ESR1 mediates estrogen-induced feminization of genetic male Chinese soft-shelled turtle†.

Exogenous estrogen have shown their feminization abilities during the specific sex differentiation period in several reptiles. However, the specific regulatory mechanism and downstream regulatory genes of estrogen remain elusive. In the present study, 17ß-estradiol (E2), as well as drugs of specific antagonists and/or agonists of estrogen receptors, were employed to figure out the molecular pathway involved in the E2-induced feminization in Chinese soft-shelled turtles, an important aquaculture species in China. E2 treatment led to typical female characteristics in the gonads of ZZ individuals, including thickened outer cortex containing a number of germ cells and degenerated medullary cords, as well as the disappearance of male marker SOX9, and the ectopic expression of ovarian regulator FOXL2 at the embryonic developmental stage 27 and 1 month after hatching. The specific ESR1 antagonist or a combination of three estrogen receptor antagonists could block the sex reversal of ZZ individuals induced by estrogen. In addition, specific activation of ESR1 by agonist also led to the feminization of ZZ gonads, which was similar to the effect of estrogen treatment. Furthermore, transcriptome data showed that the expression level of FOXL2 was significantly upregulated, whereas mRNA levels of DMRT1, SOX9, and AMH were downregulated after estrogen treatment. Taken together, our results indicated that E2 induced the feminization of ZZ Chinese soft-shelled turtles via ESR1, and decrease of male genes DMRT1, SOX9, and AMH and increase of ovarian development regulator FOXL2 might be responsible for the initiation of E2-induced feminization.

DMRT1 gene disruption alone induces incomplete gonad feminization in chicken.

Compared with the well-described XY sex determination system in mammals, the avian ZW sex determination system is poorly understood. Knockdown and overexpression studies identified doublesex and mab-3-related transcription factor 1 (DMRT1) as the testis-determining gene in chicken. However, the detailed effects of DMRT1 gene disruption from embryonic to adult development are not clear. Herein, we have generated DMRT1-disrupted chickens using the clustered regularly interspaced short palindromic repeats-associated protein 9 system, followed by an analysis of physiological, hormonal, and molecular changes in the genome-modified chickens. In the early stages of male chicken development, disruption of DMRT1 induced gonad feminization with extensive physiological and molecular changes; however, functional feminine reproductivity could not be implemented with disturbed hormone synthesis. Subsequent RNA-sequencing analysis of the DMRT1-disrupted chicken gonads revealed gene networks, including several novel genes linearly and non-linearly associated with DMRT1, which are involved in gonad feminization. By comparing the gonads of wild type with the genome-modified chickens, a set of genes were identified that is involved in the ZW sex determination system independent of DMRT1. Our results extend beyond the Z-dosage hypothesis to provide further information about the avian ZW sex determination system and epigenetic effects of gonad feminization.

2,4-Dichlorophenol induced feminization of zebrafish by down-regulating male-related genes through DNA methylation.

2,4-Dichlorophenol (2,4-DCP) is ubiquitous in aquatic environment and has potential estrogenic effect on fish. However, the effect of 2,4-DCP on sex differentiation of zebrafish (Danio rerio) and the underlying mechanism are largely unknown. To address these questions, zebrafish larvae at 20 or 30 days post fertilization (dpf) were exposed to 2,4-DCP (0, 80 and 160 µg L-1) with/without 5-aza-2'-deoxycytidine (5AZA, 50 µg L-1) for 10 days. The sex ratios and the expressions of male-related genes including amh, gata4, nr5a1a, nr5a2 and sox9a were analyzed. In addition, the DNA methylation levels of amh, nr5a2 and sox9a were examined. The results showed that 2,4-DCP exposure resulted in significant increase of female ratios both in 20-30 and 30-40 dpf groups. Correspondingly, the expressions of gata4, nr5a1a, nr5a2 and sox9a were decreased by 2,4-DCP exposure in two treatment periods. However, the transcript of amh was decreased by 2,4-DCP exposure only from 30 to 40 dpf. The DNA methylation levels of amh, nr5a2 and sox9a were increased following 2,4-DCP exposure. Moreover, the addition of 5AZA could counteract the effects including feminization, disturbance of gene expression and DNA hypermethylation caused by 2,4-DCP. These results indicated that the feminizing effect of 2,4-DCP was accomplished by regulating the expression of male-related genes through DNA methylation.

