Supergenes on steroids.

Article Figures and data Abstract Introduction Results and discussion Materials and methods Data availability References Decision letter Author response Article and author information Metrics Abstract Oxytocin-like peptides have been implicated in the regulation of a wide range of social behaviors across taxa. On the other hand, the social environment, which is composed of conspecifics that may vary in their genotypes, also influences social behavior, creating the possibility for indirect genetic effects. Here, we used a zebrafish oxytocin receptor knockout line to investigate how the genotypic composition of the social environment (Gs) interacts with the oxytocin genotype of the focal individual (Gi) in the regulation of its social behavior. For this purpose, we have raised wild-type or knock-out zebrafish in either wild-type or knock-out shoals and tested different components of social behavior in adults. GixGs effects were detected in some behaviors, highlighting the need to control for GixGs effects when interpreting results of experiments using genetically modified animals, since the genotypic composition of the social environment can either rescue or promote phenotypes associated with specific genes. Introduction Social genetic effects (aka indirect genetic effects) occur when the phenotype of an organism is influenced by the genotypes of conspecifics. Previous work has highlighted the major potential evolutionary consequences of social genetic effects (Moore et al., 1997; Wolf et al., 1998), with evidence for such effects to be present both in interactions between related (e.g. mothers and offspring Champagne and Meaney, 2006; Wilson et al., 2004) and unrelated individuals (e.g. sexual displays (Petfield et al., 2005), aggression Wilson et al., 2011; Sartori and Mantovani, 2013; Santostefano et al., 2017). More recently, the importance of social genetic effects for health and disease has also been recognized (Baud et al., 2017), which may explain the pervasiveness of the social environment as a mortality risk in humans (Holt-Lunstad et al., 2010; Holt-Lunstad et al., 2015). Interestingly, the potential consequences of social genetic effects for the interpretation of research results using genetically modified organisms (GMO) has been greatly neglected. GMOs have been widely used in behavioral neuroscience to investigate the causal role of candidate genes and behavioral phenotypes. Typically Knock-in and Knock-out transgenics and mutants have been used to causally link the gain or loss of behavioral function to a specific gene (Huang and Zeng, 2013). In recent years, the development of genome editing techniques, such as CRISPR-Cas9-and TALEN-induced mutations, have increased the interest in this approach and opened the door to studying the genetic basis of behavior in non-model organisms (Hsu et al., 2014). However, most studies using GMO in behavioral neuroscience have ignored the potential contribution of the genotypic composition of the social environment to the behavioral phenotype studied. This is because it has been assumed that if the genetic background of these mutants is identical and their environment has been kept constant, any phenotypic differences must come from the genetic manipulation. However, when GMOs are incrossed or visually screened at a very young age (e.g. using reporter genes, GFP) and thereafter raised and housed together until used in experiments, changes in their behavior might be affected by the divergent genotypic composition of social environments experienced by these mutants. In other words, modified behavior might be a result of growing with their peer mutants, rather than the canonical social environment provided by wild-type conspecifics. Such problem is particularly relevant when studying social behavior. Thus, given the rising interest in the study of social behavior in model organisms from worms to higher vertebrates, an assessment of the potential effect of the interaction between the genotype of the individual (Gi) and the genotypic composition of its social environment (Gs), on the behavioral phenotype of interest in GMOs used in social neuroscience is crucial. Despite the wide variety of species-specific social behaviors, a wealth of evidence has implicated the paralog nonapeptides vasopressin (VP) and oxytocin (OXT) and their receptors in the regulation of different aspects of social behavior across vertebrates (Donaldson and Young, 2008; Goodson and Thompson, 2010), suggesting a genetic toolkit role (sensu evo-devo, i.e. ancient genes highly conserved among taxa that control the same biological process) for these nonapeptides in social behavior. Nonapeptides are an ancestral neuropeptide family found both in vertebrates and invertebrates, that derived from a VP-like peptide, and that evolved along two parallel clades of VP- and OXT-like peptides from the duplication of the VP gene in early jawed fish (ca. 500 Mya). Both peptides have been implicated in the regulation of behavior and physiology across different taxa, with VP being more involved in aggression and agonistic behaviors and OXT-like peptides consistently acting in affiliative behaviors and species-specific social behaviors across diverse taxa (i.e. sexual behavior, social interactions) (Stoop, 2012; Goodson, 2013). Despite this wealth of evidence on the direct genetic effects of OXT on social behavior, social genetic effects (i.e. GixGs effects) of OXT genotypes have never been studied. In this study, we aimed to provide a proof of principle for GixGs effects in behavioral phenotypes observed in GMO by assessing the occurrence of such effects in a knockout line for the OXT receptor in zebrafish, a commonly used model species in behavioral neuroscience (Orger and de Polavieja, 2017), which forms social groups (aka shoals, Miller and Gerlai, 2007; Miller and Gerlai, 2012) and expresses a rich repertoire of social behavior (Zebrafish Neuroscience Research Consortium et al., 2013; Nunes et al., 2017). For this purpose, we studied the GixGs interaction in the effects of the OXT gene (oxtr) in different aspects of social behavior, by raising individual zebrafish of the WT (oxtr(+/+)) or knock-out genotype (oxtr(-/-)) in different social environments (i.e. oxtr(+/+) shoal or oxtr(-/-) shoal; Figure 1A). Since sociality encompasses motivational, cognitive and collective behavioral traits, we have selected a set of tests that aim to characterize these different aspects at a fundamental level: (Moore et al., 1997) the social preference and social habituation tests assess the motivation to approach conspecifics, and how it varies with the repeated access to conspecifics; (Wolf et al., 1998) the social recognition test, which provides an insight into the ability of zebrafish to discriminate between conspecifics based on one-trial learning; and (Champagne and Meaney, 2006) tests of shoaling behavior that assess how well the focal individual is able to integrate itself into an unfamiliar shoal and what influence it has on the behavior of the other shoal members. Figure 1 Download asset Open asset Genetic variation in the social environment affects zebrafish social behavior. The contribution of the individual genotype (Gi), the genotype of conspecifics in the social group (Gs) and the interaction between the two (GixGs) to the expression of behavioral phenotypes in zebrafish was assessed by raising oxytocin receptor mutant fish and wild types (focal fish marked with *) in shoals of either mutants or wild types (A). Social preference, measured by the time fish spend near a shoal vs. empty in a choice test (B, upper panel), showed a marginally significant effect of Gs (C; Source data file Figure 1—source data 1). Social habituation, which consisted on a consecutive social preference test exhibited a GixGs effect (D; Source data file Figure 1—source data 2). Social recognition, measured as the discrimination between a novel and a familiar conspecific (E, upper panel), shows a pure G effect (F; Source data file Figure 1—source data 3). Social integration, measured as distance to the centroid of the shoal (G), showed a GixGs effect (H; Source data file Figure 1—source data 4). Social influence, measured by the cohesion of the remaining shoal members (I), also showed a marginally significant GixGs effect (J; Source data file Figure 1—source data 5). Heatmaps show the spatial distribution of a representative oxtr(+/+) individual fish raised in a oxtr(+/+) group, during the entire trial, for both social preference (B, lower panel) and social recognition (E, lower panel). Data is presented as mean ± standard error of the mean (SEM). Sample sizes are nine for heterogeneous groups (i.e. focal individual with different genotype from the remaining individuals in the shoal; mutant focal in WT shoals and WT focal in mutant shoals) and 15 for homogeneous groups (i.e. focal individual with the same genotype of the remaining individuals in the shoal; mutant focal in mutant shoals and WT focal in WT shoals). Different letters indicate significant differences (p<0.05) between treatments as assessed by Tukey post-hoc tests following a two-way ANOVA (D,H,J; see Table 1). An asterisk indicates a Gi main effect in F. Figure 1—source data 1 Effects of individual and conspecifics genotype on Social Preference. https://cdn.elifesciences.org/articles/56973/elife-56973-fig1-data1-v2.xlsx Download elife-56973-fig1-data1-v2.xlsx Figure 1—source data 2 Effects of individual and conspecifics genotype on social habituation. https://cdn.elifesciences.org/articles/56973/elife-56973-fig1-data2-v2.xlsx Download elife-56973-fig1-data2-v2.xlsx Figure 1—source data 3 Effects of individual and conspecifics genotype on social recognition. https://cdn.elifesciences.org/articles/56973/elife-56973-fig1-data3-v2.xlsx Download elife-56973-fig1-data3-v2.xlsx Figure 1—source data 4 Effects of individual and conspecifics genotype on social integration. https://cdn.elifesciences.org/articles/56973/elife-56973-fig1-data4-v2.xlsx Download elife-56973-fig1-data4-v2.xlsx Figure 1—source data 5 Effects of individual and conspecifics genotype on social influence. https://cdn.elifesciences.org/articles/56973/elife-56973-fig1-data5-v2.xlsx Download elife-56973-fig1-data5-v2.xlsx Results and discussion Adult zebrafish, like many other social animals, express a tendency to approach and interact with conspecifics (social preference, Figure 1B; Engeszer et al., 2004). Here, we show that there was no significant effect of either genotype or GixGs interaction on social preference, but there was a marginally significant main effect of Gs (Table 1; Figure 1C). When fish were presented for a second time to a shoal to measure social habituation (i.e. expected reduction in social preference), we found a GixGs interaction, where oxtr(-/-) individuals raised in oxtr(-/-) shoals express enhanced social habituation (F1,44 = 5.642, p=0.022; Figure 1D). Thus, social motivation in zebrafish seems to be influenced by the genotype of conspecifics rather than by the genotype of the individual. Hence, the increased social habituation in oxtr(-/-) fish does not seem to be due to reduced social motivation, but rather to an heightened habituation to the stimuli, suggesting that the observed GixGs interaction effect is related to changes in single-stimulus learning mechanisms in mutant fish rather than to changes in social motivation. Table 1 Effect of genotype of the focal individual (Gi), genotype of conspecifics present in its social environment (Gs) and the interaction between the two (GixGs) on zebrafish social behavior was assessed using a two-way ANOVA. ~ indicates marginally significant, *p<0.05, **p<0.01, ***p<0.001. (Source data files Figure 1—source datas 1–5). Social preferenced.f.Mean squaresFSignificancePartial η2Gi10.0231.7310.1950.038Gs10.0503.7880.058~0.079Gi x Gs10.0010.0490.8250.001Error440.013Habituationd.f.Mean squaresFSignificancePartial η2Gi10.05813.9270.001 **0.240Gs10.0081.9360.1710.042Gi x Gs10.0245.6420.022 *0.114Error440.004Social recognitiond.f.Mean squaresFSignificancePartial η2Gi10.2137.6000.008 **0.147Gs10.0050.1890.6660.004Gi x Gs10.0010.0410.8410.001Error440.028Social group integrationd.f.Mean squaresFSignificancePartial η2Gi139.48624.370<0.001 ***0.356Gs112.5657.7550.008 **0.150Gi x Gs112.8117.9070.007 **0.152Error441.620Social group dispersiond.f.Mean squaresFSignificancePartial η2Gi1174.3664.3090.044 *0.089Gs1657.22116.240<0.001 ***0.270Gi x Gs1122.9803.0390.0880.065Error4440.469 When we tested social recognition, which is a form