activation in a progressive manner and that these responses are stronger in smaller colonies. This work adds to our knowledge of the processes that link plastic individual behavioral/physiological responses to colony-level phenotypes in social insect colonies.