DMRT1 prevents female reprogramming in the postnatal mammalian testis.

Sex in mammals is determined in the fetal gonad by the presence or absence of the Y chromosome gene Sry, which controls whether bipotential precursor cells differentiate into testicular Sertoli cells or ovarian granulosa cells. This pivotal decision in a single gonadal cell type ultimately controls sexual differentiation throughout the body. Sex determination can be viewed as a battle for primacy in the fetal gonad between a male regulatory gene network in which Sry activates Sox9 and a female network involving WNT/ß-catenin signalling. In females the primary sex-determining decision is not final: loss of the FOXL2 transcription factor in adult granulosa cells can reprogram granulosa cells into Sertoli cells. Here we show that sexual fate is also surprisingly labile in the testis: loss of the DMRT1 transcription factor in mouse Sertoli cells, even in adults, activates Foxl2 and reprograms Sertoli cells into granulosa cells. In this environment, theca cells form, oestrogen is produced and germ cells appear feminized. Thus Dmrt1 is essential to maintain mammalian testis determination, and competing regulatory networks maintain gonadal sex long after the fetal choice between male and female. Dmrt1 and Foxl2 are conserved throughout vertebrates and Dmrt1-related sexual regulators are conserved throughout metazoans. Antagonism between Dmrt1 and Foxl2 for control of gonadal sex may therefore extend beyond mammals. Reprogramming due to loss of Dmrt1 also may help explain the aetiology of human syndromes linked to DMRT1, including disorders of sexual differentiation and testicular cancer.

Androgen receptor promotes sex-independent angiogenesis in response to ischemia and is required for activation of vascular endothelial growth factor receptor signaling.

BACKGROUND: Hypoandrogenemia is associated with an increased risk of ischemic diseases. Because actions of androgens are exerted through androgen receptor (AR) activation, we studied hind-limb ischemia in AR knockout mice to elucidate the role of AR in response to ischemia. METHODS AND RESULTS: Both male and female AR knockout mice exhibited impaired blood flow recovery, more cellular apoptosis, and a higher incidence of autoamputation after ischemia. In ex vivo and in vivo angiogenesis studies, AR-deficient vascular endothelial cells showed reduced angiogenic capability. In ischemic limbs of AR knockout mice, reductions in the phosphorylation of the Akt protein kinase and endothelial nitric oxide synthase were observed despite a robust increase in hypoxia-inducible factor 1α and vascular endothelial cell growth factor (VEGF) gene expression. In in vitro studies, siRNA-mediated ablation of AR in vascular endothelial cells blunted VEGF-stimulated phosphorylation of Akt and endothelial nitric oxide synthase. Immunoprecipitation experiments documented an association between AR and kinase insert domain protein receptor that promoted the recruitment of downstream signaling components. CONCLUSIONS: These results document a physiological role of AR in sex-independent angiogenic potency and provide evidence of novel cross-talk between the androgen/AR signaling and VEGF/kinase insert domain protein receptor signaling pathways.

Microbial manipulation of host sex determination. Endosymbiotic bacteria can directly manipulate their host's sex determination towards the production of female offspring.

A recent study in the lepidopteran Ostrinia scapulalis shows that endosymbionts can actively manipulate the sex determination mechanism of their host. Wolbachia bacteria alter the sex-specific splicing of the doublesex master switch gene. In ZZ males of this female heterogametic system, the female isoform of doublesex is produced in the presence of the bacteria. The effect is a lethal feminization of genotypic males. Curing of ZW females leads to males that die, indicating that the bacteria have an obligate role in proper sex determination and development of their host. Microbial intervention with host sex determination may be a driving force behind the evolutionary turnover of sex determination mechanisms.

5 mC modification of steroid hormone biosynthesis-related genes orchestrates feminization of channel catfish induced by high-temperature.