of social memory needed for individuality in social interactions (i.e. differential expression of social behavior depending on identity of interacting individual), that is known to be modulated by oxytocin both in mammals and zebrafish (Ferguson et al., 2000; Ribeiro et al., 2020), we observed that oxtr(-/-) individuals exhibit a deficit in acquisition and retention of social recognition irrespective of the social environment (oxtr(-/-) or oxtr(+/+)) in which they were raised (F1,44 = 7.600, p=0.008; Figure 1F). Thus, in contrast to social motivation, social memory seems to rely on the individual's genotype. This result is in accordance with a recent study from our lab (Ribeiro et al., 2020) that has shown a deficit in one-trial recognition memory of both conspecifics and objects in oxt(-/-) fish, suggesting that this deficit is not specific to the social domain but is rather a general domain cognitive deficit. Given that social behavior of zebrafish mainly occurs in the context of shoaling we have also investigated two shoaling behavior parameters: social integration and social influence. Social integration assesses how well the focal individual integrates in the social group (aka shoal), and is measured by its average distance to the centroid of the shoal (Figure 1G,H). A GixGs interaction was found for social integration, where oxtr(-/-) individuals raised in oxtr(-/-) shoals exhibit a significantly lower social integration than oxtr(-/-) individuals raised in oxtr(+/+) shoals; in contrast, oxtr(+/+) individuals exhibit high levels of social integration irrespective of the shoal type in which they were raised (Table 1; Figure 1H). Social influence assesses how the focal individual affects the shoaling behavior of the remaining shoal members, by measuring the shoal dispersion as defined by the perimeter of the other shoal members (Figure 1I,J). The presence of a single WT (oxtr(+/+)) individual in a oxtr(-/-) shoal was enough to increase its dispersion, whereas the presence of a single oxtr(-/-) individual in a oxtr(+/+) shoal did not affect its dispersion (Table 1; Figure 1J). In summary, we show that distinct components of social behavior are differentially affected by the genetic composition of the social environment versus the oxtr genotype of the focal individual. Social preference shows a marginally significant influence of the genotype of conspecifics. Social recognition exhibited a pure effect of the individual genotype. And clear GixGs interactions were observed in the cases of social habituation and social integration. Social influence had a major contribution of the social environment, which is also the case, to a lesser extent, with social preference. Thus, we demonstrated that genetic variation in the social environment interacts with individual genotype during the developmental acquisition of social behavior. In other words, variation in the genotypes present in the social environment can revert particular phenotypes associated with specific genes. These results are in line with reported interactions between other aspects of the social environment and oxytocin receptor genotype in the determination of social behavior phenotypes in human populations (Thompson et al., 2011; Wade et al., 2015; McQuaid et al., 2013). Our results suggest that more caution is needed in the interpretation of studies using transgenic or mutant individuals that are raised in cohorts of the same genotype, and that some phenotypes observed in transgenic or mutant lines may in fact result from GixGs interactions. Materials and methods Key resources table Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional informationGenetic reagent, TL (Danio rerio)oxtr mutant lineNunes et al., 2020ZDB-ALT-190830–1Commercial assay or kitNucleoSpin TissueMACHEREY-NAGEL# 740952.50For oxtr mutant genotypingSequence-based reagentsense 5'-TGCGCGAGGAAAACTAGTT-3'SigmaFor oxtr mutant genotypingSequence-based reagentantisense 5'-AGCAGACACTCAGAATGGTCA-3'SigmaFor oxtr mutant genotypingSoftware, , algorithmSPSS 25.0SPSSRRID:SCR_002865Software, , algorithmImagej (Fiji)Schindelin et al., 2012RRID:SCR_003070Software, , algorithmEthovision XT 11.5Noldus Technologywww.noldus.com/ethovisionSoftware, , algorithmGraphPad Prism version 6.0 cGraphPad software, San Diego, California, USAwww.graphpad.comOtherB and W mini surveillance cameraHenelec 300BAcquisition rate of 30 fpsOtherWebcamerasLogitech HD C525Acquisition rate of 30 fps Zebrafish lines and maintenance Request a detailed protocol Zebrafish were raised and bred according to standard protocols and all experimental procedures were approved by the host institution, Instituto Gulbenkian de Ciência, and by the National Veterinary Authority (DGAV, Portugal; permit number 0421/000/000/2013). OXTR mutant zebrafish line (ZFIN ID: ZDB-ALT-190830–1) was generated and provided by Dr. Gil Levkowitz (Weizmann Institute of Science) using a TALEN-based genome editing system. The characterization of this line has been described in Nunes et al., 2020. All the experimental groups were formed at 4 days post-fertilization, based on the genotype of the progenitors, before they imprint for olfactory and visual kin recognition (Gerlach et al., 2008; Hinz et al., 2013). To evaluate genotype-environment effects, fish were raised in groups according to the experimental design in Figure 1A and both female and males tested in adulthood (3 months old). Sample sizes varied between nine for heterogeneous groups (i.e. focal individual with different genotype from the remaining individuals in the shoal) and 15 for homogeneous groups (i.e. focal individual with the same genotype of the remaining individuals in the shoal). The smaller sample size of heterogeneous groups is due to the need of genotyping all individuals in these groups to single out the focal individual. Genotyping Request a detailed protocol At 3 months old, 1-week before the behavioral screenings, genomic DNA was extracted from adult fin clips using the HotSHOT protocol (Meeker et al., 2007). All group members were fin clipped at different fin locations, to allow their identification while being maintained together. The genomic region of interest was amplified by PCR and sequenced to identify the focal fish in each group. The following primers were used: sense 5'-TGCGCGAGGAAAACTAGTT-3', antisense 5'-AGCAGACACTCAGAATGGTCA-3'. Behavioral assays Video acquisition Request a detailed protocol Fish were in a tank placed on top of an infrared lightbox and video-recorded either from above (shoal preference and social recognition tests) or laterally (group behaviour tests). Video acquisition was done with software Pinnacle Studio 14 (Corel Corporation, Ottawa, Canada). Shoal preference, social habituation and social recognition analyses were performed with EthoVision video tracking system (Noldus Information Technologies, Wageningen, The Netherlands) and group behavior analyses were done with the open source FIJI image-processing package (Schindelin et al., 2012). Social preference and social habituation Request a detailed protocol The social preference test assesses the individual's sociability by observing the interactions between conspecifics (Ribeiro et al., 2020): a focal fish was placed in a central compartment (30 × 15×10 cm) of a three-compartment tank, separated by transparent and sealed partitions. A shoal of unfamiliar fish was placed in one of the lateral compartments (15 × 10×10 cm), while the other contained only water. To avoid any side bias, the stimuli were balanced across trials. After an acclimatization period (10 min), the focal fish was released from a start box and allowed to explore the tank, while its behavior was video-recorded for 10 min. The time spent by the focal fish near (less than two body lengths) each compartment was quantified and used to calculate the social preference score (SP = Time near shoal/ [Time near shoal + Time near empty]). A score above 0.5 indicates a preference for the shoal. The social preference test was performed twice, with 24 hr in between, and social preference scores of both tests were used to calculate the habituation index (Hab. Score = 1- [SPTrial2]/[SPTrial1 + SPTrial2]). A score above 0.5 represents a decrease in preference to associate with conspecifics. Social recognition Request a detailed protocol The social recognition assay to evaluate short-term (i.e. 10 min retention) social memory was adapted from the procedure already developed in our lab for long-term (i.e. 24 hr retention) social memory in zebrafish (Gerlach et al., 2008), and has already been used successfully in previous studies (Ribeiro et al., 2020; Madeira and Oliveira, 2017). A focal fish was placed for 10 min in the central compartment of a three-compartment tank, separated by transparent and sealed partitions, to acclimatize. The focal fish was allowed to interact visually across partitions with two novel (unfamiliar) conspecifics for 10 min. After, both stimuli were removed, one was placed in the same compartment (familiar conspecific stimulus), while a novel conspecific was placed in the other compartment (novel conspecific stimulus). In a second 10 min interaction, the time spent by the focal fish near each compartment (termed novel cue or familiar cue) was quantified and used to measure the preference for the novel (Recognition Score = Time near Novel/[Time near Novel + Time near Familiar]). A recognition score of 0.5 indicates no preference between novel or familiar conspecifics. Shoaling behavior Request a detailed protocol Shoaling behavior is a common behavior present in fish models and allows to determine complex interactions between individuals. Both focal fish and social partners were recorded in the home tanks (3.5L tank). Focal fish were tagged with fin clips for easy identification. The behaviors were video-recorded from side view for 10 min. Two components of shoaling behavior were analyzed manually in time bins of 8 s, using FIJI software (Schindelin et al., 2012): (Moore et al., 1997) focal fish distance to the group centroid (social integration); and (Wolf et al., 1998) the dispersion of the remaining shoal members as measured by their perimeter (social influence). Data analysis Request a detailed protocol Data were analysed using SPSS 25.0. All data sets were tested for departures from normality with Shapiro-Wilks test. Two factor univariate ANOVA were used for comparing multiple groups. All data sets were corrected for multiple comparisons. Tukey's Test comparisons were used as post-hocs. Given that ANOVA is known to be underpowered for detecting significance of genotype x environment interaction (Wahlsten, 1990) we have decided to proceed with post-hoc tests for multiple comparisons among treatments even when GixGs interaction were only marginally significant (p<0.10). Graphs were performed with GraphPad software. Ethical approval Request a detailed protocol All experiments were performed in accordance with the relevant guidelines and regulations for the care and use of animals in research and approved by the competent Portuguese authority (Direcção Geral de Alimentação e Veterinária, permit 0421/000/000/2017). Add a comment + Open annotations. The current annotation count on this page is being calculated. Data availability Data used in this study is provided as supplemental material at this stage. Data available on Dryad at The following data sets were generated Dryad Genetic variation in the social environment affects behavioral phenotypes of oxytocin receptor mutants. References A A Genetic variation in the social environment to health and disease Champagne during care and the development of the offspring in a model and the of sociality Engeszer social preference in zebrafish Social in the oxytocin gene G A recognition in a for olfactory of the Goodson social and relevant Goodson mechanisms of social behavior and species-specific social in Hinz A G recognition in zebrafish, is based on on olfactory and visual stimuli Holt-Lunstad Social and mortality a Holt-Lunstad and social as risk for mortality on and of for genome Genetic to in the of Neuroscience Madeira social recognition memory in zebrafish Zebrafish McQuaid A of an oxytocin receptor gene and to in Neuroscience for of genomic DNA from zebrafish Miller of shoaling behaviour in zebrafish (Danio Research Miller to of collective in zebrafish (Danio Wolf phenotypes and the evolutionary direct and indirect genetic effects of social interactions Nunes Social phenotypes in zebrafish The and of Behavioral of Nunes Levkowitz G mechanisms of social in zebrafish

