XIST directly regulates X-linked and autosomal genes in naive human pluripotent cells.

X chromosome inactivation (XCI) serves as a paradigm for RNA-mediated regulation of gene expression, wherein the long non-coding RNA XIST spreads across the X chromosome in cis to mediate gene silencing chromosome-wide. In female naive human pluripotent stem cells (hPSCs), XIST is in a dispersed configuration, and XCI does not occur, raising questions about XIST's function. We found that XIST spreads across the X chromosome and induces dampening of X-linked gene expression in naive hPSCs. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences in autosomes. The dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our study identifies XIST as the regulator of X chromosome dampening, uncovers an evolutionarily conserved trans-acting role of XIST/Xist, and reveals a correlation between XIST/Xist dispersal and autosomal targeting.

Generating parallel representations of position and identity in the olfactory system.

In Drosophila, a dedicated olfactory channel senses a male pheromone, cis-vaccenyl acetate (cVA), promoting female courtship while repelling males. Here, we show that separate cVA-processing streams extract qualitative and positional information. cVA sensory neurons respond to concentration differences in a 5-mm range around a male. Second-order projection neurons encode the angular position of a male by detecting inter-antennal differences in cVA concentration, which are amplified through contralateral inhibition. At the third circuit layer, we identify 47 cell types with diverse input-output connectivity. One population responds tonically to male flies, a second is tuned to olfactory looming, while a third integrates cVA and taste to coincidentally promote female mating. The separation of olfactory features resembles the mammalian what and where visual streams; together with multisensory integration, this enables behavioral responses appropriate to specific ethological contexts.

A circuit logic for sexually shared and dimorphic aggressive behaviors in Drosophila.

Aggression involves both sexually monomorphic and dimorphic actions. How the brain implements these two types of actions is poorly understood. We have identified three cell types that regulate aggression in Drosophila: one type is sexually shared, and the other two are sex specific. Shared common aggression-promoting (CAP) neurons mediate aggressive approach in both sexes, whereas functionally downstream dimorphic but homologous cell types, called male-specific aggression-promoting (MAP) neurons in males and fpC1 in females, control dimorphic attack. These symmetric circuits underlie the divergence of male and female aggressive behaviors, from their monomorphic appetitive/motivational to their dimorphic consummatory phases. The strength of the monomorphic → dimorphic functional connection is increased by social isolation in both sexes, suggesting that it may be a locus for isolation-dependent enhancement of aggression. Together, these findings reveal a circuit logic for the neural control of behaviors that include both sexually monomorphic and dimorphic actions, which may generalize to other organisms.

Sexually Dimorphic Control of Parenting Behavior by the Medial Amygdala.

Social behaviors, including behaviors directed toward young offspring, exhibit striking sex differences. Understanding how these sexually dimorphic behaviors are regulated at the level of circuits and transcriptomes will provide insights into neural mechanisms of sex-specific behaviors. Here, we uncover a sexually dimorphic role of the medial amygdala (MeA) in governing parental and infanticidal behaviors. Contrary to traditional views, activation of GABAergic neurons in the MeA promotes parental behavior in females, while activation of this population in males differentially promotes parental versus infanticidal behavior in an activity-level-dependent manner. Through single-cell transcriptomic analysis, we found that molecular sex differences in the MeA are specifically represented in GABAergic neurons. Collectively, these results establish crucial roles for the MeA as a key node in the neural circuitry underlying pup-directed behaviors and provide important insight into the connection between sex differences across transcriptomes, cells, and circuits in regulating sexually dimorphic behavior.

Multimodal Analysis of Cell Types in a Hypothalamic Node Controlling Social Behavior.

The ventrolateral subdivision of the ventromedial hypothalamus (VMHvl) contains ∼4,000 neurons that project to multiple targets and control innate social behaviors including aggression and mounting. However, the number of cell types in VMHvl and their relationship to connectivity and behavioral function are unknown. We performed single-cell RNA sequencing using two independent platforms-SMART-seq (∼4,500 neurons) and 10x (∼78,000 neurons)-and investigated correspondence between transcriptomic identity and axonal projections or behavioral activation, respectively. Canonical correlation analysis (CCA) identified 17 transcriptomic types (T-types), including several sexually dimorphic clusters, the majority of which were validated by seqFISH. Immediate early gene analysis identified T-types exhibiting preferential responses to intruder males versus females but only rare examples of behavior-specific activation. Unexpectedly, many VMHvl T-types comprise a mixed population of neurons with different projection target preferences. Overall our analysis revealed that, surprisingly, few VMHvl T-types exhibit a clear correspondence with behavior-specific activation and connectivity.