To elucidate the mechanism behind channel catfish feminization induced by high temperature, gonad samples were collected from XY pseudo-females and wild-type females and subjected to high-throughput sequencing for Whole-Genome-Bisulfite-Seq (WGBS) and transcriptome sequencing (RNA-Seq). The analysis revealed 50 differentially methylated genes between wild-type females and XY pseudo-females, identified through the analysis of KEGG pathways and GO enrichment in the promoter of the genome and differentially methylated regions (DMRs). Among these genes, multiple differential methylation sites observed within the srd5a2 gene. Repeatability tests confirmed 7 differential methylation sites in the srd5a2 gene in XY pseudo-females compared to normal males, with 1 specific differential methylation site (16608174) distinguishing XY pseudo-females from normal females. Interestingly, the expression of these genes in the transcriptome showed no difference between wild-type females and XY pseudo-females. Our study concluded that methylation of the srd5a2 gene sequence leads to decreased expression, which inhibits testosterone synthesis while promoting the synthesis of 17ß-estradiol from testosterone. This underscores the significance of the srd5a2 gene in the sexual differentiation of channel catfish, as indicated by the ipu00140 KEGG pathway analysis.

Ethynylestradiol feminizes gene expression partly in testis developing as ovotestis and disrupts asymmetric Müllerian duct development by eliminating asymmetric gene expression in Japanese quail embryos.

In avian embryos, xenoestrogens induce abnormalities in reproductive organs, particularly the testes and Müllerian ducts (MDs). However, the molecular mechanisms remain poorly understood. We investigated the effects of ethynylestradiol (EE2) exposure on gene expression associated with reproductive organ development in Japanese quail embryos. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis revealed that the left testis containing ovary-like tissues following EE2 exposure highly expressed the genes for steroidogenic enzymes (P450scc, P45017α, lyase, and 3ß-HSD) and estrogen receptor-ß, compared to the right testis. No asymmetry was found in these gene expression without EE2. EE2 induced hypertrophy in female MDs and suppressed atrophy in male MDs on both sides. RNA sequencing analysis of female MDs showed 1,366 differentially expressed genes between developing left MD and atrophied right MD in the absence of EE2, and these genes were enriched in Gene Ontology terms related to organogenesis, including cell proliferation, migration and differentiation, and angiogenesis. However, EE2 reduced asymmetrically expressed genes to 21. RT-qPCR analysis indicated that genes promoting cell cycle progression and oncogenesis were more highly expressed in the left MD than in the right MD, but EE2 eliminated such asymmetric gene expression by increasing levels on the right side. EE2-exposed males showed overexpression of these genes in both MDs. This study reveals part of the molecular basis of xenoestrogen-induced abnormalities in avian reproductive organs, where EE2 may partly feminize gene expression in the left testis, developing as the ovotestis, and induce bilateral MD malformation by canceling asymmetric gene expression underlying MD development.

The potential regulatory role of the non-coding RNAs in regulating the exogenous estrogen-induced feminization in Takifugu rubripes gonad.

Estrogen plays a pivotal role in the early stage of sex differentiation in teleost. However, the underlying mechanisms of estrogen-induced feminization process are still needed for further clarification. Here, the comparative analysis of whole-transcriptome RNA sequencing was conducted between 17beta-Estradiol induced feminized XY (E-XY) gonads and control gonads (C) in Takifugu rubripes. A total of 57 miRNAs, 65 lncRNAs, and 4 circRNAs were found to be expressed at lower levels in control-XY (C-XY) than that in control-XX (C-XX), and were up-regulated in XY during E2-induced feminization process. The expression levels of 24 miRNAs, and 55 lncRNAs were higher in C-XY than that in C-XX, and were down-regulated in E2-treated XY. Furthermore, a correlation analysis was performed between miRNA-seq and mRNA-seq data. In C-XX/C-XY, 114 differential expression (DE) miRNAs were predicted to target to 904 differential expression genes (DEGs), while in C-XY/E-XY, 226 DEmiRNAs were predicted to target to 2,048 DEGs. In C-XX/C-XY, and C-XY/E-XY, KEGG pathway enrichment analysis showed that those targeted genes were mainly enriched in MAPK signaling, calcium signaling, steroid hormone biosynthesis and ovarian steroidogenesis pathway. Additionally, the competitive endogenous RNA (ceRNA) regulatory network was constructed by 24 miRNAs, 21 lncRNAs, 4 circRNAs and 5 key sex-related genes. These findings suggested that the expression of critical genes in sex differentiation were altered in E2-treated XY T. rubripes may via the lncRNA-miRNA-mRNA regulation network to facilitate the differentiation and maintenance of ovaries. Our results provide a new insight into the comprehensive understanding of the effects of estrogen signaling pathways on sex differentiation in teleost gonads.