Article Figures and data Abstract Editor's evaluation Introduction Results Discussion Materials and methods Data availability References Decision letter Author response Article and author information Metrics Abstract In white-throated sparrows, two alternative morphs differing in plumage and behavior segregate with a large chromosomal rearrangement. As with sex chromosomes such as the mammalian Y, the rearranged version of chromosome two (ZAL2m) is in a near-constant state of heterozygosity, offering opportunities to investigate both degenerative and selective processes during the early evolutionary stages of ‘supergenes.’ Here, we generated, synthesized, and analyzed extensive genome-scale data to better understand the forces shaping the evolution of the ZAL2 and ZAL2m chromosomes in this species. We found that features of ZAL2m are consistent with substantially reduced recombination and low levels of degeneration. We also found evidence that selective sweeps took place both on ZAL2m and its standard counterpart, ZAL2, after the rearrangement event. Signatures of positive selection were associated with allelic bias in gene expression, suggesting that antagonistic selection has operated on gene regulation. Finally, we discovered a region exhibiting long-range haplotypes inside the rearrangement on ZAL2m. These haplotypes appear to have been maintained by balancing selection, retaining genetic diversity within the supergene. Together, our analyses illuminate mechanisms contributing to the evolution of a young chromosomal polymorphism, revealing complex selective processes acting concurrently with genetic degeneration to drive the evolution of supergenes. Editor's evaluation In this important paper, the authors generate and analyze new genome and gene expression data to understand better the evolution of the white-throated sparrow supergene region, which contains 1000 genes and determines whether a bird has a tan or a white stripe. The study convincingly illustrates how the cessation of recombination that results from a chromosomal inversion can become a source of evolutionary novelty. The lack of recombination can result in the accumulation of deleterious variation leading to degeneration, but it can also (as here) facilitate genomic diversification and adaptation. The results will be of interest to a broad array of researchers studying genome architecture and phenotypic diversity and evolution. https://doi.org/10.7554/eLife.79387.sa0 Decision letter Reviews on Sciety eLife's review process Introduction Supergenes comprise closely linked genetic variants that are maintained due to suppressed recombination (Charlesworth, 2016; Thompson and Jiggins, 2014). Their evolution presents an interesting paradox, in that the suppression of recombination that occurs inside supergenes reduces the efficacy of natural selection, leading to genetic degeneration. At the same time, supergenes are associated with dramatically divergent, adaptive phenotypes. These divergent phenotypes, which include classic examples of Batesian mimicry and self-incompatibility in flowering plants (Charlesworth, 2016; Thompson and Jiggins, 2014; Otto and Lenormand, 2002) and striking polymorphisms in social behavior (Wang et al., 2013; Huang et al., 2018; Yan et al., 2020; Martinez-Ruiz et al., 2020; Farrell et al., 2013; Küpper et al., 2016; Lamichhaney et al., 2016), have long inspired both theoretical and empirical studies of their evolution. Recent genome-scale studies have illuminated wide-ranging impacts of supergene evolution on complex phenotypes across diverse taxa (e.g. Schwander et al., 2014; Pearse et al., 2019; Hager et al., 2022; Joron et al., 2011; Kunte et al., 2014; Kess et al., 2019; Lundberg et al., 2017; Roberts et al., 2009; Sanchez-Donoso et al., 2022; Funk et al., 2021). Currently, the mechanisms by which functionally divergent supergene haplotypes evolve in the face of multiple evolutionary forces remain poorly understood, presenting a critical gap in knowledge. One notable example of a supergene associated with social behavior is found in white-throated sparrows (Zonotrichia albicollis), in which a large supergene co-segregates with parental behavior and aggression (Tuttle, 2003; Maney et al., 2015; Horton et al., 2013; Tuttle et al., 2016; Sun et al., 2018; Merritt et al., 2020). White-throated sparrows occur in two alternative plumage morphs, white- and tan-striped (Lowther, 1961). These morphs differ not only in their plumage coloration, but also in their social behavior, with white-striped birds exhibiting increased aggression and more frequent extra-pair copulations, and tan-striped birds engaging in more parental care compared with birds of the white-striped morph (Tuttle, 2003; Maney et al., 2015; Maney, 2008; Horton et al., 2012). These alternative morphs are linked to a large (~100 Mbp, >1 k genes) rearrangement on the second largest chromosome, called ZAL2m, so named because the rearranged chromosome is metacentric. White-striped birds are heterozygous for ZAL2m and the sub-metacentric chromosomal arrangement, ZAL2, whereas tan-striped birds are homozygous for ZAL2 (30, 31). In addition to highly divergent social behavior, this relatively young supergene (estimated to have arisen 2–3 million years ago Tuttle et al., 2016; Thomas et al., 2008; Huynh et al., 2010), is also associated with a remarkable disassortative mating system that maintains the ‘balanced’ morph frequencies in the population. Almost all breeding pairs consist of one bird of each morph, earning the species the moniker ‘the bird with four sexes’ (Campagna, 2016). Breeding pairs consisting of two individuals of the same morph are estimated to occur less than 1% of the time (Tuttle et al., 2016; Thorneycroft, 1966) and only six ZAL2m homozygotes (i.e. ‘super-white’ birds) have ever been identified (Horton et al., 2013; Tuttle et al., 2016; Thorneycroft, 1975; Falls and Kopachena, 2020) out of thousands of birds karyotyped or genotyped. Given that ZAL2m exists in a near-constant state of heterozygosity, it is in a state of suppressed recombination, similar to the Y and W sex chromosomes in mammals and birds, respectively. The suppression of recombination on ZAL2m is expected to reduce the efficacy of natural selection, leading to reduced genetic diversity and the degeneration of the chromosome (Barton and Charlesworth, 1998; Charlesworth, 2012). On the other hand, the tight linkage of alleles within the ZAL2m supergene may contribute to adaptive phenotypes (Tuttle et al., 2016; Sun et al., 2018; Maney et al., 2020). Therefore, this system provides a unique opportunity to investigate the evolution of a supergene underlying social and mating behavior (Tuttle et al., 2016; Sun et al., 2018; Merritt et al., 2020; Sun et al., 2021). Here we aim to better understand the evolutionary forces shaping the ZAL2 and ZAL2m chromosomes. Our goal was to address two unanswered questions. First, to what extent has ZAL2m degenerated? Early analyses of the rearrangement (Davis et al., 2011) did not show signals of degeneration, such as pseudogenization or the accumulation of repetitive sequences. However, Tuttle et al., 2016 found a weak signal of excess non-synonymous polymorphism for genes inside the rearranged region on ZAL2m and reduced allelic expression for ZAL2m genes, which could be consistent with functional degradation of ZAL2m (Tuttle et al., 2016). (Sun et al., 2018) similarly found a slightly higher number of non-synonymous substitutions and an increased ratio of non-synonymous to synonymous substitution rates (dN/dS) on ZAL2m compared with ZAL2. (Sun et al., 2018) also found reduced expression of ZAL2m alleles in brain tissue, perhaps suggesting that the accumulation of deleterious mutations has led to reduced expression of genes from ZAL2m. Their additional finding of reduced accumulation of mutations in functional regions suggested that ZAL2m has, in fact, experienced purifying selection to remove deleterious alleles. Thus, while there is some evidence that ZAL2m has degenerated, these results have been inconsistent and somewhat inconclusive. Second, what are the selective forces shaping the genomic landscapes of both ZAL2 and the ZAL2m supergene? The signals of both purifying and positive selection have been relatively weak in previous genomic analyses of ZAL2 and ZAL2m (Tuttle et al., 2016; Sun et al., 2018). Yet, by definition, ZAL2m must contain variation that underlies the differences between the white- and tan-striped morphs (Fisher, 1931; Bull, 1983; Charlesworth and Charlesworth, 1980; Rice, 1987b; Rice, 1987a). There is already some evidence that