Limbic Neurons Shape Sex Recognition and Social Behavior in Sexually Naive Males.

Sexually naive animals have to distinguish between the sexes because they show species-typical interactions with males and females without meaningful prior experience. However, central neural pathways in naive mammals that recognize sex of other individuals remain poorly characterized. We examined the role of the principal component of the bed nucleus of stria terminalis (BNSTpr), a limbic center, in social interactions in mice. We find that activity of aromatase-expressing BNSTpr (AB) neurons appears to encode sex of other animals and subsequent displays of mating in sexually naive males. Silencing these neurons in males eliminates preference for female pheromones and abrogates mating success, whereas activating them even transiently promotes male-male mating. Surprisingly, female AB neurons do not appear to control sex recognition, mating, or maternal aggression. In summary, AB neurons represent sex of other animals and govern ensuing social behaviors in sexually naive males.

Brain-wide Maps Reveal Stereotyped Cell-Type-Based Cortical Architecture and Subcortical Sexual Dimorphism.

The stereotyped features of neuronal circuits are those most likely to explain the remarkable capacity of the brain to process information and govern behaviors, yet it has not been possible to comprehensively quantify neuronal distributions across animals or genders due to the size and complexity of the mammalian brain. Here we apply our quantitative brain-wide (qBrain) mapping platform to document the stereotyped distributions of mainly inhibitory cell types. We discover an unexpected cortical organizing principle: sensory-motor areas are dominated by output-modulating parvalbumin-positive interneurons, whereas association, including frontal, areas are dominated by input-modulating somatostatin-positive interneurons. Furthermore, we identify local cell type distributions with more cells in the female brain in 10 out of 11 sexually dimorphic subcortical areas, in contrast to the overall larger brains in males. The qBrain resource can be further mined to link stereotyped aspects of neuronal distributions to known and unknown functions of diverse brain regions.

piRNA-mediated gene regulation and adaptation to sex-specific transposon expression in D. melanogaster male germline.

Small noncoding piRNAs act as sequence-specific guides to repress complementary targets in Metazoa. Prior studies in Drosophila ovaries have demonstrated the function of the piRNA pathway in transposon silencing and therefore genome defense. However, the ability of the piRNA program to respond to different transposon landscapes and the role of piRNAs in regulating host gene expression remain poorly understood. Here, we comprehensively analyzed piRNA expression and defined the repertoire of their targets in Drosophila melanogaster testes. Comparison of piRNA programs between sexes revealed sexual dimorphism in piRNA programs that parallel sex-specific transposon expression. Using a novel bioinformatic pipeline, we identified new piRNA clusters and established complex satellites as dual-strand piRNA clusters. While sharing most piRNA clusters, the two sexes employ them differentially to combat the sex-specific transposon landscape. We found two piRNA clusters that produce piRNAs antisense to four host genes in testis, including CG12717/pirate, a SUMO protease gene. piRNAs encoded on the Y chromosome silence pirate, but not its paralog, to exert sex- and paralog-specific gene regulation. Interestingly, pirate is targeted by endogenous siRNAs in a sibling species, Drosophila mauritiana, suggesting distinct but related silencing strategies invented in recent evolution to regulate a conserved protein-coding gene.

Interactions between U and V sex chromosomes during the life cycle of Ectocarpus.

In many animals and flowering plants, sex determination occurs in the diploid phase of the life cycle with XX/XY or ZW/ZZ sex chromosomes. However, in early diverging plants and most macroalgae, sex is determined by female (U) or male (V) sex chromosomes in a haploid phase called the gametophyte. Once the U and V chromosomes unite at fertilization to produce a diploid sporophyte, sex determination no longer occurs, raising key questions about the fate of the U and V sex chromosomes in the sporophyte phase. Here, we investigate genetic and molecular interactions of the UV sex chromosomes in both the haploid and diploid phases of the brown alga Ectocarpus. We reveal extensive developmental regulation of sex chromosome genes across its life cycle and implicate the TALE-HD transcription factor OUROBOROS in suppressing sex determination in the diploid phase. Small RNAs may also play a role in the repression of a female sex-linked gene, and transition to the diploid sporophyte coincides with major reconfiguration of histone H3K79me2, suggesting a more intricate role for this histone mark in Ectocarpus development than previously appreciated.

Fossil evidence sheds light on sexual selection during the early evolution of birds.