Dysregulation of neonatal hippocampal cell genesis in the androgen insensitive Tfm rat.

The first two weeks of life are a critical period for hippocampal development. At this time gonadal steroid exposure organizes sex differences in hippocampal sensitivity to activational effects of steroids, hippocampal cell morphology and hippocampus dependent behaviors. Our laboratory has characterized a robust sex difference in neonatal neurogenesis in the hippocampus that is mediated by estradiol. Here, we extend our knowledge of this sex difference by comparing the male and female hippocampus to the androgen insensitive testicular feminized mutant (Tfm) rat. In the neonatal Tfm rat hippocampus, fewer newly generated cells survive compared to males or females. This deficit in cell genesis is partially recovered with the potent androgen DHT, but is more completely recovered following estradiol administration. Tfm rats do not differ from males or females in the level of endogenous estradiol in the neonatal hippocampus, suggesting other mechanisms mediate a differential sensitivity to estradiol in male, female and Tfm hippocampus. We also demonstrate disrupted performance on a hippocampal-dependent contextual fear discrimination task. Tfm rats generalize fear across contexts, and do not exhibit significant loss of fear during extinction exposure. These results extend prior reports of exaggerated response to stress in Tfm rats, and following gonadectomy in normal male rats.

Sex-specific DNA methylation and transcription of zbtb38 and effects of gene-environment interactions on its natural antisense transcript in zebrafish.

There is increasing evidence for the involvement of epigenetics in sex determination, maintenance, and plasticity, from plants to humans. In our previous work, we reported a transgenerational feminization of a zebrafish population for which the first generation was exposed to cadmium, a metal with endocrine disrupting effects. In this study, starting from the previously performed whole methylome analysis, we focused on the zbtb38 gene and hypothesized that it could be involved in sex differentiation and Cd-induced offspring feminization. We observed sex-specific patterns of both DNA methylation and RNA transcription levels of zbtb38. We also discovered that the non-coding exon 3 of zbtb38 encodes for a natural antisense transcript (NAT). The activity of this NAT was found to be influenced by both genetic and environmental factors. Furthermore, increasing transcription levels of this NAT in parental gametes was highly correlated with offspring sex ratios. Since zbtb38 itself encodes for a transcription factor that binds methylated DNA, our results support a non-negligible role of zbtb38 not only in orchestrating the sex-specific transcriptome (i.e., sex differentiation) but also, via its NAT, offspring sex ratios.

Effective silencing of Sry gene with RNA interference in developing mouse embryos resulted in feminization of XY gonad.

Delivering siRNA or shRNA into the developing embryos is still a main challenge to use of RNAi in mammalian systems. Here we analyze several factors influencing RNAi-mediated silencing of Sry gene, which is a tightly controlled spatiotemporal expressed gene and only shortly expressed in developing mouse embryo gonad. A Sry gene-specific shRNAs expression vector (pSilencer4.1/Sry565) was constructed. The shRNA constructs were mixed with polyethylenimines (PEIs) to form a complex and then injected into pregnant mice though tail vein. Our results showed that Sry gene was downregulated significantly in developing embryos. Further study revealed that knocking-down of Sry expression resulted in feminization of gonad development in mouse embryos and the expression level of Sox9 and Wt1 gene was also significantly changed by downregulation of Sry. The transfection efficiency is associated with the amount of plasmid DNA injection, injection time, injection speed, and volume. Our studies suggest that transplacental RNAi could be implemented by tail vein injection of plasmid vector into pregnant mice.

Comparative Genomics of Strictly Vertically Transmitted, Feminizing Microsporidia Endosymbionts of Amphipod Crustaceans.