this variation affects behavior; allelic differences in the promoter region of the gene encoding estrogen-receptor alpha (ESR1) are likely to alter expression (Merritt et al., 2020), and the expression of this gene was shown to be necessary for aggressive behavior typical of the white-striped morph (Merritt et al., 2020). ZAL2m is also associated with differential expression of a key neuromodulator, vasoactive intestinal peptide (Horton et al., 2020), known to be causal for aggression in songbirds (Goodson et al., 2012). Investigations of young heteromorphic sex chromosomes suggest that the accumulation of sexually antagonistic genes (i.e. genes that are beneficial to one sex and harmful to the other) may in fact drive the evolution of sex chromosomes (Bachtrog, 2004; Bachtrog, 2006; Zhou and Bachtrog, 2012). For example, positive selection at a small number of antagonistic alleles was shown to be a potent force shaping evolution of the young Y chromosomes in Drosophila miranda (Bachtrog, 2004) even in the face of degeneration of other genes elsewhere on the chromosome. In white-throated sparrows, evidence of positive selection on both ZAL2 and ZAL2m has been quite limited (Tuttle et al., 2016; Sun et al., 2018). Nonetheless, the discovery of ZAL2- and ZAL2m-specific alleles that benefit the tan- and white-striped morphs, respectively (Merritt et al., 2020; Horton et al., 2020), suggests that antagonistic selection likewise contributes to the evolution of both ZAL2 and ZAL2m. In addition to antagonistic selection, balancing selection may be implicated in the evolution of ZAL2m. The negative assortative mating system in white-throated sparrows, which maintains the chromosomal polymorphism, is a canonical example of balancing selection (Huynh et al., 2010). However, balancing selection is also a way of maintaining advantageous genetic diversity in populations, which may be especially critical in the context of a non-recombining chromosome. Indeed, balancing selection appears to be more common in self-fertilizing (selfing) vs non-selfing species, which are likewise characterized by reduced genetic diversity, increased linkage disequilibrium, and reduced efficacy of selection (Glémin, 2021; Glémin et al., 2019; Delph and Kelly, 2014; Gaut et al., 2015). Therefore, balancing selection may maintain multiple alleles inside non-recombining regions of chromosomes like ZAL2m. Previous studies have been limited in the extent to which they could test directly for degeneration, adaptive changes on ZAL2m, and selection at the genome level. These limitations stemmed from low sample sizes of sequencing data, the reduced intraspecies variability, and a low-quality ZAL2m assembly that prevented detection of long-range haplotypes (Tuttle et al., 2016; Sun et al., 2018; Thomas et al., 2008). Here, we overcome these challenges by analyzing extensive genomic, transcriptomic, and population data, providing insight into the evolution of young supergenes. Results Novel and extensive genomic and population data from white-throated sparrows To better understand the evolutionary history of the ZAL2m chromosomal rearrangement, we generated additional sequence data from a rare, ‘super-white’ (ZAL2m homozygote) bird (Horton et al., 2013; Sun et al., 2018). We generated variable fragment size libraries consisting of 150 bp paired-end reads (insert size of 300 bp and 500 bp) and 125 bp mate pair reads (insert size of 1 kb, 4–7 kb, 7–10 kb, and 10–15 kb). We performed whole-genome sequencing of an additional 62 birds (49 white-striped birds and 13 tan-striped birds sampled from a variety of locations around the U.S.) (Materials and methods, Supplementary file 1). White-striped birds, which are heterozygous for the rearrangement (ZAL2/2m), were sequenced at higher coverage than tan-striped birds (ZAL2 homozygotes) so that we could obtain sufficient reads to separate ZAL2 and ZAL2m alleles in white-striped individuals (average mean depth coverages were 41.5 × vs 28.4 × for white- and tan-striped birds, respectively, Supplementary file 2). Genomic variants were called according to the guidelines of Genome Analysis Toolkit (GATK) (ver. 4.1) (Materials and methods), leading to the discovery of a total of 11,382,994 single nucleotide polymorphisms (SNPs). None of the samples showed evidence of family relationships when we computed relatedness estimates between individuals. Consequently, we used all samples in the subsequent analyses. We found a significantly higher number of polymorphic sites within white-striped birds than tan-striped birds exclusively for ZAL2/2m chromosomal regions (Figure 1—figure supplement 1). Nucleotide diversity of the rearranged region of the ZAL2/2m chromosomes was elevated in white-striped birds compared with tan-striped birds, suggesting distinctive patterns between the two plumage morphs (Figure 1a). Figure 1 with 2 supplements see all Download asset Open asset Genomic data from newly sequenced tan- and white-striped birds. (A) Nucleotide diversity of macro-chromosomes for tan-striped (TS) and white-striped (WS) birds. White-striped birds (ZAL2/2m) show elevated nucleotide diversity for the ZAL2/2m inverted (INV, i.e. rearranged) regions (ZAL2/2m inv), while TS birds (ZAL2/2) show overall reduced nucleotide diversity for the inverted regions compared with other chromosomes. Note that panel (A) shows the comparison across morph. The comparison across the ZAL2 and ZAL2m alleles is shown in Figure 2a. (B) Scatterplots of eigenvector 1 (PC1) and eigenvector 2 (PC2) from principal component analysis of all single-nucleotide variants (left panel). (C) Principal component analysis (PCA) excluding single nucleotide polymorphisms (SNPs) on the ZAL2 chromosomes (right panel). The sex chromosomes and the ZAL3 chromosome (which includes an additional chromosomal inversion) were excluded from both PCA analyses. Note that ‘location’ here refers to the site of collection or capture of the bird: Georgia (GA), Illinois (IL), or Maine (ME). Breeding locations for GA and IL birds are unknown. Figure 1—source data 1 Nucleotide diversity between tan- and white-striped birds. Figure 1B and C: Supplementary file 1 (PCAs performed using variant call format (vcf) data from whole genome sequencing). https://cdn.elifesciences.org/articles/79387/elife-79387-fig1-data1-v1.txt Download elife-79387-fig1-data1-v1.txt Among the total SNPs identified, 12.6% (N=1,439,991) resided on scaffolds we have previously assigned to the ZAL2/2m chromosome (Sun et al., 2018). Principal component analysis (PCA) of these ZAL2/2m SNPs revealed distinct clusters corresponding to the morphs (Figure 1b). The first principal component (PC1), which explained 6.7% of the variation in the data, clearly separated tan- and white-striped birds, with the lone super-white individual (ZAL2m/2m homozygote) as a clear outlier. In contrast, other available phenotypic information, including sex and geographic origin of samples, did not show meaningful variation with the principal components, and other PCs had little explanatory power (Figure 1b). Tests for admixture also failed to identify significant population substructures by geographical origin of samples (Figure 1—figure supplement 2). This lack of population structure is unsurprising, as 35 of the 63 samples (56%) were from birds that were migrating, and, thus, the breeding location of these birds is unknown. Features of the ZAL2m chromosome consistent with reduced efficacy of natural selection and low levels of recombination We examined several genomic features of the ZAL2m chromosome using the additional genomic resources we generated. We first performed a de novo genome assembly of the super-white bird, employing newly generated sequence data, to study the ZAL2m chromosome with an assembly derived entirely from a bird homozygous for the ZAL2m chromosome. The total assembly size was 1058 Mbp (N50 length of 3.1 Mbp, longest scaffold 27 Mbps), comparable to that of the ZAL2/2 reference assembly (1052 Mbp, N50 scaffold length of 4.86 Mbp, longest scaffold 45 Mbp) (see Supplementary file 3 for more details). There were 160 putatively ZAL2m-linked scaffolds (Materials and methods), with a total length (110.99 Mbp) comparable with that of ZAL2-linked scaffolds from the reference assembly (108.5 Mbp Tuttle et al., 2016). Despite this similarity in total length, however, the average length of the individual ZAL2m-linked scaffolds was significantly shorter than scaffolds on other chromosomes in the super-white assembly (p<0.001, Mann-Whitney U-test). It was also shorter than the average scaffold length on the ZAL2 chromosome in the ZAL2/2 