The impact of sexual selection on the evolution of birds has been widely acknowledged. Although sexual selection has been hypothesized as a driving force in the occurrences of numerous morphological features across theropod evolution, this hypothesis has yet to be comprehensively tested due to challenges in identifying the sex of fossils and by the limited sample size. Confuciusornis sanctus is arguably the best-known early avialan and is represented by thousands of well-preserved specimens from the Early Cretaceous Jehol lagerstätte, which provides us with a chance to decipher the strength of sexual selection on extinct vertebrates. Herein, we present a morphometric study of C. sanctus based on the largest sample size of this taxon collected up to now. Our results indicate that the characteristic elongated paired rectrices is a sexually dimorphic trait and statistically robust inferences of the sexual dimorphism in size, shape, and allometry that have been established, providing the earliest known sexual dimorphism in avian evolution. Our findings suggest that sexual selection, in conjunction with natural selection, does act upon body size and limb length ratio in early birds, thereby promoting a deeper understanding of the role of sexual selection in large-scale phylogenetic evolution.

Sexual conflict drive in the rapid evolution of new gametogenesis genes.

The evolutionary forces underlying the rapid evolution in sequences and functions of new genes remain a mystery. Adaptation by natural selection explains the evolution of some new genes. However, many new genes perform sex-biased functions that have rapidly evolved over short evolutionary time scales, suggesting that new gene evolution may often be driven by conflicting selective pressures on males and females. It is well established that such sexual conflict (SC) plays a central role in maintaining phenotypic and genetic variation within populations, but the role of SC in driving new gene evolution remains essentially unknown. This review explores the connections between SC and new gene evolution through discussions of the concept of SC, the phenotypic and genetic signatures of SC in evolving populations, and the molecular mechanisms by which SC could drive the evolution of new genes. We synthesize recent work in this area with a discussion of the case of Apollo and Artemis, two extremely young genes (<200,000 years) in Drosophila melanogaster, which offered the first empirical insights into the evolutionary process by which SC could drive the evolution of new genes. These new duplicate genes exhibit the hallmarks of sexually antagonistic selection: rapid DNA and protein sequence evolution, essential sex-specific functions in gametogenesis, and complementary sex-biased expression patterns. Importantly, Apollo is essential for male fitness but detrimental to female fitness, while Artemis is essential for female fitness but detrimental to male fitness. These sexually antagonistic fitness effects and complementary changes to expression, sequence, and function suggest that these duplicates were selected for mitigating SC, but that SC has not been fully resolved. Finally, we propose Sexual Conflict Drive as a self-driven model to interpret the rapid evolution of new genes, explain the potential for SC and sexually antagonistic selection to contribute to long-term evolution, and suggest its utility for understanding the rapid evolution of new genes in gametogenesis.

SPIN: sex-specific and pathway-based interpretable neural network for sexual dimorphism analysis.

Sexual dimorphism in prevalence, severity and genetic susceptibility exists for most common diseases. However, most genetic and clinical outcome studies are designed in sex-combined framework considering sex as a covariate. Few sex-specific studies have analyzed males and females separately, which failed to identify gene-by-sex interaction. Here, we propose a novel unified biologically interpretable deep learning-based framework (named SPIN) for sexual dimorphism analysis. We demonstrate that SPIN significantly improved the C-index up to 23.6% in TCGA cancer datasets, and it was further validated using asthma datasets. In addition, SPIN identifies sex-specific and -shared risk loci that are often missed in previous sex-combined/-separate analysis. We also show that SPIN is interpretable for explaining how biological pathways contribute to sexual dimorphism and improve risk prediction in an individual level, which can result in the development of precision medicine tailored to a specific individual's characteristics.

Pelvic shape change in adult Japanese macaques and implications for childbirth at old age.

Pelvic morphology exhibits a particular sexual dimorphism in humans, which reflects obstetrical constraints due to the tight fit between neonates and mothers. Huseynov et al. [Proc. Natl. Acad. Sci. U.S.A. 113, 5227-5232 (2016)] showed that in humans, pelvic sexual dimorphism is greatest around the age of highest fertility, and it becomes less marked in association with menopause in females. They proposed that this reflects changes of obstetrical versus locomotor functional demands in females. It remains unknown whether such developmental adjustment of the pelvic morphology is unique to humans. Macaques exhibit human-like cephalopelvic proportions, but they lack menopause and usually maintain fertility throughout adulthood. Here, we track pelvic development in Japanese macaques from neonate to advanced ages using computed tomography-based data. We show that female pelvic morphology changes throughout adult life, reaching the obstetrically most favorable shape at advanced ages rather than around primiparity. We hypothesize that pelvic morphology in Japanese macaques is developmentally adjusted to childbirth at advanced ages, where obstetrical risks are potentially higher than at younger ages. Our data contribute to the growing evidence that the female primate pelvis changes its morphology during the whole lifespan, possibly adjusting for changing functional demands during adulthood.

