Expression of sex steroid hormone-related genes in the embryo of the leopard gecko (original) (raw)
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Comparative Biochemistry and Physiology B-biochemistry & Molecular Biology, 2003
Sex steroid hormones play a central role in the reproduction of all vertebrates. These hormones function through their specific receptors, so the expression levels of the receptors may reflect the responsibility of target organs. However, there was no effective method to quantify the expression levels of these receptors in reptilian species. In this study, we established the competitive-PCR assay systems for the quantification of the mRNA expression levels of three sex steroid hormone receptors in the leopard gecko. These assay systems were successfully able to detect the mRNA expression level of each receptor in various organs of male adult leopard geckoes. The expression levels of mRNA of these receptors were highly various depending on the organs assayed. This is the first report regarding the tissue distributions of sex steroid hormone receptor expressions in reptile. The effects of environmental conditions on these hormone receptor expressions were also examined. After the low temperature and short photoperiod treatment for 6 weeks, only the androgen receptor expression was significantly increased in the testes. The competitive-PCR assay systems established in this report should be applicable for various studies of the molecular mechanism underlying the reproductive activity of the leopard gecko.
Estrogen signalling is critical for ovarian differentiation in reptiles with temperature-dependent sex determination (TSD). To elucidate the involvement of estrogen in this process, adrenal–kidney–gonadal (AKG) expression of estrogen receptor (ERa) was studied at female-producing temperature (FPT) in the developing embryos of the lizard, Calotes versicolor which exhibits a distinct pattern of TSD. The eggs of this lizard were incubated at 31.5 ± 0.5 °C (100% FPT). The torso of embryos containing adrenal– kidney–gonadal complex (AKG) was collected during different stages of development and subjected to Western blotting and immunohistochemistry analysis. The ERa antibody recognized two protein bands with apparent molecular weight 55and55 and 55and45 kDa in the total protein extracts of embryonic AKG complex of C. versicolor. The observed results suggest the occurrence of isoforms of ERa. The differential expression of two different protein isoforms may reveal their distinct role in cell proliferation during gonadal differentiation. This is the first report to reveal two isoforms of the ERa in a reptile during development. Immunohistochemical studies reveal a weak, but specific, cytoplasmic ERa immunostaining exclusively in the AKG during late thermo-sensitive period suggesting the responsiveness of AKG to estrogens before gonadal differentiation at FPT. Further, cytoplasmic as well as nuclear expression of ERa in the medulla and in oogonia of the cortex (faint activity) at gonadal differentiation stage suggests that the onset of gonadal estrogen activity coincides with sexual differentiation of gonad. Intensity and pattern of the immunoreactions of ERa in the medullary region at FPT suggest endogenous production of estrogen which may act in a paracrine fashion to induce neighboring cells into ovarian differentiation pathway.
Sex steroid hormones play an important role in mediating physiological responses and developmental processes through their receptors across all vertebrates. Chinese alligator (Alligator sinensis) is a critically endangered reptile species unique to China. In this study, we have cloned one of the sex steroid hormone receptor genes, androgen receptor (AR) from the brain of Chinese alligator for the first time. The full-length AR cDNA is 2717 bp in length with an open reading frame (ORF) encoding 722 amino acids. Amino acid alignment analyses indicated that the ARs exhibit highly conserved functional domains. Especially, the P-box and D-box, which are essential to ensure that receptor binding to the androgen response elements, are completely conserved in selected species. Using the quantitative real-time PCR (qPCR), the spatial expression of four receptor mRNAs in all newborn brain tissues and temporal expression of them in the cerebrum during the embryonic development in Chinese alligators were investigated. The results of qPCR showed ubiquitous expression of the four receptor mRNAs in all newborn brain tissues examined and significant changes in the expression levels of these receptor mRNAs in the embryonic development. These results suggest that sex steroid hormones might play an important role in the regulation of complex neuroendocrine activities in newborn Chinese alligator. Furthermore, these data provide an important foundation for further studies on endocrinology and molecular biology of non-mammalian sex steroid hormone receptors.