reference assembly (Figure 2a). We did not observe such a pattern in the other chromosomes of similar size when comparing between the two assemblies (Figure 2a). This result was consistent with the presence of repetitive DNA sequences on ZAL2m causing more assembly breaks compared with the ZAL2/2 reference genome. We found evidence that the ZAL2m chromosome contained more repeat elements and was especially enriched for long terminal repeat elements (2.4 Mbp vs 2.1 Mbp) and interspersed repeats (5.8 Mbp vs 5.5 Mbp), compared with the ZAL2 chromosome. The number of these repeat elements is likely to be underestimated, given that the ZAL2m assembly is highly fragmented. Additionally, we found that ZAL2 and ZAL2m had accumulated a higher proportion of structural variants (insertions and deletions) compared with other chromosomes (Figure 2b). Figure 2 with 1 supplement see all Download asset Open asset Genetic divergence between ZAL2 and ZAL2m chromosomes. (A) The scaffolds for the ZAL2m chromosome in the super-white (SWS) assembly tend to be fragmented compared with those for the ZAL2 chromosome in the tan-striped (TS) assembly. ** p<0.001 (Mann-Whitney U-test); ns, not significant (B) Fraction of structural variants (SV), both insertion and deletion events, for the 4 largest chromosomes, using the tan-striped assembly as a reference. The fraction of SV is computed as a total base affected by variants divided by the length of the chromosome. (C) Number of fixed mutations derived in ZAL2 and ZAL2m in protein-coding regions (D) Sliding window (window size of 20 genes with step size of 5 genes) analysis of the ratio of nonsynonymous to synonymous nucleotide diversity (πN/πS) within the ZAL2 and ZAL2m chromosomes. The ZAL2m outlier region is highlighted (colored background). (E) Site frequency spectrum of polymorphic sites. (F) Decay of linkage disequilibrium. (G) Proportion of the ZAL2m alleles expressed for each tissue set. The proportion of the ZAL2m alleles expressed is less than the null hypothesis of 0.5 for all tissues except nestling AMV using false discovery rate (FDR) correction. Hyp, hypothalamus; AMV, ventromedial arcopallium. Figure 2—source data 1 Scaffold length. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data1-v1.txt Download elife-79387-fig2-data1-v1.txt Figure 2—source data 2 Structural variant proportions. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data2-v1.txt Download elife-79387-fig2-data2-v1.txt Figure 2—source data 3 Variant information. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data3-v1.txt Download elife-79387-fig2-data3-v1.txt Figure 2—source data 4 Haplotype phased nucleotide diversity data. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data4-v1.txt Download elife-79387-fig2-data4-v1.txt Figure 2—source data 5 Minor alleles. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data5-v1.txt Download elife-79387-fig2-data5-v1.txt Figure 2—source data 6 Linkage disequilibrium. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data6-v1.txt Download elife-79387-fig2-data6-v1.txt Figure 2—source data 7 RNAseq allele specific expression data in long format. https://cdn.elifesciences.org/articles/79387/elife-79387-fig2-data7-v1.txt Download elife-79387-fig2-data7-v1.txt Figure 3 with 5 supplements see all Download asset Open asset Genetic diversity and patterns of divergence across the rearranged region of the ZAL2m chromosome and in the ZAL2m outlier region. (A) Tajima’s D and nucleotide diversity across the ZAL2 and ZAL2m chromosomes. The ZAL2m outlier region is highlighted (colored background). (B) Phylogenetic tree of randomly selected regions (left panel) and the ZAL2m outlier region (right panel). The ZAL2m chromosome shows multiple haplotype structures and has longer branch lengths within the population compared with ZAL2 chromosomes. (C) Single nucleotide polymorphism (SNP) genotype plot of a scaffold inside the ZAL2m outlier region (Scaffold NW_005189516.1, 1900001–1950001). The plot shows two haplogroups. Major allele SNPs (A, same genotype as the super-white ZAL2m/2m genome) are represented in purple, and minor allele SNPs (a, different from the super-white genome) in red. Tan that there were fixed SNPs to ZAL2 vs ZAL2m in data. (D) Genetic divergence for a of the rearrangement. between the ZAL2 chromosome and 1 is in between ZAL2 and 2 in and between and in Figure data 1 for scaffold Figure Supplementary file 1 D and nucleotide diversity from variant call format (vcf) data from whole genome sequencing). Download Figure data 2 for scaffold Download Figure data 3 data for scaffold Download Figure data 4 between ZAL2m haplotypes and ZAL2. Download using the large of newly generated population genomic data, we examined patterns of SNPs on ZAL2m alleles from those on ZAL2 haplotype using fixed differences between the two chromosome (Materials and We found that the total number of genetic variants was reduced on the ZAL2m alleles compared with ZAL2 alleles vs SNPs and vs on ZAL2m and ZAL2, respectively, after excluding The mean nucleotide diversity was similarly reduced on the ZAL2m chromosome compared with the ZAL2 chromosome vs for ZAL2m vs ZAL2, of the genetic variants on the ZAL2 and ZAL2m alleles showed evidence of only weak genetic degeneration. The ratio of non-synonymous to synonymous fixed differences inside the rearranged region was but elevated for compared with fixed differences (Figure which is consistent with positive selection or purifying selection on ZAL2m. We found that the ratio of non-synonymous to synonymous nucleotide diversity (πN/πS) was significantly increased on ZAL2m compared with ZAL2 × Mann-Whitney Figure The minor allele site frequency spectrum for the ZAL2m synonymous and non-synonymous sites showed a large proportion of variants and an of allele frequency as the minor allele (Figure also suggesting reduced efficacy of purifying selection on ZAL2m. the rates of the two chromosomes are the ratio of population size between the ZAL2m and ZAL2 can be by the ratio of nucleotide diversity of synonymous sites between the ZAL2 and ZAL2m. The proportion of between ZAL2m and ZAL2 is which is than the expected ratio of the ZAL2m chromosome is as frequent as the ZAL2, given ‘balanced’ morph frequencies in the This proportion suggests that of ZAL2m has than expected from the size in the consistent with the of reduced We that the linkage between variants on ZAL2m the classic with (Figure at some of these results are consistent with but not entirely recombination on the ZAL2m chromosome. We examined whether degeneration inside the rearranged region on ZAL2m has in reduced expression of the alleles by the ZAL2m supergene To we used multiple large RNAseq from a variety of tissues in birds sampled from different geographic locations and of (see Materials and methods, 1). As and consistent with what was previously (Sun et al., we found evidence of reduced expression of the ZAL2m alleles in of tissue (Figure We for an between the number of accumulated mutations and in the promoter on ZAL2m and allelic bias in expression of the ZAL2m alleles within each tissue, which genetic degeneration within or genes to reduced expression of ZAL2m. We found evidence that allelic bias in gene expression was associated with the rate of non-synonymous fixed differences the size was small (Figure supplement 1a). the rate of synonymous fixed differences Figure supplement the number of fixed differences within 1 of the site Figure supplement were associated with allelic Thus, the overall in expression of the alleles by the ZAL2m supergene is associated with an increased number of non-synonymous fixed nucleotide changes within The limited and weak of the however, that the pattern of gene expression may have been affected also by other for example selection as nucleotide selection at more such as differences between ZAL2 and ZAL2m in DNA or (see Sun et al., 2021). 1 of sequencing data size early in the breeding et al., 2015; Sun et al., early in the breeding from during the breeding and during in on long or to breeding vs at two time during the et al., of balancing selection on the ZAL2m chromosome the of genetic diversity was overall reduced on ZAL2m, it was elevated in one region corresponding to 5 This region, to as the ZAL2m outlier region 2 and includes at protein-coding genes that are as single genes across 13 species 2). On nucleotide diversity in ZAL2m across this region was which is higher than the mean nucleotide diversity of ZAL2m and even the nucleotide diversity in the corresponding region within ZAL2 by 2 of protein-coding genes inside the ZAL2m outlier region.

