Fertility differences between two wild-type Drosophila melanogaster lines correlate with differences in the expression of the Jheh1 gene, which codes for an enzyme degrading juvenile hormone.

Juvenile hormone plays a "status quo" role in Drosophila melanogaster larvae, preventing the untimely metamorphosis, and performs a gonadotropic function in imagoes, ensuring the ovaries' preparedness for vitellogenesis. The decreased level of juvenile hormone results in reproductive disorders in D. melanogaster females including a delay in the oviposition onset and a fertility decrease. Another factor that can affect the insect reproduction is an infection with the maternally inherited symbiotic α-proteobacterium Wolbachia. The present study is devoted to the analysis of the expression of two juvenile hormone metabolism genes encoding enzymes of its synthesis and degradation, juvenile hormone acid O-methyltransferase ( jhamt) and juvenile hormone epoxide hydrase (Jheh1), respectively, in four wild-type D. melanogaster lines, two of them being infected with Wolbachia. Lines w153 and Bi90 were both derived from an individual wild-caught females infected with Wolbachia, while lines w153T and Bi90T were derived from them by tetracycline treatment and are free of infection. Line Bi90 is known to be infected with the Wolbachia strain wMel, and line w153, with the Wolbachia strain wMelPlus belonging to the wMelCS genotype. It was found that infection with either Wolbachia strain does not affect the expression of the studied genes. At the same time, it was shown that the w153 and w153T lines differ from the Bi90 and Bi90T lines by an increased level of the Jheh1 gene expression and do not differ in the jhamt gene expression level. Analysis of the fertility of these four lines showed that it does not depend on Wolbachia infection either, but differs between lines with different nuclear genotypes: in w153 and w153T, it is significantly lower than in lines Bi90 and Bi90T. The data obtained allow us to reasonably propose that the inter-line D. melanogaster polymorphism in the metabolism of the juvenile hormone is determined by its degradation (not by its synthesis) and correlates with the fertility level.

Insulin-like peptide DILP6 regulates juvenile hormone and dopamine metabolism in Drosophila females.

Insulin-like peptide DILP6 is a component of the insulin/insulin-like growth factor signalling pathway of Drosophila. Juvenile hormone (JH) and dopamine (DA) are involved in the stress response and in the control of reproduction. In this study, we investigate whether DILP6 regulates the JH and DA levels by studying the effect of a strong hypomorphic mutation dilp641 on JH and DA metabolism in D. melanogaster females. We show that DILP6 regulates JH and DA metabolism: the mutation dilp641 results in a reduction in JH-hydrolysing activity and an increase in the activities of DA synthesis enzymes (alkaline phosphatase (ALP) and tyrosine hydroxylase (TH)). In the mutant females, we also found increased fecundity in addition to the intensity of the response (stress reactivity) of ALP and TH to heat stress. As we showed previously, this suggests an increased level of JH synthesis. We confirm this suggestion by treating the mutant females with the JH inhibitor, precocene, which restors the activity and stress reactivity of ALP and TH as well as fecundity to levels similar to those in the control flies. The data suggest a feedback system in the interaction between JH and DILP6 in which DILP6 negatively regulates the JH titre via an increase in the hormone degradation and a decrease in its synthesis.

[Gene dilp6 regulates octopamine metabolism in Drosophila melanogaster].

The effect of strong hypomorphic mutation of the insulin-like protein gene (dilp6) on metabolism of octopamine (one of the main biogenic amines in insects) was studied in Drosophila melanogaster males and females. The activity of tyrosine decarboxylase (the key enzyme of octopamine synthesis) and the activity of octopamine-dependent N-acetyltransferase (the enzyme of its degradation) were measured. It was demonstrated that the activity of both studied enzymes is decreased under normal conditions in the dilp6 41 mutants (as we previously demonstrated, this is correlated with an increased level of octopamine). It was also found that hypomorphic mutation of the dilp6 gene decreases the intensity of tyrosine decarboxylase response to heat stress. Thus, it was demonstrated for the first time that insulin-like DILP6 protein in drosophila influences the level of octopamine (regulating the activity of the enzyme degrading octopamine).

The insulin-like receptor gene expression in the tissues synthesizing gonadotropic hormones at sexual maturation of Drosophila melanogaster females].

