Wnt Signaling Cross-Talks with JH Signaling by Suppressing Met and gce Expression

Juvenile hormone (JH) plays key roles in controlling insect growth and metamorphosis. However, relatively little is known about the JH signaling pathways. Until recent years, increasing evidence has suggested that JH modulates the action of 20-hydroxyecdysone (20E) by regulating expression of broad (br), a 20E early response gene, through Met/Gce and Kr-h1. To identify other genes involved in JH signaling, we designed a novel Drosophila genetic screen to isolate mutations that derepress JH-mediated br suppression at early larval stages. We found that mutations in three Wnt signaling negative regulators in Drosophila, Axin (Axn), supernumerary limbs (slmb), and naked cuticle (nkd), caused precocious br expression, which could not be blocked by exogenous JHA. A similar phenotype was observed when armadillo (arm), the mediator of Wnt signaling, was overexpressed. qRT-PCR revealed that Met, gce and Kr-h1expression was suppressed in the Axn, slmb and nkd mutants as well as in arm gain-of-function larvae. Furthermore, ectopic expression of gce restored Kr-h1 expression but not Met expression in the arm gain-of-function larvae. Taken together, we conclude that Wnt signaling cross-talks with JH signaling by suppressing transcription of Met and gce, genes that encode for putative JH receptors. The reduced JH activity further induces down-regulation of Kr-h1expression and eventually derepresses br expression in the Drosophila early larval stages

Methyl Farnesoate Plays a Dual Role in Regulating \u3cem\u3eDrosophila\u3c/em\u3e Metamorphosis

Corpus allatum (CA) ablation results in juvenile hormone (JH) deficiency and pupal lethality in Drosophila. The fly CA produces and releases three sesquiterpenoid hormones: JH III bisepoxide (JHB3), JH III, and methyl farnesoate (MF). In the whole body extracts, MF is the most abundant sesquiterpenoid, followed by JHB3 and JH III. Knockout of JH acid methyl transferase (jhamt) did not result in lethality; it decreased biosynthesis of JHB3, but MF biosynthesis was not affected. RNAi-mediated reduction of 3-hydroxy-3-methylglutaryl CoA reductase (hmgcr) expression in the CA decreased biosynthesis and titers of the three sesquiterpenoids, resulting in partial lethality. Reducing hmgcr expression in the CA of the jhamt mutant further decreased MF titer to a very low level, and caused complete lethality. JH III, JHB3, and MF function through Met and Gce, the two JH receptors, and induce expression of Kr-h1, a JH primary-response gene. As well, a portion of MF is converted to JHB3 in the hemolymph or peripheral tissues. Topical application of JHB3, JH III, or MF precluded lethality in JH-deficient animals, but not in the Met gce double mutant. Taken together, these experiments show that MF is produced by the larval CA and released into the hemolymph, from where it exerts its anti-metamorphic effects indirectly after conversion to JHB3, as well as acting as a hormone itself through the two JH receptors, Met and Gce

Methyl Farnesoate Plays a Dual Role in Regulating \u3cem\u3eDrosophila\u3c/em\u3e Metamorphosis

Corpus allatum (CA) ablation results in juvenile hormone (JH) deficiency and pupal lethality in Drosophila. The fly CA produces and releases three sesquiterpenoid hormones: JH III bisepoxide (JHB3), JH III, and methyl farnesoate (MF). In the whole body extracts, MF is the most abundant sesquiterpenoid, followed by JHB3 and JH III. Knockout of JH acid methyl transferase (jhamt) did not result in lethality; it decreased biosynthesis of JHB3, but MF biosynthesis was not affected. RNAi-mediated reduction of 3-hydroxy-3-methylglutaryl CoA reductase (hmgcr) expression in the CA decreased biosynthesis and titers of the three sesquiterpenoids, resulting in partial lethality. Reducing hmgcr expression in the CA of the jhamt mutant further decreased MF titer to a very low level, and caused complete lethality. JH III, JHB3, and MF function through Met and Gce, the two JH receptors, and induce expression of Kr-h1, a JH primary-response gene. As well, a portion of MF is converted to JHB3 in the hemolymph or peripheral tissues. Topical application of JHB3, JH III, or MF precluded lethality in JH-deficient animals, but not in the Met gce double mutant. Taken together, these experiments show that MF is produced by the larval CA and released into the hemolymph, from where it exerts its anti-metamorphic effects indirectly after conversion to JHB3, as well as acting as a hormone itself through the two JH receptors, Met and Gce