A chromosomal inversion predicts the expression of sex steroid-related genes in a species with alternative behavioral phenotypes

Despite growing interest in animal social networks, surprisingly little is known about whether individuals are consistent in their social network characteristics. Networks are rarely repeatedly sampled; yet an assumption of individual consistency in social behaviour is often made when drawing conclusions about the consequences of social processes and structure. A characterization of such social phenotypes is therefore vital to understanding the significance of social network structure for individual fitness outcomes, and for understanding the evolution and ecology of individual variation in social behaviour more broadly. Here, we measured foraging associations over three winters in a large PIT-tagged population of great tits, and used a range of social network metrics to quantify individual variation in social behaviour. We then examined repeatability in social behaviour over both short (week to week) and long (year to year) timescales, and investigated variation in repeatability across age and sex classes. Social behaviours were significantly repeatable across all timescales, with the highest repeatability observed in group size choice and unweighted degree, a measure of gregariousness. By conducting randomizations to control for the spatial and temporal distribution of individuals, we further show that differences in social phenotypes were not solely explained by within-population variation in local densities, but also reflected fine-scale variation in social decision making. Our results provide rare evidence of stable social phenotypes in a wild population of animals. Such stable social phenotypes can be targets of selection and may have important fitness consequences, both for individuals and for their social-foraging associates.

