Diversification of takeout, a male-biased gene family in Drosophila

The display of courtship behavior has evolved in response to sexual selection driven by competition to obtain mates. Sexually dimorphic mate selection rituals are likely controlled at least in part by genes with sex-biased patterns of expression. In Drosophila melanogaster, male courtship behavior has been well described and consists of a series of stereotyped behaviors. The takeout gene is predominantly expressed in males and affects male courtship behavior. In this study, we examine the patterns of expression and evolution in takeout and the family of related proteins. We show that a number of genes in the takeout gene family show male-biased expression in D. melanogaster, largely in non-reproductive tissues. Phylogenetic analysis reveals that this gene family is conserved across insects. As expected for genes with male-biased expression, we also find evidence of positive selection in some lineages. Our results suggest that the genes in this family may have evolutionarily conserved sex specific roles in male mating behavior across insects.     

Genes expressed in the Drosophila head reveal a role for fat cells in sex-specific physiology

The downstream effectors of the Drosophila sex determination cascade are mostly unknown and thought to mediate all aspects of sexual differentiation, physiology and behavior. Here, we employed serial analysis of gene expression (SAGE) to identify male and female effectors expressed in the head, and report 46 sex-biased genes (>4-fold/P < 0.01). We characterized four novel, male- or female-specific genes and found that all are expressed mainly in the fat cells in the head. Tsx (turn on sex-specificity), sxe1 and sxe2 (sex-specific enzyme 1/2) are expressed in males, but not females, and are dependent on the known sex determination pathway, specifically transformer (tra) and its downstream target doublesex (dsx). Female-specific expression of the fourth gene, fit (female-specific independent of transformer), is not controlled by tra and dsx, suggesting an alternative pathway for the regulation of some effector genes. Our results indicate that fat cells in the head express sex-specific effectors, thereby generating distinct physiological conditions in the male and female head. We suggest that these differences have consequences on the male and female brain by modulating sex-specific neuronal processes.     

Courtship song, male agonistic encounters, and female mate choice in the house cricket,Acheta domesticus (Orthoptera: Gryllidae)

Whether female crickets choose among males based on characteristics of the courtship song is uncertain, but in many species, males not producing courtship song do not mate. In the house cricket,Acheta domesticus, we examined whether a female chose or rejected a male based on his size, latency to chirp, latency to produce courtship song, or rate of the high-frequency pulse of courtship song (“court rate”). We confirmed that females mated only with males that produced courtship song, but we found no evidence that the other factors we measured affected a female’s decision to mate. In addition, we investigated whether the outcome of male agonistic encounters affected the subsequent production of courtship song. In one experiment, we observed courtship and mating behavior when a single female was placed with a pair of males following a 10-min interaction period between the two males. Winners of male agonistic encounters had higher mating success. However, winners and losers of agonistic encounters were not different in their likelihood or latency to produce courtship song or in the number of times they were disrupted by the other male in the pair. In a second experiment, we allowed two males to interact for a 10-min period, but following this interaction period, we placed a female with each male separately and observed courtship and mating behavior. The mating success of winners and losers was not different under these circumstances, and we found no differences between winners and losers in any subsequent courtship or mating behavior examined. We conclude that winning agonistic encounters influences a male’s mating success in ways other than his production of courtship song and this effect is lost when winning and losing males are separated and each is given an opportunity to mate.     

Male Siamese Fighting Fish, Betta splendens, Increase Rather than Conceal Courtship Behavior when a Rival is Present

The impact of social environment on mating success is especially pronounced in species where both intraspecific and interspecific selection influence reproduction, such as the Siamese fighting fish. Males alter male–male interactions when either a male or female audience is present, but how males change their behavior toward a female when a rival male is present is unknown. This study addresses whether males alter their behavior toward a female in a way that would prevent a rival male from interrupting courtship. The behavior of male Siamese fighting fish toward a dummy female was examined under various degrees of visual cover, both in the presence and absence of a rival male, to investigate whether males use concealment provided by the structural environment to their advantage. While males did not use barriers to conceal courtship as hypothesized, males altered their behavior by increasing courtship and monitoring their nest when a rival was visible. This increase in courtship is in contrast to most studies on courtship in the presence of a rival that find a reduction in courtship behavior. Males spent more time opercular gill flaring when no barriers were present, suggesting that males may be trying to court the female and communicate to the rival simultaneously. There was also a trend for aggression toward the female and the rival to decrease as screen length increased. Thus, males compensate for the presence of a rival by adjusting their courtship and aggressive behaviors, which could have important implications for courtship success.     