The Journal of Experimental Zoology, 1998
Sex determination in egg-laying amniotes may be fundamentally different from that of placental mammals. The mammalian ovary differentiates normally in the absence of estrogen, whereas estrogen seems to be crucial for proper ovarian development in birds, reptiles, and lower vertebrates. Estrogens are produced normally by the biosynthetic conversion of androgens by the enzyme aromatase (CYP19), which is the sole mediator of this reaction. Aromatase inhibitors are capable of reversing females to males in turtles and chickens; therefore, a role for aromatase as the female sex determinant has been postulated for species in which sex determination is temperature-dependent. The entire aromatase coding sequence (1,509 base pairs) from adult terrapin ovaries was cloned, and Northern analysis indicates a single transcript (2.4 kb) for adult ovaries, whereas male and female brains express a 2.4-kb as well as a 9.6-kb transcript. Using a sensitive (attomole sensitivity) competitive RT-PCR technique, aromatase transcript abundance was quantified during embryonic development for embryos treated with and without estrogen. Aromatase is transcribed, well before the temperature-sensitive, (stage 12), at both male and female temperatures in the brain. There is a switch to lower aromatase transcript abundance in the female brain concurrent with an exponential rise of aromatase transcript in the putative ovary. Transcripts remain below the detection limits in the putative testes but exhibit female levels of aromatase transcript when treated with estrogen. Aromatase mRNA levels are generally reduced in the brain by estradiol application. On the basis of these findings, we have postulated a model based on the competition between 5α-reductase and P450 aromatase for androgen substrate in both the brain and the undifferentiated gonad to explain the TSD phenomenon in reptiles.
Journal of Endocrinology and Reproduction, 2020
Steroidal sex hormones play critical rolest during gonadal sex differentiation and development. Among the sex steroids, estradiol acts at the central level in the sexual differentiation of many reptilian species exhibiting temperature-dependent sex determination (TSD). In recent years, the growing amount of information has highlighted the importance of aromatase (CYP19) in irreversibly catalyzing conversion of androgens into estrogens by aromatization during ovarian morphogenesis. To elucidate the involvement of aromatase in this process we investigated the ontogenic and cellular expression of aromatase in the embryos of the lizard Calotes versicolor that exhibits a unique pattern of TSD. The eggs of this lizard were incubated at 31.5±0.5°C which is 100% Female-Producing Temperature (FPT). The torso of embryos containing Adrenal- Kidney-Gonadal (AKG) complex was collected during different stages of development and subjected to Western blotting and immunohistochemical analysis. Weste...
Sex steroid receptors in the ovarian follicles of the lizard Sceloporus torquatus
Zygote
SummaryEstradiol and progesterone have been recognized as important mediators of reproductive events in the female mainly via binding to their receptors. This study aimed to characterize the immunolocalization of the estrogen receptor alfa (ERα), estrogen receptor beta (ERβ) and progesterone receptor (PR) in the ovarian follicles of the lizard Sceloporus torquatus. The localization of steroid receptors has a spatio-temporal pattern that depends on the stage of follicular development. The immunostaining intensity of the three receptors was high in the pyriform cells and the cortex of the oocyte of previtellogenic follicles. During the vitellogenic phase, the granulosa and theca immunostaining was intense even with the modification of the follicular layer. In the preovulatory follicles, the receptors were found in yolk and additionally, ERα was also located in the theca. These observations suggest a role for sex steroids in regulating follicular development in lizards, like other vert...
