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Hippo signaling instructs ectopic but not normal organ growth [Drosophila NonPupatingLarvae]

ABSTRACT: We fed wildtype larvae a special diet that lacks precursor molecules necessary for the synthesis of the molting hormone ecdysone required for pupation, thus keeping animals in the larval stage. Importantly, in these larvae, the imaginal discs grow until their terminal size is reached but they do not proceed with differentiation as this requires a pulse of ecdysone. This method thus allowed us to directly compare actively growing discs with discs at the same developmental stage that terminated proliferation after reaching their proper size.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE211457 | GEO | 2022/11/19

REPOSITORIES: GEO

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Project description:Molting is an important physiological process in the larval stage of Bombyx mori and is controlled by various hormones and peptides. The silkworm mutant that exhibits the phenotype of non-molting in the 2nd instar (nm2) is incapable of molting in the 2nd instar and dies after seven or more days. The ecdysone titer in the nm2 mutant is lower than that in the wildtype, and the mutant can be rescued by feeding with 20E and cholesterol. The results of positional cloning indicated that structural alteration of BmCPG10 is responsible for the phenotype of the nm2 mutant. To explore the possible relationship between BmCPG10 and the ecdysone titer as well as the genes affected by BmCPG10, digital gene expression (DGE) profile analysis was conducted in the nm2 mutant, with the wildtype strain C603 serving as the control. The results revealed 1727 differentially expressed genes, among which 651 genes were upregulated and 1076 were downregulated in nm2. BLASTGO analysis showed that these differentially expressed genes were involved in various biological processes, cellular components and molecular functions. KEGG analysis indicated an enrichment of these differentially expressed genes in 240 pathways, including metabolic pathways, pancreatic secretion, protein digestion and absorption, fat digestion and absorption and glycerolipid metabolism. To verify the accuracy of the DGE results, quantitative reverse transcription PCR (qRT-PCR) was performed, focusing on key genes in several related pathways, and the results were highly consistent with the DGE results. Our findings indicated significant differences in cuticular protein genes, ecdysone biosynthesis genes and ecdysone-related nuclear receptors genes, but no significant difference in juvenile hormone and chitin biosynthesis genes was detected. Considering the physiological characteristics of nm2 together with the results of DGE analysis, we speculate that there may be a relationship between BmCPG10 and ecdysone. In the mutant, the original function of BmCPG10 was lost due to the change in its protein structure. It appeared that BmCPG10 might influence the synthesis of ecdysone by controlling the production of substrate cholesterol. However, under a low titer of ecdysone, the expression of BmCPG10 was affected, as one of a series of effectors of the ecdysone signaling pathway. Blocking ecdysone biosynthesis resulted in non-molting silkworms and the formation of the nm2 mutant.

Project description:Although radiation effects have been extensively studied, the biological effects of low-dose radiation (LDR) are controversial. This study investigates LDR-induced alterations in locomotive behavior and gene expression profiles of Drosophila melanogaster. We measured locomotive behavior using larval pupation height and rapid iterative negative geotaxis (RING) assay after exposure to 0.1 Gy gamma-radiation (dose rate of 16.7 mGy/h). We also observed chronic LDR effects on development (pupation and eclosion rates) and longevity (life span). To identify chronic LDR effects on gene expression, we performed whole-genome expression analysis using gene-expression microarrays, and confirmed the results using quantitative real-time PCR. Pupation height was significantly higher after LDR treatment at the first larval instar. Locomotive behavior of male flies was significantly greater approximately 3M-BM--5 weeks after LDR, but pupation and eclosion rates and life spans were not significantly different. Genome-wide expression analysis identified 344 genes that were differentially expressed in irradiated larvae compared with those of controls. We identified several genes belonging to larval behavior functional groups such as locomotive behavior and oxidation reduction, and genes involved in conventional functional groups modulated by irradiation such as defense response, sensory and perception. Four candidate genes were confirmed as differentially expressed genes in irradiated larvae using qRT-PCR. These data suggest that LDR stimulates locomotion-related genes, and these genes can be used as potential markers for LDR. Eggs were collected from 5-day-old female flies and cultivated for 24 h on standard medium. Then, twenty larvae were manually selected and seeded on fresh standard medium in a new vial. After transfer, the experimental group of first-instar larvae was immediately irradiated with chronic gamma-radiation at a dose rate of 16.7 mGy/h. After treatment, gamma-irradiated flies and non-irradiated control flies were maintained in the same incubator at 25degC.

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Hippo signaling instructs ectopic but not normal organ growth [Drosophila NonPupatingLarvae]

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