Microsporidia are obligate intracellular eukaryotic parasites of vertebrates and invertebrates. Microsporidia are usually pathogenic and undergo horizontal transmission or a mix of horizontal and vertical transmission. However, cases of nonpathogenic microsporidia, strictly vertically transmitted from mother to offspring, have been reported in amphipod crustaceans. Some of them further evolved the ability to feminize their nontransmitting male hosts into transmitting females. However, our understanding of the evolution of feminization in microsporidia is hindered by a lack of genomic resources. We report the sequencing and analysis of three strictly vertically transmitted microsporidia species for which feminization induction has been demonstrated (Nosema granulosis) or is strongly suspected (Dictyocoela muelleri and Dictyocoela roeselum), along with a draft genome assembly of their host Gammarus roeselii. Contrary to horizontally transmitted microsporidia that form environmental spores that can be purified, feminizing microsporidia cannot be easily isolated from their host cells. Therefore, we cosequenced symbiont and host genomic DNA and devised a computational strategy to obtain genome assemblies for the different partners. Genomic comparison with feminizing Wolbachia bacterial endosymbionts of isopod crustaceans indicated independent evolution of feminization in microsporidia and Wolbachia at the molecular genetic level. Feminization thus represents a remarkable evolutionary convergence of eukaryotic and prokaryotic microorganisms. Furthermore, a comparative genomics analysis of microsporidia allowed us to identify several candidate genes for feminization, involving functions such as DNA binding and membrane fusion. The genomic resources we generated contribute to establish Gammarus roeselii and its microsporidia symbionts as a new model to study the evolution of symbiont-mediated feminization.

Effects of 2,4-dichlorophenol exposure on zebrafish: Implications for the sex hormone synthesis.

2,4-Dichlorophenol (2,4-DCP), an estrogenic endocrine disruptor, is widely spread in aquatic environments and may interfere with normal physiological functions in fish. However, the influence of this chemical on the synthesis of sex hormones is not well understood. In the present study, zebrafish (Danio rerio) were exposed to 2,4-DCP (80 and 160 µg/L) with or without fadrozole (an aromatase inhibitor which inhibits the synthesis of estradiol) from 20 to 40 days post fertilization. Then, the sex ratio, the content of vitellogenin (VTG) and sex hormones (androstenedione (ASD), estrone (E1), 17ß-estradiol (E2), estriol (E3), testosterone (T) and 11-ketotestosterone (11-KT)) were studied. Furthermore, the expression of genes involved in synthesis of sex hormones (cyp19a1a, cyp19a1b, 17ß-hsd, 11ß-hsd and cyp11b) along with the DNA methylation in cyp19a1a and cyp19a1b promoters was analyzed. The results showed that 2,4-DCP exposure led to female-biased ratio, increased the content of ASD, E2 and VTG, as well as the ratio of E2/11-KT, while decreased the levels of androgens (T and 11-KT). The sex hormonal change can be explained by the significant up-regulation of cyp19a1a, cyp19a1b, 17ß-hsd and 11ß-hsd genes. In addition, hypomethylation of cyp19a1a promoter was involved in this process. Notably, fadrozole can partly attenuate 2,4-DCP-induced feminization, and recover the levels of ASD, E2 and 11-KT. Thus, these results demonstrate that 2,4-DCP induces feminization in fish by disrupting the synthesis of sex hormones.

Searching for sex-reversals to explain population demography and the evolution of sex chromosomes.