Deep conservation and co-option of programmed cell death facilitates evolution of alternative phenotypes at multiple biological levels.

Alternative phenotypes, such as polyphenisms and sexual dimorphisms, are widespread in nature and appear at all levels of biological organization, from genes and cells to morphology and behavior. Yet, our understanding of the mechanisms through which alternative phenotypes develop and how they evolve remains understudied. In this review, we explore the association between alternative phenotypes and programmed cell death, a mechanism responsible for the elimination of superfluous cells during development. We discuss the ancient origins and deep conservation of programmed cell death (its function, forms and underlying core regulatory gene networks), and propose that it was co-opted repeatedly to generate alternative phenotypes at the level of cells, tissues, organs, external morphology, and even individuals. We review several examples from across the tree of life to explore the conditions under which programmed cell death is likely to facilitate the evolution of alternative phenotypes.

Two C. elegans DM domain proteins, DMD-3 and MAB-3, function in late stages of male somatic gonad development.

To bring about sexual dimorphism in form, information from the sex determination pathway must trigger sex-specific modifications in developmental programs. DM-domain encoding genes have been found to be involved in sex determination in a multitude of animals, often at the level of male somatic gonad formation. Here we report our findings that the DM-domain transcription factors MAB-3 and DMD-3 function together in multiple steps during the late stages of C. elegans male somatic gonad development. Both mab-3 and dmd-3 are expressed in the linker cell and hindgut of L4 males and dmd-3 is also expressed in presumptive vas deferens cells. Furthermore, dmd-3, but not mab-3, expression in the linker cell is downstream of nhr-67, a nuclear hormone receptor that was previously shown to control late stages of linker cell migration. In mab-3; dmd-3 double mutant males, the last stage of linker cell migration is partially defective, resulting in aberrant linker cell shapes and often a failure of the linker cell to complete its migration to the hindgut. When mab-3; dmd-3 double mutant linker cells do complete their migration, they fail to be engulfed by the hindgut, indicating that dmd-3 and mab-3 activity are essential for this process. Furthermore, linker cell death and clearance are delayed in mab-3; dmd-3 double mutants, resulting in the linker cell persisting into adulthood. Finally, DMD-3 and MAB-3 function to activate expression of the bZIP transcription factor encoding gene zip-5 and downregulate the expression of the zinc metalloprotease ZMP-1 in the linker cell. Taken together, these results demonstrate a requirement for DM-domain transcription factors in controlling C. elegans male gonad formation, supporting the notion that the earliest DM-domain genes were involved in male somatic gonad development in the last common ancestor of the bilaterians.

Genetic Basis and Evolutionary Forces of Sexually Dimorphic Color Variation in a Toad-Headed Agamid Lizard.

In the animal kingdom, sexually dimorphic color variation is a widespread phenomenon that significantly influences survival and reproductive success. However, the genetic underpinnings of this variation remain inadequately understood. Our investigation into sexually dimorphic color variation in the desert-dwelling Guinan population of the toad-headed agamid lizard (Phrynocephalus putjatai) utilized a multidisciplinary approach, encompassing phenotypic, ultrastructural, biochemical, genomic analyses, and behavioral experiments. Our findings unveil the association between distinct skin colorations and varying levels of carotenoid and pteridine pigments. The red coloration in males is determined by a genomic region on chromosome 14, housing four pigmentation genes: BCO2 and three 6-pyruvoyltetrahydropterin synthases. A Guinan population-specific nonsynonymous single nucleotide polymorphism in BCO2 is predicted to alter the electrostatic potential within the binding domain of the BCO2-ß-carotene complex, influencing their interaction. Additionally, the gene MAP7 on chromosome 2 emerges as a potential contributor to the blue coloration in subadults and adult females. Sex-specific expression patterns point to steroid hormone-associated genes (SULT2B1 and SRD5A2) as potential upstream regulators influencing sexually dimorphic coloration. Visual modeling and field experiments support the potential selective advantages of vibrant coloration in desert environments. This implies that natural selection, potentially coupled with assortative mating, might have played a role in fixing color alleles, contributing to prevalence in the local desert habitat. This study provides novel insights into the genetic basis of carotenoid and pteridine-based color variation, shedding light on the evolution of sexually dimorphic coloration in animals. Moreover, it advances our understanding of the driving forces behind such intricate coloration patterns.

Sexual Dimorphism of Parental Care: From Genes to Behavior.