Consistent individual differences in social phenotypes have been observed in many animal species. Changes in demographics, dominance hierarchies or ecological factors, such as food availability or disease prevalence, are expected to influence decision-making processes regarding social interactions. Therefore, it should be expected that individuals show flexibility rather than stability in social behaviour over time to maximize the fitness benefits of social living. Understanding the processes that create and maintain social phenotypes requires data encompassing a range of socioecological settings and variation in intrinsic state or life-history stage or strategy. Using observational data spanning up to 19 years for some individuals, we demonstrate that multiple types of social behaviour are repeatable over the long term in wild chimpanzees, a long-lived species with complex fission–fusion societies. We controlled for temporal, ecological and demographic changes, limiting pseudo-repeatability. We conclude that chimpanzees living in natural ecological settings have relatively stable long-term social phenotypes over years that may be independent of life-history or reproductive strategies. Our results add to the growing body of the literature suggesting consistent individual differences in social tendencies are more likely the rule rather than the exception in group-living animals.

BackgroundRASopathies are a group of disorders that result from mutations in genes coding for proteins involved in regulating the Ras-MAPK signaling pathway, and have an increased incidence of autism spectrum disorder (ASD). Legius syndrome is a rare RASopathy caused by loss-of-function mutations in the SPRED1 gene. The patient phenotype is similar to, but milder than, Neurofibromatosis type 1—another RASopathy caused by loss-of-function mutations in the NF1 gene. RASopathies exhibit increased activation of Ras-MAPK signaling and commonly manifest with cognitive impairments and ASD. Here, we investigated if a Spred1-/- mouse model for Legius syndrome recapitulates ASD-like symptoms, and whether targeting the Ras-MAPK pathway has therapeutic potential in this RASopathy mouse model.MethodsWe investigated social and communicative behaviors in Spred1-/- mice and probed therapeutic mechanisms underlying the observed behavioral phenotypes by pharmacological targeting of the Ras-MAPK pathway with the MEK inhibitor PD325901.ResultsSpred1-/- mice have robust increases in social dominance in the automated tube test and reduced adult ultrasonic vocalizations during social communication. Neonatal ultrasonic vocalization was also altered, with significant differences in spectral properties. Spred1-/- mice also exhibit impaired nesting behavior. Acute MEK inhibitor treatment in adulthood with PD325901 reversed the enhanced social dominance in Spred1-/- mice to normal levels, and improved nesting behavior in adult Spred1-/- mice.LimitationsThis study used an acute treatment protocol to administer the drug. It is not known what the effects of longer-term treatment would be on behavior. Further studies titrating the lowest dose of this drug that is required to alter Spred1-/- social behavior are still required. Finally, our findings are in a homozygous mouse model, whereas patients carry heterozygous mutations. These factors should be considered before any translational conclusions are drawn.ConclusionsThese results demonstrate for the first time that social behavior phenotypes in a mouse model for RASopathies (Spred1-/-) can be acutely reversed. This highlights a key role for Ras-MAPK dysregulation in mediating social behavior phenotypes in mouse models for ASD, suggesting that proper regulation of Ras-MAPK signaling is important for social behavior.