Sex determination can be purely genetic (as in mammals and birds), purely environmental (as in many reptiles), or genetic but reversible by environmental factors during a sensitive period in life, as in many fish and amphibians (Wallace et al. 1999; Baroiller et al. 2009a; Stelkens & Wedekind 2010). Such environmental sex reversal (ESR) can be induced, for example, by temperature changes or by exposure to hormone-active substances. ESR has long been recognized as a means to produce more profitable single-sex cultures in fish farms (Cnaani & Levavi-Sivan 2009), but we know very little about its prevalence in the wild. Obviously, induced feminization or masculinization may immediately distort population sex ratios, and distorted sex ratios are indeed reported from some amphibian and fish populations (Olsen et al. 2006; Alho et al. 2008; Brykov et al. 2008). However, sex ratios can also be skewed by, for example, segregation distorters or sex-specific mortality. Demonstrating ESR in the wild therefore requires the identification of sex-linked genetic markers (in the absence of heteromorphic sex chromosomes) followed by comparison of genotypes and phenotypes, or experimental crosses with individuals who seem sex reversed, followed by sexing of offspring after rearing under non-ESR conditions and at low mortality. In this issue, Alho et al. (2010) investigate the role of ESR in the common frog (Rana temporaria) and a population that has a distorted adult sex ratio. They developed new sex-linked microsatellite markers and tested wild-caught male and female adults for potential mismatches between phenotype and genotype. They found a significant proportion of phenotypic males with a female genotype. This suggests environmental masculinization, here with a prevalence of 9%. The authors then tested whether XX males naturally reproduce with XX females. They collected egg clutches and found that some had indeed a primary sex ratio of 100% daughters. Other clutches seemed to result from multi-male fertilizations of which at least one male had the female genotype. These results suggest that sex-reversed individuals affect the sex ratio in the following generation. But how relevant is ESR if its prevalence is rather low, and what are the implications of successful reproduction of sex-reversed individuals in the wild?

Genetic feminization of the thoracic nervous system disrupts courtship song in male Drosophila melanogaster.

Despite the growing research investigating the sex-specific organization of courtship behavior in Drosophila melanogaster, much remains to be understood about the sex-specific organization of the motor circuit that drives this behavior. To investigate the sex-specification of a tightly patterned component of courtship behavior, courtship song, the authors used the GAL4/UAS targeted gene expression system to feminize the ventral ganglia in male Drosophila and analyzed the acoustic properties of courtship song. More specifically, the authors used the thoracic-specifying teashirt promoter (tsh(GAL4)) to express feminizing transgenes specifically in the ventral ganglia. When tsh(GAL4) drove expression of transformer (tra), males were unable to produce prolonged wing extensions. Transgenic expression of an RNAi construct directed against male-specific fruitless (fru(M)) transcripts resulted in normal wing extension, but highly defective courtship song, with 58% of males failing to generate detectable courtship song. Of those that did sing, widths of individual pulses were significantly broader than controls, suggesting thoracic fru(M) function serves to mediate proprioceptive-dependent wing vibration damping during pulse song. However, the most critical signal in the song, the interpulse interval, remained intact. The inability to phenocopy this effect by reducing fru(M) expression in motor neurons and proprioceptive neurons suggests thoracic interneurons require fru(M) for proper pulse song execution and patterning of pulse structure, but not for pulse timing. This provides evidence that genes establishing sex-specific activation of complex behaviors may also be used in establishing pattern-generating motor networks underlying these sex-specific behaviors.

Profiling spermatogenic failure in adult testes bearing Sox9-deficient Sertoli cells identifies genes involved in feminization, inflammation and stress.

BACKGROUND: Sox9 (Sry box containing gene 9) is a DNA-binding transcription factor involved in chondrocyte development and sex determination. The protein's absence in testicular Sertoli nurse cells has been shown to disrupt testicular function in adults but little is known at the genome-wide level about molecular events concomitant with testicular break-down. METHODS: To determine the genome-wide effect on mRNA concentrations triggered by the absence of Sox9 in Sertoli cells we analysed adult testicular tissue from wild-type versus mutant mice with high-density oligonucleotide microarrays and integrated the output of this experiment with regulatory motif predictions and protein-protein network data. RESULTS: We report the genome-wide mRNA signature of adult testes lacking Sox9 in Sertoli cells before and after the onset of late spermatogenic failure as compared to fertile controls. The GeneChip data integrated with evolutionarily conserved Sox9 DNA binding motifs and regulatory network data identified genes involved in feminization, stress response and inflammation. CONCLUSIONS: Our results extend previous observations that genes required for female gonadogenesis are up-regulated in the absence of Sox9 in fetal Sertoli cells to the adult stage. Importantly, we identify gene networks involved in immunological processes and stress response which is reminiscent of a phenomenon occurring in a sub-group of infertile men. This suggests mice lacking Sox9 in their Sertoli cells to be a potentially useful model for adult human testicular failure.

Studies on feminization, sex determination, and differentiation of the Southern catfish, Silurus meridionalis--a review.

