Is there any charm, who has the final say?
Sex differentiation and hormones are critical to the production of animal sexual signals and involve complex top-down gene regulatory networks. However, it is unclear whether core genes can integrate upstream regulatory signals, regulatory signal output, and terminal behavioral responses.
Cockroach Love Skill: Dance wings for love
As the driving force of animal sexual selection, sexual attraction includes both the generation and release of sexual signals and the effective reception of sexual signals by another individual. Many insects rely on volatile pheromones to search for mates and use contact pheromones (CSPs) for close courtship recognition. Blattella germanica is the most common worldwide household pest of cockroaches, with distinct sexual characteristics and clear sexual pheromone components, which is a good model for studying the chemical communication mechanism of gender. For males, what drives them crazy is the long chain (up to C29) lipid mixture on the surface of the female's body, which is very different from the well-known volatile pheromones, which must be physically contacted by the male and female to be felt by the male, stimulating the male to court at close range and inducing sexual mating behavior, of which the male rotates his body 180 degrees and raises his wings is the most typical courtship feature (Figure 1a). The most abundant sexually mature female is 3,11-dimethyl-2-ketone (C29 methyl ketone), which is formed by the precursor substance 3,11-dimethylexacurane by C2-bit hydroxylation and oxidation reaction. Studies have shown that the hydroxylation reaction described above is a female-specific step and is promoted by sparing juvenile hormones. The step has been thought to have been responsible for a female-specific expression of the P450 gene since 1992, but it has not yet been identified.
Molecular 'code' for controlling pheromone synthesis
Professor Li Sheng's team comprehensively used behavioral, biochemical, molecular, genetic and biogenetic analysis methods to screen and identify the CYP4PC1 gene CYP4PC1, which is a key CSP synthesis pathway of German cockroaches, which is expressed only in female insects, and the expression amount continues to rise with the process of sexual maturation. In addition, the gene is highly expressed in the antennae and wings of female insects, thus overturning the previously believed view that "the pigment cells under the skin of the abdomen are the main site of CSP synthesis". The use of RNAi knocked down this gene, resulting in a significant decrease in the CSP content of female insects, and male insects no longer show courtship behavior in these females. After the introduction of females into the control group, the males still exhibited normal courtship behavior. These results suggest that CYP4PC1 is necessary for the maintenance of CSP synthesis and sexual attractiveness in females, and that it is highly likely to be involved in 2-hydroxylation of CSP precursors.
Sexually mature female cockroaches are more attractive
As one of the two endogenous hormones of insects, JH not only inhibits nymph metamorphosis, but also promotes adult reproduction (gonadotropin). The JH titer and JH signal after female feathering rise with the process of sexual maturation. Studies have shown that the 2-hydroxylation step in the CSP synthesis pathway is promoted by JH. To further demonstrate that CYP4PC1 controls this speed-limiting step, the team then examined whether the gene was regulated by JH signaling. Treatment of females with exogenous JH significantly induces CYP4PC1 expression, while RNAi knocks down JH signaling elements (intracellular receptor Met and primary response gene Kr-h1) weakens female sexual attraction. After the males supplement JH, a small part can attract males to courtship (Figure 1b). Therefore, the expression of CYP4PC1 is promoted by the JH-Met-Kr-h1 signaling pathway, thereby maintaining a high sexual pheromone content of females after sexual maturity, stimulating courtship mating of males.
Male cockroaches 'lights out' on the same sex
So why can't males synthesize sex pheromones (even though males also have high levels of CSP precursor compounds)? Both CYP4PC1 expression and CSP synthesis are typically female-specific and are classic sex-type II phenomena. Although high doses of exogenous JH supplementation can induce substantial expression of CYP4PC1 and synthesis of CSP in males, the effect is very limited, suggesting that there may be more important factors inhibiting CYP4PC1 expression in males. It was found that the expression of CYP4PC1 in female insects is regulated by the sex differentiation signaling pathway, and the male-specific product of the intersex gene doublesex (dsx) DsxM protein (DMRT transcription factor) can bind to the CYP4PC1 promoter to inhibit its transcription. It is important to note that knocking down dsxM in male adults can lead to typical homosexual behavior (Figure 1c), and this behavior can be rescued by CYP4PC1 knockdown, indicating that DsxM avoids male attraction to each other by inhibiting CSP synthesis. Unlike existing studies, which have focused mostly on neurological changes, the above results highlight the role of sexual attraction changes in mediating sexual behavior, providing new insights into the mechanisms of same-sex sexual behavior in animals.
Conceptual framework
Through a series of competitive courtship behavioral analyses, CYP4PC1 controls CSP production and regulates male courtship index in a dose-dependent manner (Figures 2a, b). On this basis, the team proposed a theoretical system for the regulation of sexual attraction molecules: the sex differentiation gene dsxM plays a 'brake' role in males (directly inhibits CYP4PC1 expression), transformer guides dsxF variable splicing in females to remove brakes, and JH signals are further exerted in females' Accelerator' role (promotes high expression of CYP4PC1 during sexual maturation in females), thereby mediating sex and age-specific sexual attraction (Figure 2c). It is worth noting that by simultaneously manipulating sex differentiation and hormone signaling in males (removing the 'brakes' and applying 'accelerators'), males can be made to produce sexual attraction beyond sexually mature females (Figure 3), further demonstrating the decisive role of the above pathway in regulatory pheromone synthesis.
Figure 3. Behavioral analysis of competitive courtship