Differences in behavioural phenotype between parental deletion and maternal uniparental disomy in Prader–Willi syndrome: an ERP study

Activity of the hypothalamic–pituitary–gonadal axis differs between behavioral phenotypes in female white-throated sparrows (Zonotrichia albicollis)

Human life history (LH) strategies are theoretically regulated by developmental exposure to environmental cues that ancestrally predicted LH-relevant world states (e.g., risk of morbidity–mortality). Recent modeling work has raised the question of whether the association of childhood family factors with adult LH variation arises via (i) direct sampling of external environmental cues during development and/or (ii) calibration of LH strategies to internal somatic condition (i.e., health), which itself reflects exposure to variably favorable environments. The present research tested between these possibilities through three online surveys involving a total of over 26,000 participants. Participants completed questionnaires assessing components of self-reported environmental harshness (i.e., socioeconomic status, family neglect, and neighborhood crime), health status, and various LH-related psychological and behavioral phenotypes (e.g., mating strategies, paranoia, and anxiety), modeled as a unidimensional latent variable. Structural equation models suggested that exposure to harsh ecologies had direct effects on latent LH strategy as well as indirect effects on latent LH strategy mediated via health status. These findings suggest that human LH strategies may be calibrated to both external and internal cues and that such calibrational effects manifest in a wide range of psychological and behavioral phenotypes.

Arid1b is a high confidence risk gene for autism spectrum disorder that encodes a subunit of a chromatin remodeling complex expressed in neuronal progenitors. Haploinsufficiency causes a broad range of social, behavioral, and intellectual disability phenotypes, including Coffin-Siris syndrome. Recent work using transgenic mouse models suggests pathology is due to deficits in proliferation, survival, and synaptic development of cortical neurons. However, there is conflicting evidence regarding the relative roles of excitatory projection neurons and inhibitory interneurons in generating abnormal cognitive and behavioral phenotypes. Here, we conditionally knocked out either one or both copies of Arid1b from excitatory projection neuron progenitors and systematically investigated the effects on intrinsic membrane properties, synaptic physiology, social behavior, and seizure susceptibility. We found that disrupting Arid1b expression in excitatory neurons alters their membrane properties, including hyperpolarizing action potential threshold; however, these changes depend on neuronal subtype. Using paired whole-cell recordings, we found increased synaptic connectivity rate between projection neurons. Furthermore, we found reduced strength of excitatory synapses to parvalbumin (PV)-expression inhibitory interneurons. These data suggest an increase in the ratio of excitation to inhibition. However, the strength of inhibitory synapses from PV interneurons to excitatory neurons was enhanced, which may rebalance this ratio. Indeed, Arid1b haploinsufficiency in projection neurons was insufficient to cause social deficits and seizure phenotypes observed in a preclinical germline haploinsufficient mouse model. Our data suggest that while excitatory projection neurons likely contribute to autistic phenotypes, pathology in these cells is not the primary cause.

Endocrine correlates of alternative phenotypes in the white-throated sparrow ( Zonotrichia albicollis)

The White-throated Sparrow is a passerine bird of the American sparrow family Emberizidae, which has been studied extensively as it maintains a 100Mb inversion polymorphism on chromosome 2 via disassortative mating. The inverted arrangement is maintained in a near constant state of heterozygosity. Approximately half of the population is homozygous for the ZAL2 arrangement, whereas the other half of the population has the heterozygous condition. The exceptional inversion polymorphism (ZAL2m) in the white-throated sparrow (Zonotrichia albicollis) is linked to variation in plumage, social behavior and mate choice, and is maintained in the population by negative assortative mating.

Coevolution of Genome Architecture and Social Behavior

The social behavior of male rats administered an adult-onset calorie restriction regimen

Here we employed the partner preference test (PPT) to examine how naked mole-rat non-breeding individuals of different behavioral phenotypes make social decisions. Naked mole-rats from six colonies were classified into three behavioral phenotypes (soldiers, dispersers, and workers) using a battery of behavioral tests. They then participated in a 3 h long PPT, where they could freely interact with a tethered familiar or tethered unfamiliar conspecific. By comparing the three behavioral phenotypes, we tested the hypothesis that the PPT can be used to interrogate social decision-making in this species, revealing individual differences in behavior that are consistent with discrete social phenotypes. We also tested whether a shorter, 10 min version of the paradigm is sufficient to capture group differences in behavior. Overall, soldiers had higher aggression scores toward unfamiliar conspecifics than both workers and dispersers at the 10 min and 3 h comparison times. At the 10 min comparison time, workers showed a stronger preference for the familiar animal’s chamber, as well as for investigating the familiar conspecific, compared to both dispersers and soldiers. At the 3 h time point, no phenotype differences were seen with chamber or investigation preference scores. Overall, all phenotypes spent more time in chambers with another animal vs. being alone. Use of the PPT in a comparative context has demonstrated that the test identifies species and group differences in affiliative and aggressive behavior toward familiar and unfamiliar animals, revealing individual differences in social decision-making and, importantly, capturing aspects of species-specific social organization seen in nature.