Drosophila HNF4 Regulates Lipid Mobilization and β-Oxidation

Abstract

Summary Drosophila HNF4 (dHNF4) is the single ancestral ortholog of a highly conserved subfamily of nuclear receptors that includes two mammalian receptors, HNFα and HNFγ, and 269 members in C. elegans. We show here that dHNF4 null mutant larvae are sensitive to starvation. Starved mutant larvae consume glycogen normally, but retain lipids in their midgut and fat body, and have increased levels of long chain fatty acids, suggesting that they are unable to efficiently mobilize stored fat for energy. Microarray studies support this model, indicating reduced expression of genes that control lipid catabolism and β-oxidation. A GAL4-dHNF4;UAS-lacZ ligand sensor can be activated by starvation or exogenous long chain fatty acids, suggesting that dHNF4 is responsive to dietary signals. Taken together, our results support a feed-forward model for dHNF4, in which fatty acids released from triglycerides activate the receptor, inducing enzymes that drive fatty acid oxidation for energy production.

An obligatory role for neurotensin in high-fat-diet-induced obesity

“…with saline or NT (3600 nmol/kg body weight), and, following sacrifice 60 min later, proximal intestinal fragments were dissected as described above and further divided into proximal and distal halves that were either processed for ORO staining or used for TG quantification, respectively, as described previously with modifications 44 . Briefly, intestinal tissues were homogenized in 200 μl of 0.5% Tween-20 in PBS, heated for 5 min at 70°C, homogenates were microcentrifuged (3 min), and TG measurements were performed on the supernatants using a Triglyceride Determination Kit (Sigma-Aldrich) 44 according to the manufacturer’s instructions.…”

“…Briefly, flies (5d after emerging) were collected and maintained on standard fly food for an additional 5d, split into two groups, and fed either standard fly food or standard fly food containing 20% (w/v) coconut oil for a further 5d, followed by dissection and lipid staining using Bodipy or Nile Red. voila -NT and voila -Gal4-w118 control flies were fed HFD or standard fly food as described above for TG measurement (25 flies/sample) as described 44 . Fluorescence signals were acquired on an Olympus confocal microscope and images processed with Olympus FV10-ASW Ver.3.1b.…”

Curcumin enhances parental reproductive lifespan and progeny viability in Drosophila melanogaster

“…The tissue from 20 female flies was pooled for the assay of glycogen. Glycogen levels were determined as described by using a standard protocol (Palanker et al 2009) with minor modifications. In brief, flies were homogenized in ice-cold homogenization buffer (20 mM HEPES, 0.2 % sodium azide, 0.2 mM PMSF, and 1 mM EDTA, pH 7.0) and centrifuged at 17,000 rpm for 3 min at 4°C, and 25 μL of the supernatant was incubated with 100 μL glucose reagent (Sigma) to measure the glucose level, or 100 μL glucose reagent and 0.5 U amyloglucosidase (Sigma) to measure the glycogen level.…”

“…Triglyceride levels were determined by using a standard protocol (Palanker et al 2009) with minor modifications. Twenty female flies were homogenized in 100 μL phosphate buffered saline (PBS) and 0.5 % Tween 20 and immediately incubated at 70°C for 5 min.…”

Systemic Activin signaling independently regulates sugar homeostasis, cellular metabolism, and pH balance in Drosophila melanogaster

“…TAG, glycogen, and hemolymph sugar assays were performed as described elsewhere (17,48). For hemolymph pH assay, 3 μL of hemolymph was mixed with 2 μL of pyranine dye (Invitrogen; 1.2 mM final concentration).…”