RESUMEN
Regulation of cell and organ sizes is fundamental for all organisms, but its molecular basis is not fully understood. Here we performed a gain-of-function screen and identified larp4B whose overexpression reduces cell and organ sizes in Drosophila melanogaster. Larp4B is a member of La-related proteins (LARPs) containing an LA motif and an adjacent RNA recognition motif (RRM), and play diverse roles in RNA metabolism. However, the function of Larp4B has remained poorly characterized. We generated transgenic flies overexpressing wild-type Larp4B or a deletion variant lacking the LA and RRM domains, and demonstrated that the RNA-binding domains are essential for Larp4B to reduce cell and organ sizes. We found that the larp4B-induced phenotype was suppressed by dMyc overexpression, which promotes cell growth and survival. Furthermore, overexpression of larp4B decreased dMyc protein levels, whereas its loss-of-function mutation had an opposite effect. Our results suggest that Larp4B is a negative regulator of dMyc.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila melanogaster/crecimiento & desarrollo , Drosophila melanogaster/genética , Factores de Transcripción/genética , Regulación hacia Arriba , Animales , Tamaño de la Célula , Proteínas de Unión al ADN/genética , Proteínas de Drosophila/química , Drosophila melanogaster/química , Drosophila melanogaster/citología , Femenino , Regulación del Desarrollo de la Expresión Génica , Genotipo , Tamaño de los Órganos , Fenotipo , Dominios Proteicos , Factores de Transcripción/químicaRESUMEN
Succinyl-CoA synthetase/ligase (SCS) is a mitochondrial enzyme that catalyzes the reversible process from succinyl-CoA to succinate and free coenzyme A in TCA cycle. SCS deficiencies are implicated in mitochondrial hepatoencephalomyopathy in humans. To investigate the impact of SCS deficiencies in Drosophila, we generated a null mutation in Scs alpha subunit (Scsα) using the CRISPR/Cas9 system, and characterized their phenotype. We found that the Drosophila SCS deficiency, designated ScsαKO, contained a high level of succinyl-CoA, a substrate for the enzyme, and altered levels of various metabolites in TCA cycle and glycolysis, indicating that the energy metabolism was impaired. Unlike SCSα deficiencies in humans, there was no reduction in lifespan, indicating that Scsα is not critical for viability in Drosophila. However, they showed developmental delays, locomotor activity defects, and reduced survival under starvation. We also found that glycogen breakdown occurred during development, suggesting that the mutant flies were unable to produce sufficient energy to promote normal growth. These results suggested that SCSα is essential for proper energy metabolism in Drosophila. The ScsαKO flies should be useful as a model to understand the physiological role of SCSα as well as the pathophysiology of SCSα deficiency.
Asunto(s)
Acilcoenzima A/deficiencia , Proteínas de Drosophila/deficiencia , Drosophila melanogaster/fisiología , Metabolismo Energético , Privación de Alimentos , Locomoción , Acilcoenzima A/genética , Animales , Animales Modificados Genéticamente , Conducta Animal , Supervivencia Celular , Ciclo del Ácido Cítrico , Proteínas de Drosophila/genética , Drosophila melanogaster/enzimología , Drosophila melanogaster/genética , Glucógeno/metabolismo , Glucólisis , Masculino , Mitocondrias/metabolismo , Fenotipo , InaniciónRESUMEN
Insulin/insulin-like growth factor (IGF) plays an important role as a systemic regulator of metabolism in multicellular organisms. Hyperinsulinemia, a high level of blood insulin, is often associated with impaired physiological conditions such as hypoglycemia, insulin resistance, and diabetes. However, due to the complex pathophysiology of hyperinsulinemia, the causative role of excess insulin/IGF signaling has remained elusive. To investigate the biological effects of a high level of insulin in metabolic homeostasis and physiology, we generated flies overexpressing Drosophila insulin-like peptide 2 (Dilp2), which has the highest potential of promoting tissue growth among the Ilp genes in Drosophila. In this model, a UAS-Dilp2 transgene was overexpressed under control of sd-Gal4 that drives expression predominantly in developing imaginal wing discs. Overexpression of Dilp2 caused semi-lethality, which was partially suppressed by mutations in the insulin receptor (InR) or Akt1, suggesting that dilp2-induced semi-lethality is mediated by the PI3K/Akt1 signaling. We found that dilp2-overexpressing flies exhibited intensive autophagy in fat body cells. Interestingly, the dilp2-induced autophagy as well as the semi-lethality was partially rescued by increasing the protein content relative to glucose in the media. Our results suggest that excess insulin/IGF signaling impairs the physiology of animals, which can be ameliorated by controlling the nutritional balance between proteins and carbohydrates, at least in flies.
RESUMEN
The insulin/insulin-like growth factor (IGF) and the target of rapamycin (TOR) signaling pathways are known to regulate lifespan in diverse organisms. However, only a limited number of genes involved in these pathways have been examined regarding their effects on lifespan. Through a gain-of-function screen in Drosophila, we found that overexpression of the wdb gene encoding a regulatory subunit of PP2A, and overexpression of the lkb1 gene encoding a serine/threonine kinase, reduced organ size and extended lifespan. Overexpression of wdb also reduced the level of phosphorylated AKT, while overexpression of lkb1 increased the level of phosphorylated AMPK and decreased the level of phosphorylated S6K. Taken together, our results suggest that wdb- and lkb1-dependent lifespan extension is mediated by downregulation of S6K, a downstream component of the insulin/IGF and TOR signaling pathways.
Asunto(s)
Proteínas de Drosophila/fisiología , Drosophila melanogaster/fisiología , Longevidad/genética , Proteínas Quinasas/fisiología , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/anatomía & histología , Drosophila melanogaster/genética , Tamaño de los Órganos/genética , Proteínas Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Alas de Animales/anatomía & histologíaRESUMEN
MachiBase (http://machibase.gi.k.u-tokyo.ac.jp/) provides a comprehensive and freely accessible resource regarding Drosophila melanogaster 5'-end mRNA transcription at different developmental states, supporting studies on the variabilities of promoter transcriptional activities and gene-expression profiles in the fruitfly. The data were generated in conjunction with the recently developed high-throughput genome sequencer Illumina/Solexa using a newly developed 5'-end mRNA collection method.