The sex ratio of the feral Southern catfish was reported to be about 1:1, while the fish obtained by artificial fertilization were always female. Hence, we examined the possible influence of the micro-environment during artificial insemination (pH of the ovarian fluid and concentration of the semen) and early development (feed, hatching temperature, and water) on the sex ratio of Southern catfish fry. In order to examine the possibility of the occurrence of gynogenesis during artificial propagation, cytological observations on the insemination processes and the artificial induction of gynogenesis were also performed. However, no male fish were obtained even in these experiments, excluding the possibilities of these micro-environmental changes on catfish sex ratio and the occurrence of gynogenesis during artificial propagation. Female-to-male sex reversal was achieved by treatment with fadrozole (an aromatase inhibitor) and tamoxifen (an estrogen receptor antagonist). Histological analyses on the gonadal development of both female and induced male fish were subsequently performed. Moreover, several genes involved in sex differentiation, such as dmrt1, foxl2, and cyp19, and three subunits of gonadotropin (gth), i.e., gthalpha, lhbeta, and fshbeta, were isolated. Their expression patterns were studied under normal gonadal development and sex reversal conditions. The results revealed that dmrt1, foxl2, and cyp19a were closely related to catfish sex differentiation, and the gth subunits were possibly related to ovarian differentiation and oocyte development. Taken together, we hypothesized that estrogen was highly responsible for the ovarian differentiation and feminization of catfish fry under artificial propagation, although the mechanism involved remains elusive.

AI- modelling of molecular identification and feminization of wolbachia infected Aedes aegypti.

BACKGROUND: The genetic control strategies of vector borne diseases includes the replacement of a vector population by "disease-refractory" mosquitoes and the release of mosquitoes with a gene to control the vector's reproduction rates. Wolbachia are common intracellular bacteria that are found in arthropods and nematodes. Wolbachia infected male mosquitos have been used in different experimental trials around the world to suppress the target population of Aedes aegypti and this genetic control strategy has proved to be a promising alternative to other treatment strategies. Due to certain limitations, the successful application of this strategy is still awaited. METHODS: Mathematical frame work for Wolbachia induced genetic control strategy has been developed in this article. With the aid of Artificial Intelligence (AI) tools, accurate parametric values are depicted. For the first time, the model is well synchronized with the experimental findings. The model is comprised of the generalized varying coefficient and multiple mating rates between infected and uninfected compartments of Aedes aegypti dengue to forecast the disease control. RESULTS: Two mathematical models are developed in this article to demonstrate different mating rates of the genetic control strategy. The important parameters and time varying coefficients are well demonstrated with the aid of numerical computations. The resulting thresholds and forecasting may prove to be a useful tool for future experimental studies. CONCLUSIONS: From our analysis, we have concluded that the genetic control strategy is a promising technique and the role of Wolbachia infected male mosquitos, in genetic control strategies, can be better interpreted in an inexpensive manner with the aid of a theoretical model.

teiresias, a Fruitless target gene encoding an immunoglobulin-superfamily transmembrane protein, is required for neuronal feminization in Drosophila.

This study aims at identifying transcriptional targets of FruitlessBM (FruBM), which represents the major isoform of male-specific FruM transcription factors that induce neural sexual dimorphisms. A promoter of the axon-guidance factor gene robo1 carries the 16-bp palindrome motif Pal1, to which FruM binds. Our genome-wide search for Pal1-homologous sequences yielded ~200 candidate genes. Among these, CG17716 potentially encodes a transmembrane protein with extracellular immunoglobulin (Ig)-like domains similar to Robo1. Indeed, FruBM overexpression reduced CG17716 mRNA and protein expression. In the fru-expressing mAL neuron cluster exhibiting sexual dimorphism, we found that CG17716 knockdown in female neurons completely transformed all neurites to the male-type. Conversely, CG17716 overexpression suppressed male-specific midline crossing of fru-expressing sensory axons. We renamed CG17716 teiresias (tei) based on this feminizing function. We hypothesize that Tei interacts with other Ig superfamily transmembrane proteins, including Robo1, to feminize the neurite patterns in females, whereas FruBM represses tei transcription in males.