RESUMEN

Kisspeptin plays a vital role in mediating the stress-induced reproductive regulation. Cortisol, known as a stress-related hormone, is involved in gonadal development and sexual differentiation by binding with glucocorticoid receptor (GR) to regulate the expression of kiss gene. In the present study, cortisol treatment in yellowtail clownfish (Amphiprion clarkii) showed that the expression of kiss (kiss1 and kiss2) and gr (gr1 and gr2) genes were increased significantly. We demonstrated that the yellowtail clownfish Kiss neurons co-express the glucocorticoid receptors in the telencephalon, mesencephalon, cerebellum, and hypothalamus. We further cloned the promoter of kiss2 gene in yellowtail clownfish and identified the presence of putative binding sites for glucocorticoid receptors, estrogen receptors, androgen receptors, progesterone receptors, AP1, and C/EBP. Applying transient transfection in HEK293T cells of the yellowtail clownfish kiss2 promoter, cortisol (dexamethasone) treatment was shown to enhance the promoter activities of the yellowtail clownfish kiss2 gene in the presence of GRs. Deletion analysis of kiss2 promoter indicated that cortisol-induced promoter activities were located between position -660 and -433 with GR1, and -912 and -775 with GR2, respectively. Finally, point mutation studies on the kiss2 promoter showed that cortisol-stimulated promoter activity was mediated by one GRE site located at position -573 in the presence of GR1 and by each GRE site located at position -883, -860, -851, and -843 in the presence of GR2. Results of the present study provide novel evidence that cortisol could regulate the transcription of kiss2 gene in the yellowtail clownfish via GRE-dependent GR pathway.

Asunto(s)

Perciformes , Receptores de Glucocorticoides , Animales , Células HEK293 , Humanos , Hidrocortisona/metabolismo , Hidrocortisona/farmacología , Perciformes/genética , Regiones Promotoras Genéticas , Receptores de Glucocorticoides/genética

RESUMEN

Kisspeptin system was shown to be a key factor in mediating social stress and reproduction. Yellowtail clownfish, Amphiprion clarkii, is a hermaphrodite fish, whose sex determination and gonadal development are affected by the social status of individuals. The yellowtail clownfish is a fantastic animal model to explore sex determination, but the social status and precise distribution of kiss mRNAs in the brain of this species are unknown. Hererin, a novel in situ hybridization technique, RNAscope, was used to investigate the distribution of kiss1 and kiss2 expressions in the brain of yellowtail clownfish. The coronal planes of brain showed that the kiss1 signal was mainly present in dorsal habenular nucleus (NHd) and kiss2 mRNA was widely expressed in telencephalon, midbrain, and hypothalamus, especially in dorsal part of the nucleus of the lateral recess (NRLd). Additionally, kiss1 and kiss2 signals have sexually dimorphic distribution. The kiss1 mRNA was distributed in NHd, the telencephalon, and lateral part of the diffuse nucleus of the inferior lobe (NDLIl) of females but in NHd and NDLIl of males. kiss2 signals were stronger in females than that in males. The distribution of kiss1 and kiss2 neurons in NHd of habenula and NRLd of hypothalamus may suggest that kiss genes associate environmental signaling and reproductive function in yellowtail clownfish.

RESUMEN

The khapra beetle, Trogoderma granarium Everts, is defined as one of the most important quarantine pests globally, and fumigation with methyl bromide, an ozone-depleting substance, is a common phytosanitary measure currently used. The modified atmosphere (MA), irradiation, and their combination treatments of T. granarium larvae and adults were performed at room temperature (24-26 ℃) to develop an ecofriendly phytosanitary disinfestation measure and to shorten the exposure time and overcome treatment disadvantages of irradiation. Late-stage larvae are determined as the most tolerant stage resulted in large LT99.9968 values of 32.6 (29.2-37.5) and 38.0 (35.1-41.7) days treated under 1% and 2% O2 (with N2 balance) atmosphere, respectively. Ionizing radiation was used to enhance the effect of MA and the mortality was highly significantly affected by all the interaction effects, indicating that the synergistic effects present in all the combined treatments. The synergistic ratios, which is defined as the estimated lethal time for MA treatment (LD90, LD99, and LD99.9968), divided by that of combined treatment, were between 1.47 and 2.47. In the confirmatory tests, no individuals recovered from a sum of 111,366 late-stage larvae treated under 1% O2 atmosphere for 14- or 15-d after 200 Gy irradiation, which resulted in validating the probit estimations and achieving an efficacy of 99.9973% mortality at 95% confidence level. Therefore, these treatment schedules are recommended to disinfest T. granarium infecting commodities for phytosanitary purposes under the warehouse, MA packaging, or in combination with international transportation by train or sea container.