Biochemical and insecticidal efficacy of clove and basil essential oils and two photosensitizers and their combinations on Aphis gossypii glover (Hemiptera: Aphididae)

The present study investigates the insecticidal and biochemical effects of two essential oils (EOs) and two photosensitizers against cotton aphids in a laboratory setting. The EOs evaluated were clove (Syzygium aromaticum L.) and basil (Ocimum basilicum), while the photosensitizers were rose bengal and rhodamine B. The individual median lethal concentrations (LC50) revealed that clove was ~4.44 times more potent than basil, and rhodamine B was ~1.34 times more potent than rose bengal. The mortality rates increased using higher concentrations of the photosensitizers and prolonging exposure time to sunlight. The most effective combination against adult aphids was found to be a mixture of sub-lethal concentrations of clove and rhodamine B, resulting in a mortality rate of 92.31%. Conversely, the combination of basil and rose bengal exhibited the lowest efficacy with a mortality rate of 33.33%. Biochemical analyses indicate that Rhodamine B, basil, and the basil-rhodamine B mixture (mixture C) significantly reduced trehalase activity. However, the protease activity significantly increased in aphids treated with rose bengal, clove, and the clove-rose bengal mixtures (mixtures A and B). The lipase activity is notably decreased upon treatment with rhodamine B and clove. Glutathione S-transferase (GST) activity decreased in aphids treated with rose bengal and the basil-rhodamine B mixtures (mixtures C and D), suggesting that GST did not play a role in detoxifying these compounds, thereby explaining the susceptibility of A. gossypii. Overall, the combination of essential oils and photosensitizers has demonstrated a synergistic effect in controlling Aphis gossypii, offering great potential as an effective strategy for aphid management

<i>GAL4-PG12</i> resembles endogenous <i>br</i> expression patterns.

<p>(A) Protein extracts isolated from wild type animals at different developmental stages were separated by SDS-PAGE. Br proteins were assessed by Western blotting using a Br-core antibody. Tubulin-β was used as a loading control. The Br proteins were only detected in the late 3^rd instar larval stage to pupal stage. All Br isoforms were expressed in the late 3^rd instar larvae and early pupae, but only Z1 and/or Z3 isoforms were expressed in the late pupae. (B–F) Expression of <i>GAL4-PG12</i> was marked by <i>GAL4-PG12,UAS-mCD8GFP</i>. Constitutive expression of <i>GAL4-PG12</i> in salivary glands (arrows) and auto-fluorescence of fly food in the midgut (arrowheads) are indicated. In tissues other than those from the salivary gland, <i>GAL4-PG12/UAS-mCD8GFP</i> was only expressed in late 3^rd instar larval and during early pupal stages (G and H). (B′–F′) White light images of the same organisms are shown in [B–F]. (G–I) <i>GAL4-PG12</i> expression was monitored by mCD8GFP (green) [G–I]. Endogenous Br proteins were recognized by a Br-core antibody (red) [G′–I′] and nuclei were marked with DAPI (blue) [G″–H″]. Neither endogenous Br nor <i>GAL4-PG12</i> were expressed in FB of the 2^nd instar or early 3^rd instar [G-G″ and H-H″], but both were expressed in FB of the late 3^rd instar [I-I″]. [I″] is a merged image of [I] and [I′].</p

Ectopic expression of JHE induces precocious <i>br</i> expression in the 2^nd instar larvae.