The genetics of mating recognition between Drosophila simulans and D. sechellia

During courtship, visual and chemical signals are often exchanged between the sexes. The proper exchange of such signals ensures intraspecific recognition. We have examined the genetic basis of interspecific differences in male mating behaviour and pheromone concentration between Drosophila simulans and D. sechellia by using Drosophila simulans/D. sechellia introgression lines. Our results show a majority of quantitative trait loci (QTLs) explaining variation in both male mating behaviour and pheromone concentration to be located on the third chromosome. One QTL found on the third chromosome explains variation in time needed to start courtship and copulation as well as time spent courting. The position of such QTL (approximately 84A-88B) with effects on courtship and copulation aspects of mating includes the candidate sex determination gene doublesex (84E5-6) and Voila (86E1-2), a gene that affects male courtship in D. melanogaster. One additional third chromosome QTL explained variation in 7-tricosene pheromone concentrations among males. The interval mapping position of this QTL (approximately 68E-76E) did not overlap with the position detected for differences in mating behaviour and the intervals did not include candidate genes previously identified as having an effect on D. melanogaster cuticular hydrocarbon production. We did not detect any directionality of the effect of Drosophila sechellia allele introgressions in male mating recognition.     

fruitless在桔小实蝇求偶和交配行为中的作用

【目的】fruitless (fru)是昆虫求偶和交配行为中的关键基因,但是其在桔小实蝇Bactrocera dorsalis中的具体功能尚不清楚。本研究旨在明确fru在桔小实蝇求偶交配行为中的作用,进一步明确fru的生物学意义。【方法】根据NCBI上桔小实蝇fru的预测序列设计两端引物,以桔小实蝇羽化10 d已交配的成虫头部cDNA为模板克隆fru基因的全长cDNA,并对其编码蛋白的结构域进行预测。通过实时荧光定量PCR(RT-qPCR)技术检测该基因在桔小实蝇交配前后表达量的变化。待合成fru基因的dsRNA后,注射到羽化9 d后的桔小实蝇雄成虫腹部进行RNAi,分别于注射后48 h和72 h取样,检测fru基因的相对表达量;并于RNAi后72 h进行求偶交配行为观察,测定桔小实蝇的交配频率和交配持续时间。【结果】克隆获得桔小实蝇fru的全长cDNA,长2 865 bp,编码954个氨基酸,预测蛋白分子量104.1 kD,等点为6.01。该基因所编码的蛋白具有特殊结构域,分别为一个BTB (Broad-complex, Tramtrack and Bric-a-bric)结构域和两个锌指(zinc finger)结构域。RT-qPCR结果表明,相比未交配的桔小实蝇成虫,fru基因在雄成虫头部中的表达量在与雌成虫交配后显著上升。相对于空白对照组(注射DEPC水)和阴性对照组(注射dsGFP),注射dsfru的RNAi处理组的雄性求偶时间延长(延长了25~35 min),且交配频率下降(降低了17%~22%),说明fru基因在雄性桔小实蝇的求偶交配行为中发挥着重要作用。【结论】本研究结果为明确fru在非模式昆虫桔小实蝇求偶交配行为中的功能提供参考,并为深入研究桔小实蝇的求偶交配行为和绿色防控提供了理论基础。     

Gene expression changes in male accessory glands during ageing are accompanied by reproductive decline in Drosophila melanogaster

Senescence is accompanied by loss of reproductive functions. Here, we studied reproductive ageing in Drosophila melanogaster males and asked whether the expected decline in male reproductive success is due to diminished functionality of the male accessory gland (AG). The male AG produces the majority of seminal fluid proteins (SFPs) transferred to the female at mating. SFPs induce female postmating changes and are key to male reproductive success. We measured age‐dependent gene expression changes for five representative SFP genes in males from four different age groups ranging from 1 to 6 weeks after eclosion. Simultaneously, we also measured male reproductive success in postmating traits mediated by transfer of these five SFPs. We found a decreased in male SFP gene expression with advancing age and an accompanying decline in male postmating success. Hence, male reproductive senescence is associated with a decline in functionality of the male AG. While overall individual SFP genes decreased in expression, our results point towards the idea that the composition of an ejaculate might change with male age as the rate of change was variable for those five genes.     

