Ski3/TTC37 deficiency associated with trichohepatoenteric syndrome causes mitochondrial dysfunction in Drosophila.

Tetratricopeptide repeat protein 37 (TTC37) is a causative gene of trichohepatoenteric syndrome (THES). However, little is known about the pathogenesis of this disease. Here, we characterize the phenotype of a Drosophila model in which ski3, a homolog of TTC37, is disrupted. The mutant flies are pupal lethal, and the pupal lethality is partially rescued by transgenic expression of wild-type ski3 or human TTC37. The mutant larvae show growth retardation, heart arrhythmia, triacylglycerol accumulation, and aberrant metabolism of glycolysis and the TCA cycle. Moreover, mitochondrial membrane potential and respiratory chain complex activities are significantly reduced in the mutants. Our results demonstrate that ski3 deficiency causes mitochondrial dysfunction, which may underlie the pathogenesis of THES.

Guarana improves behavior and inflammatory alterations triggered by methylmercury exposure: an in vivo fruit fly and in vitro neural cells study.

Methylmercury (MeHg) is a well-known environmental pollutant associated with neurological and developmental deficits in animals and humans. However, epidemiological data showed that people living in the Amazon region although exposed to MeHg do not present these effects probably due to the protective effect of certain foods. We hypothesized here if guarana, a highly caffeinated fruit and consumed on a daily basis by Amazon people, could have some protective effect against MeHg toxicity using two complementary approaches. To assess locomotor impairment and sleep disruption, we used fruit fly (Drosophila melanogaster) model, and to evaluate neuroinflammation, we used human SH-SY5Y neural cells by measuring inflammatory cytokines levels. Results showed that guarana had a protective effect on the locomotor activity of male fruit flies reducing the excessive sleepiness caused by MeHg and increasing daily activity. Also, guarana increased the viability of flies and attenuated neural cells mortality. In addition, guarana reduced all pro-inflammatory cytokines levels increased by MeHg, along with caspase-1, caspase -3, caspase-8, and 8-dOHG levels, whereas increased the anti-inflammatory (IL-10) cytokine levels, which was decreased by MeHg. Our study provides new insights on the protective effects of guarana on the viability, locomotor activity, sleep, and activity patterns in vivo and the in vitro neuronal anti-inflammatory effect against MeHg toxicity.

Epigenetic regulation of the glucose transporter gene Slc2a1 by ß-hydroxybutyrate underlies preferential glucose supply to the brain of fasted mice.

We carried out liquid chromatography-tandem mass spectrometry analysis of metabolites in mice. Those metabolome data showed that hepatic glucose content is reduced, but that brain glucose content is unaffected, during fasting, consistent with the priority given to brain glucose consumption during fasting. The molecular mechanisms for this preferential glucose supply to the brain are not fully understood. We also showed that the fasting-induced production of the ketone body ß-hydroxybutyrate (ß-OHB) enhances expression of the glucose transporter gene Slc2a1 (Glut1) via histone modification. Upon ß-OHB treatment, Slc2a1 expression was up-regulated, with a concomitant increase in H3K9 acetylation at the critical cis-regulatory region of the Slc2a1 gene in brain microvascular endothelial cells and NB2a neuronal cells, shown by quantitative PCR analysis and chromatin immunoprecipitation assay. CRISPR/Cas9-mediated disruption of the Hdac2 gene increased Slc2a1 expression, suggesting that it is one of the responsible histone deacetylases (HDACs). These results confirm that ß-OHB is a HDAC inhibitor and show that ß-OHB plays an important role in fasting-induced epigenetic activation of a glucose transporter gene in the brain.

Deficiency of succinyl-CoA synthetase α subunit delays development, impairs locomotor activity and reduces survival under starvation in Drosophila.

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.

A Comprehensive Genomic Analysis Reveals the Genetic Landscape of Mitochondrial Respiratory Chain Complex Deficiencies.

Mitochondrial disorders have the highest incidence among congenital metabolic disorders characterized by biochemical respiratory chain complex deficiencies. It occurs at a rate of 1 in 5,000 births, and has phenotypic and genetic heterogeneity. Mutations in about 1,500 nuclear encoded mitochondrial proteins may cause mitochondrial dysfunction of energy production and mitochondrial disorders. More than 250 genes that cause mitochondrial disorders have been reported to date. However exact genetic diagnosis for patients still remained largely unknown. To reveal this heterogeneity, we performed comprehensive genomic analyses for 142 patients with childhood-onset mitochondrial respiratory chain complex deficiencies. The approach includes whole mtDNA and exome analyses using high-throughput sequencing, and chromosomal aberration analyses using high-density oligonucleotide arrays. We identified 37 novel mutations in known mitochondrial disease genes and 3 mitochondria-related genes (MRPS23, QRSL1, and PNPLA4) as novel causative genes. We also identified 2 genes known to cause monogenic diseases (MECP2 and TNNI3) and 3 chromosomal aberrations (6q24.3-q25.1, 17p12, and 22q11.21) as causes in this cohort. Our approaches enhance the ability to identify pathogenic gene mutations in patients with biochemically defined mitochondrial respiratory chain complex deficiencies in clinical settings. They also underscore clinical and genetic heterogeneity and will improve patient care of this complex disorder.

Overexpression of dilp2 causes nutrient-dependent semi-lethality in Drosophila.

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.