<p>Flies carrying two copies of <i>hs-jhe</i> transgenes (<i>GAL4-PG12, UAS-mCD8GFP/Fm7C; hs-jhe¹, hs-jhe²/+</i>) were reared on normal (−JHA) or 0.1 ppm pyriproxifen-containing (+JHA) food and were treated with (HS) or without (no-HS) heat shocking twice a day for 40 min at 37°C. <i>Br</i> expression was monitored by <i>GAL4-PG12,UAS-mCD8GFP</i> [A–D] and FB Br-core antibody staining in 2^nd instar larvae [E–H]. Precocious <i>br</i> expression occurred in 2^nd instar larvae that were reared on normal food and treated with heat shocking [B-B′ and F-F′]. However, this phenotype was blocked by JHA treatment [D-D′ and H-H′].</p

A genetic screen identifies that Axn, Slmb and Nkd regulate <i>br</i> expression.

<p>(A). Schematic diagram of genetic crosses for isolating mutations that derepress <i>br</i> expression in young larvae. <i>GAL4-PG12,UAS-mCD8GFP</i> (X chromosome) was used to monitor <i>br</i> expression. The lethal mutation or <i>P</i>-insertion on the 2^nd or 3^rd chromosome is represented by an asterisk (*). (B–E). GFP images show the expression of <i>GAL4-PG12,UASmCD8GFP</i> in 2^nd instar larvae. GFP was only expressed in the salivary gland of the wild type [B], but widely expressed in all tissues of <i>Axn^EY10228</i> [C], <i>slmb⁰⁰²⁹⁵</i> [D], and <i>nkd²</i> [E] mutant larvae. (B′–E′) White light images of the same organisms are shown in [B–E].</p

Precocious <i>br</i> expression occurs in <i>Axn</i> mutants in a tissue-specific manner.

<p>2^nd instar larvae of <i>Oregon</i> R and <i>Axn^EY10228</i> were dissected and stained with a Br-core antibody (red). Nuclei were labeled with DAPI (blue). Images show central nervous system (CNS) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0026772#pone-0026772-g006" target="_blank">Fig. 6A and D</a>), fat body (FB) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0026772#pone-0026772-g006" target="_blank">Fig. 6B and E</a>) and midgut (MG) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0026772#pone-0026772-g006" target="_blank">Fig. 6C and F</a>).</p

<i>GAL4-PG12</i> carries a <i>P</i>-element insertion in the first intron of <i>br</i> gene.

<p>(A) The flanking sequence of the <i>GAL4-PG12 P</i>-element insertion site identified by inverse PCR analysis. (B) The insertion site of <i>GAL4-PG12</i> was located within the first intron of the <i>br</i> gene by comparing the sequence with the <i>Drosophila</i> genome.</p

Phytochemical Investigation of Three <i>Cystoseira</i> Species and Their Larvicidal Activity Supported with In Silico Studies

Culex pipiens mosquitoes are transmitters of many viruses and are associated with the transmission of many diseases, such as filariasis and avian malaria, that have a high rate of mortality. The current study draws attention to the larvicidal efficacy of three methanolic algal extracts, Cystoseira myrica, C. trinodis, and C. tamariscifolia, against the third larval instar of Cx. pipiens. The UPLC-ESI-MS analysis of three methanol fractions of algal samples led to the tentative characterization of twelve compounds with different percentages among the three samples belonging to phenolics and terpenoids. Probit analysis was used to calculate the lethal concentrations (LC50 and LC90). The highest level of toxicity was attained after treatment with C. myrica extract using a lethal concentration 50 (LC50) of 105.06 ppm, followed by C. trinodis (135.08 ppm), and the lowest level of toxicity was achieved by C. tamariscifolia (138.71 ppm) after 24 h. The elevation of glutathione-S-transferase (GST) and reduction of acetylcholine esterase (AChE) enzymes confirm the larvicidal activity of the three algal extracts. When compared to untreated larvae, all evaluated extracts revealed a significant reduction in protein, lipid, and carbohydrate contents, verifying their larvicidal effectiveness. To further support the observed activity, an in silico study for the identified compounds was carried out on the two tested enzymes. Results showed that the identified compounds and the tested enzymes had excellent binding affinities for each other. Overall, the current work suggests that the three algal extractions are a prospective source for the development of innovative, environmentally friendly larvicides