Experimentally induced variation in neuroendocrine processes affects male reproductive behaviour,sperm characteristics and social interactions

While extensive research has focused on how social interactions evolve, the fitness consequences of the neuroendocrine mechanisms underlying these interactions have rarely been documented, especially in the wild. Here, we measure how the neuroendocrine mechanisms underlying male behaviour affect mating success and sperm competition in the ocellated wrasse (Symphodus ocellatus). In this species, males exhibit three alternative reproductive types. “Nesting males” provide parental care, defend territories and form cooperative associations with unrelated “satellites,” who cheat by sneaking fertilizations but help by reducing sperm competition from “sneakers” who do not cooperate or provide care. To measure the fitness consequences of the mechanisms underlying these social interactions, we used “phenotypic engineering” that involved administering an androgen receptor antagonist (flutamide) to wild, free‐living fish. Nesting males treated with flutamide shifted their aggression from sneakers to satellite males and experienced decreased submissiveness by sneaker males (which correlated with decreased nesting male mating success). The preoptic area (POA), a region controlling male reproductive behaviours, exhibited dramatic down‐regulation of androgen receptor (AR) and vasotocin 1a receptor (V1aR) mRNA following experimental manipulation of androgen signalling. We did not find a direct effect of the manipulation on male mating success, paternity or larval production. However, variation in neuroendocrine mechanisms generated by the experimental manipulation was significantly correlated with changes in behaviour and mating success: V1aR expression was negatively correlated with satellite‐directed aggression, and expression of its ligand arginine vasotocin (AVT) was positively correlated with courtship and mating success, thus revealing the potential for sexual selection on these mechanisms.     

Highly tissue specific expression of Sphinx supports its male courtship related role in Drosophila melanogaster

Sphinx is a lineage-specific non-coding RNA gene involved in regulating courtship behavior in Drosophila melanogaster. The 5' flanking region of the gene is conserved across Drosophila species, with the proximal 300 bp being conserved out to D. virilis and a further 600 bp region being conserved amongst the melanogaster subgroup (D. melanogaster, D. simulans, D. sechellia, D. yakuba, and D. erecta). Using a green fluorescence protein transformation system, we demonstrated that a 253 bp region of the highly conserved segment was sufficient to drive sphinx expression in male accessory gland. GFP signals were also observed in brain, wing hairs and leg bristles. An additional ～800 bp upstream region was able to enhance expression specifically in proboscis, suggesting the existence of enhancer elements. Using anti-GFP staining, we identified putative sphinx expression signal in the brain antennal lobe and inner antennocerebral tract, suggesting that sphinx might be involved in olfactory neuron mediated regulation of male courtship behavior. Whole genome expression profiling of the sphinx knockout mutation identified significant up-regulated gene categories related to accessory gland protein function and odor perception, suggesting sphinx might be a negative regulator of its target genes.     

Exploring the influence of individual courtship behaviors on male mating success in a blow fly

Male courtship behavior and displays influence female mating decisions, and therefore affect mating success in a diverse range of organisms. While there is substantial evidence confirming that females prefer males who invest more in courtship, less attention has been paid to the relative importance of individual behaviors, and the discrete sequences of courtship that result in mating success. The small hairy maggot blow fly, Chrysomya varipes, performs stereotyped courtship behaviors, involving orienting, tapping, waving, arching, wing vibration and mounting. This study aimed to quantify male investment in specific courtship behaviors, and compare courtship investment and behavioral transitions between males who gained mating success (successful males) and those that did not (unsuccessful males). Our results show that mating success was influenced by the behaviors orienting, tapping, arching and mounting. Behavioral transitions revealed a distinct pattern of behaviors leading to a mating attempt, and some differences were observed between successful and unsuccessful males. Overall, our findings suggest that female mating decisions were based on differences in specific male courtship behaviors. This detailed observational study has quantified multiple courtship behaviors for the first time in C. varipes, and highlights the importance of considering multiple behavioral traits when exploring the influence of male courtship on mating success.