The insulin/insulin-like growth factor signaling pathway is involved in the regulation of the synthesis of insect gonadotropic hormones, juvenile (JH) and 20-hydroxyecdysone (20E). We carried out the immunohistochemical analysis of the insulin receptor (InR) expression in the corpus allatum (the JH-producing gland) and in the ovarian follicular cells (a site for the synthesis of 20E precursor, ecdysone) in the process of sexual maturation of D. melanogaster females and examined the influence of exogenous JH on the InR expression in these tissues. For the first time, it was demonstrated that InR was expressed in follicular cells and that its expression in corpus allatum and follicular cells of Drosophila females was stage-specific, i.e., the expression intensity in young females greatly exceeded that in mature individuals. We also found a negative feedback loop in the regulation of JH levels by the insulin signaling pathway in Drosophila adults: the experimental increase in the JH titers in young females dramatically reduced the InR expression intensity in corpus allatum and follicular cells.

[InR gene expression and octopamine metabolism in Drosophila melanogaster females].

The influence of suppression of the expression of the Drosophila insulin-like receptor gene (InR) in corpus allatum (the gland-synthesizing juvenile hormone) on octopamine and juvenile hormone metabolism and on the development of the stress-reaction in Drosophila melanogaster females was studied. It was demonstrated that the suppression of InR gene expression in corpus allatum induces in D. melanogaster females an increase in the activity of the enzyme that limits the rate of octopamine synthesis (tyrosine decarboxylase), as well as in the level of juvenile hormone degradation and the intensity of the response of octopamine and juvenile hormone metabolism systems to heat stress. It was mentioned that a decrease in InR gene expression in corpus allatum does not influence the activity of OA-dependent N-acetyltransferase (the enzyme that degrades octopamine). It was established that the influence of suppression of the InR gene expression in corpus allatum on octopamine metabolism is mediated by juvenile hormone, since the processing of flies by exogenous juvenile hormone restores the activity oftyrosine decarboxylase in flies with decreased InR expression in corpus allatum up to the normal level.

Decrease in juvenile hormone level as a result of genetic ablation of the corpus allatum cells affects the synthesis and metabolism of stress related hormones in Drosophila.

Previous studies have shown that juvenile hormone (JH) regulates dopamine (DA) and octopamine (OA) content in Drosophila, and we have shown the influence of an increase in JH level on DA and OA metabolism in young females of Drosophila virilis and Drosophila melanogaster. Here we investigate the effects of genetic ablation of a subset of cells in the Corpusallatum (CA, endocrine gland synthesizing JH) on the DA levels and activities of alkaline phosphatase (ALP), tyrosine hydroxylase (TH), DA-dependent arylalkylamine N-acetyltransferase (DAT) and tyrosine decarboxylase (TDC) in young D. melanogaster females under normal conditions and upon heat stress (38°Ð¡). We show that ablation of СА cells causes: (1) a decrease in ALP, TH and DAT activities, (2) an increase in DA level and (3) an increase in TDC activity in young females. The CA ablation was also found to modulate ALP, TH and TDC responses to heat stress. Mechanisms of regulation of DA and OA levels by JH in Drosophila females are discussed.

Mechanisms of age-specific regulation of dopamine metabolism by juvenile hormone and 20-hydroxyecdysone in Drosophila females.

20-hydroxyecdysone (20E) and the juvenile hormone (JH) have an age-specific effect on total dopamine (DA) content in Drosophila (Gruntenko and Rauschenbach 2008). Earlier we studied the mechanism of influence of 20E and JH on DA metabolism in young females (Rauschenbach et al. in J Insect Physiol 53:587-591, 2007a: Arch Insect Biochem Physiol 65:95-102, 2008a; Gruntenko et al. in Arch Insect Biochem Physiol 72:263-269, 2009). Here we investigate the effects of 20E and JH on the activities of the alkaline phosphatase (ALP), tyrosine hydroxylase (TH) and DA-dependent arylalkylamine N-acetyltransferase (AANAT) in mature females of wild type D. virilis under normal conditions and under heat stress (38°C). 20E feeding of the flies led to a substantial decrease in ALP and TH activities and to an increase in AANAT activity in mature females. JH application resulted in an increasing of ALP and TH activities, but did not influence AANAT activity in mature females. A rise in JH and 20E levels was found to change ALP and TH stress reactivities. Mechanisms of age-specific regulation of DA level by 20E and JH in Drosophila females are discussed.

Altered juvenile hormone metabolism, reproduction and stress response in Drosophila adults with genetic ablation of the corpus allatum cells.

Juvenile hormone (JH), which controls many developmental and physiological processes in Drosophila melanogaster, is synthesized de novo in the specialized endocrine glands, corpus allatum (CA). The present study concerns JH metabolism, reproduction and stress resistance in Drosophila with genetic ablation of a part of CA cells. The correlated regulation of JH biosynthesis and degradation in Drosophila adults has been found: ablation of CA cells led to (1) a dramatic decrease in activity of the key regulatory enzyme of JH biosynthesis, juvenile hormone acid methyl transferase and (2) a considerable increase in JH-hydrolyzing activity. It has been also shown that ablation of CA cells caused three significant physiological changes: (1) an increase in the intensity of response of JH degradation system to heat stress; (2) a disturbance of reproduction; (3) a decrease in stress resistance. Pharmacological rise of JH level rescued JH-hydrolyzing activity, fecundity and stress resistance in CA-ablated females. Pronouncedly, all the physiological effects caused by CA ablation were significant in females but not in males indicating a sexual dimorphism of JH physiological roles in Drosophila adults.