General and Comparative Endocrinology, 2003
Successful reproduction requires the coordination of reproductive physiology with behavior. The neural correlates of reproductive behavior have been elucidated in a variety of amphibians, mammals, and birds but relatively few studies have examined reptiles. Here we investigate differences in androgen receptor (AR) mRNA expression in the forebrain and oviduct between previtellogenic and late vitellogenic female leopard geckos, Eublepharis macularius. Plasma concentrations of testosterone (T) are low when females are previtellogenic and sexually unreceptive but increase dramatically during late vitellogenesis when females are receptive. In addition, receptivity can be induced by treatment with exogenous T. The relative abundance of AR-mRNA across various nuclei was greater in late vitellogenic than in previtellogenic females. This general pattern was observed in the medial preoptic area, anterior hypothalamus, external nucleus of the amygdala, dorsolateral aspect of the ventromedial hypothalamus, lateral septum, and periventricular hypothalamus. There were also clear differences in AR-mRNA expression among these nuclei. The pattern of gene expression observed in the brain was reversed within stromal cells of the oviduct where expression of AR-mRNA decreased from the previtellogenic stage to the late vitellogenic stage. Overall, these data demonstrate that T concentration in the plasma, abundance of AR-mRNA in the brain and oviduct, and sexual behavior change coordinately during the reproductive cycle of female leopard geckos. Although the function of AR in the female leopard gecko is not yet clear, our results are in accord with growing evidence that androgens regulate numerous aspects of female physiology and behavior in vertebrates.
General and Comparative Endocrinology, 2005
Incubation temperature during embryonic development determines gonadal sex in the leopard gecko (Eublepharis macularius). Incubation temperature and gonadal sex jointly inXuence the display of sexual and agonistic behavior in adult leopard geckos. These diVerences in adult behavior are organized prior to sexual maturity, and it is plausible that post-natal hormones inXuence neural and behavioral diVerentiation. Here we assessed incubation temperature and sex eVects on sex steroid levels in leopard geckos at 2, 10, and 25 weeks of age and monitored the development of male secondary sex structures. Males had signiWcantly higher androgen concentrations at all time points, whereas females had signiWcantly higher 17-estradiol (E2) concentrations only at 10 and 25 weeks. Within males, age but not incubation temperature aVected steroid levels and morphological development. Male androgen levels increased modestly by 10 and dramatically by 25 weeks of age, whereas E2 levels remained unchanged over this period. Most males had signs of hemipenes at 10 weeks of age, and all males had hemipenes and open preanal pores by 25 weeks of age. In females, age and incubation temperature aVected E2 and dihydrotestosterone (DHT) but not T concentrations. Controlling for age, females from 34°C have higher DHT and lower E2 levels than females from 30°C. Further, E2 concentrations increased signiWcantly from 2 to 10 weeks, after which E2 levels remained steady. Together, these results indicate that sexually dimorphic levels of steroids play a major role in the development of leopard gecko behavior and morphology. Furthermore, these data suggest that the organizational eVects of incubation temperature on adult female phenotype could be, in part, mediated by incubation temperature eVects on steroid hormone levels during juvenile development.
General and Comparative Endocrinology, 2003
We examined differences in gonadal steroid production and biosynthetic pathway activity with changing reproductive condition and between sexes in the scincid lizard, Tiliqua nigrolutea. We observed clear seasonal and sexual variation in the production of androgens and steroid conjugates, but detected no 17b-estradiol or 5a-dihydrotestosterone produced by the gonads. An alternative steroid, more polar than estradiol, was detected: an investigation of this steroid is reported separately [Gen. Comp. Endocrinol. 129 ]. There were seasonal and sex-related differences in steroid biosynthetic pathway activity. The D5 pathway metabolite, dehydroepiandrosterone, was detected only in males, and only from incubations using regressed testicular tissue. There was also a seasonal difference between the sexes in rates of progesterone accumulation, although the absence of corresponding elevated plasma concentrations suggests that the role of progesterone switches from a directly acting hormone to a precursor for others during the reproductive cycle in females. These results suggest that within the traditional view that vertebrate biosynthetic pathway activity and end-products are phylogenetically conserved, there is likely to be considerably species-and/or genus-specific variation.