Parental care is found in species across the animal kingdom, from small insects to large mammals, with a conserved purpose of increasing offspring survival. Yet enormous variability exists between different species and between the sexes in the pattern and level of parental investment. Here, we review the literature on the neurobiological mechanisms underlying maternal and paternal care, especially in rodents, and discuss the relationship between sex differences in behavior and sexual dimorphism in the brain. We argue that although several brain regions and circuits regulating parental care are shared by both sexes, some of the fundamental components comprising the maternal brain are innate and sex specific. Moreover, we suggest that a more comprehensive understanding of the underlying mechanisms can be achieved by expanding the methodological toolbox, applying ethologically relevant approaches such as nontraditional wild-derived animal models and complex seminatural experimental set-ups.

Sex-specific impact of psychosocial stress on hematopoiesis and blood leukocytes.

Stress exposure has been shown to modulate innate and adaptive immune responses. Indeed, stress favors myelopoiesis and monocyte generation and contributes to cardiovascular disease development. As sex hormones regulate innate and adaptive immune responses, we decided to investigate whether stress exposure leads to a different immune response in female and male mice. Our data demonstrated that psychosocial stressinduced neutrophilia in male, but not female mice. Importantly, we identified that B-cell numbers were reduced in female, but not male mice upon exposure to stress. Thus, our study revealed that the stress-induced immune alterations are sex-dependent, and this is an important feature to consider for future investigations.

Nociceptors are functionally male or female: from mouse to monkey to man.

The prevalence of many pain conditions often differs between sexes. In addition to such quantitative distinctions, sexual dimorphism may also be qualitative reflecting differences in mechanisms that promote pain in men and women. A major factor that influences the likelihood of pain perception is the threshold for activation of nociceptors. Peripheral nociceptor sensitization has been demonstrated to be clinically relevant in many pain conditions. Whether peripheral nociceptor sensitization can occur in a sexually dimorphic fashion, however, has not been extensively studied. To address this fundamental knowledge gap, we used patch clamp electrophysiology to evaluate the excitability of dorsal root ganglion neurones from male or female rodents, non-human primates, and humans following exposure to putative sensitizing agents. Previous studies from our laboratory, and others, have shown that prolactin promotes female-selective pain responses in rodents. Consistent with these observations, dorsal root ganglion neurones from female, but not male, mice were selectively sensitized by exposure to prolactin. The sensitizing action of prolactin was also confirmed in dorsal root ganglion neurones from a female macaque monkey. Critically, neurones recovered from female, but not male, human donors were also selectively sensitized by prolactin. In the course of studies of sleep and pain, we unexpectedly observed that an orexin antagonist could normalize pain responses in male animals. We found that orexin B produced sensitization of male, but not female, mouse, macaque, and human dorsal root ganglion neurones. Consistent with functional responses, increased prolactin receptor and orexin receptor 2 expression was observed in female and male mouse dorsal root ganglia, respectively. Immunohistochemical interrogation of cultured human sensory neurones and whole dorsal root ganglia also suggested increased prolactin receptor expression in females and orexin receptor 2 expression in males. These data reveal a functional double dissociation of nociceptor sensitization by sex, which is conserved across species and is likely directly relevant to human pain conditions. To our knowledge, this is the first demonstration of functional sexual dimorphism in human sensory neurones. Patient sex is currently not a common consideration for the choice of pain therapy. Precision medicine, based on patient sex could improve therapeutic outcomes by selectively targeting mechanisms promoting pain in women or men. Additional implications of these findings are that the design of clinical trials for pain therapies should consider the proportions of male or female patients enrolled. Lastly, re-examination of selected past failed clinical trials with subgroup analysis by sex may be warranted.

The doublesex gene regulates dimorphic sexual and aggressive behaviors in Drosophila.

Most animal species display dimorphic sexual behaviors and male-biased aggressiveness. Current models have focused on the male-specific product from the fruitless (fruM) gene, which controls male courtship and male-specific aggression patterns in fruit flies, and describe a male-specific mechanism underlying sexually dimorphic behaviors. Here we show that the doublesex (dsx) gene, which expresses male-specific DsxM and female-specific DsxF transcription factors, functions in the nervous system to control both male and female sexual and aggressive behaviors. We find that Dsx is not only required in central brain neurons for male and female sexual behaviors, but also functions in approximately eight pairs of male-specific neurons to promote male aggressiveness and approximately two pairs of female-specific neurons to inhibit female aggressiveness. DsxF knockdown females fight more frequently, even with males. Our findings reveal crucial roles of dsx, which is broadly conserved from worms to humans, in a small number of neurons in both sexes to establish dimorphic sexual and aggressive behaviors.