Dopamine down-regulates activity of alkaline phosphatase in Drosophila: the role of D2-like receptors.

The effect of a rise in dopamine (DA) level as a result of a mutation, stress or pharmacological treatment on the activity of the enzyme of its synthesis, alkaline phosphatase (ALP) in females of Drosophila virilis and Drosophila melanogaster has been studied. It has been found that regardless of its nature, a rise in DA level has a negative effect on ALP activity, which indicates that DA down-regulates activity of the enzyme. The effects of bromocriptine (an agonist of Drosophila dopamine 2-like receptor (DD2R)) on ALP activity have been studied. ALP activity was found to drop in response to bromocriptine in flies. Conversely ALP activity was increased in flies with reduced DD2R expression (i.e. Actin5C-Gal4>UAS-ds-DD2R RNA-interference flies) vs. corresponding controls (i.e. Actin5C-Gal4>w1118 flies). Bromocriptine treatment of RNAi flies rescues ALP activity to the level typical of Actin5C-Gal4>w1118 flies. A change in DD2R number or availability was found not to prevent the response of ALP to heat stress, but to change the intensity of its response to the stress exposure. The role of D2-like receptors in down-regulation of ALP activity by DA and in ALP response to stressor in Drosophila is discussed.

20-hydroxyecdysone and juvenile hormone influence tyrosine hydroxylase activity in Drosophila females under normal and heat stress conditions.

The effects of exogenous 20-hydroxyecdysone (20E) and the juvenile hormone (JH) on the activity of the tyrosine hydroxylase (TH), the first and rate-limiting DA biosynthetic enzyme, has been studied in young females of wild type D. virilis and D. melanogaster under normal conditions and under heat stress (38 degrees C). Both 20E feeding of the flies and JH application led to a substantial rise in TH activity. A rise in JH and 20E levels was found not to prevent the response of TH to heat stress, but to change the intensity of its response to the stress exposure. Putative mechanisms of regulation of DA level by 20E and JH in Drosophila females are discussed.

Effects of 20-hydroxyecdysone and juvenile hormone on octopamine metabolism in females of Drosophila.

The effect of exogenous 20-hydroxyecdysone (20E) and juvenile hormone (JH) on the activities of the tyrosine decarboxylase (TDC), the first enzyme in octopamine (OA) synthesis, has been studied in young females of wild type D. virilis and D. melanogaster under normal and heat stress (38 degrees C) conditions. Flies fed 20E expressed increased TDC activity in both species. JH application decreased TDC activity in both species. A rise in JH and 20E levels did not prevent a TDC response to heat stress, but changed the response intensity. A long-term increase in JH titre had no effect on the activity of main OA catabolyzing enzyme, arylalkylamine N-acetyltransferase, in females of both species. A possible mechanism of regulation of OA levels by 20E and JH in Drosophila females is discussed.

Role of ecdysone 20-monooxygenase in regulation of 20-hydroxyecdysone levels by juvenile hormone and biogenic amines in Drosophila.

The effects of increased levels of dopamine (feeding flies with dopamine precursor, L: -dihydroxyphenylalanine) and octopamine (feeding flies with octopamine) on ecdysone 20-monooxygenase activity in young (2 days old) wild type females (the strain wt) of Drosophila virilis have been studied. L: -dihydroxyphenylalanine and octopamine feeding increases ecdysone 20-monooxygenase activity by a factor of 1.6 and 1.7, respectively. Ecdysone 20-monooxygenase activity in the young (1 day old) octopamineless females of the strain Tbetah ( nM18 ), in females of the strain P845 (precursor of Tbetah ( nM18 ) strain) and in wild type females (Canton S) of Drosophila melanogaster have been measured. The absence of octopamine leads to a considerable decrease in the enzyme activity. We have also studied the effects of juvenile hormone application on ecdysone 20-monooxygenase activity in 2-day-old wt females of D. virilis and demonstrated that an increase in juvenile hormone titre leads to an increase in the enzyme activity. We discuss the supposition that ecdysone 20-monooxygenase occupies a key position in the regulation of 20-hydroxyecdysone titre under the conditions that lead to changes in juvenile hormone titre and biogenic amine levels.