Mio acts in the Drosophila brain to control nutrient storage and feeding

Abstract

Animals recognize the availability of nutrients and regulate the intake and storage of these nutrients accordingly. However, the molecular mechanisms underlying nutrient sensing and subsequent changes in behavior and metabolism are not fully understood. Mlx interactor (Mio), the Drosophila homolog of carbohydrate response element binding protein (ChREBP), functions as a transcription factor in the fat body of the fly to control triglyceride storage as well as feeding, suggesting that Mio may act in a nutrient-sensing pathway to coordinate food consumption and metabolism. Here, we show that Mio functions in neurons in Drosophila to regulate feeding and nutrient storage. Pan-neuronal disruption of Mio function leads to increased triglyceride and glycogen storage, and this phenotype is not due to increased food consumption. Interestingly, targeted disruption of Mio specifically in the insulin-producing cells (IPCs) has little effect on nutrient storage, but increases food consumption suggesting that Mio acts in these neurons to control feeding behavior. Since Mio is a transcription factor, one possible way Mio may act in the IPCs to control feeding is through regulating the expression of Drosophila insulin-like peptides (dilps) or drosulfakinin (dsk), neuropeptides produced in the IPCs. Consistent with this hypothesis, IPC-specific knockdown of Mio leads to an increase in dilp3 expression, while not affecting dilp2, 5 or dsk levels. Together, this study indicates a new function for Mio in the Drosophila brain and specifically in the IPCs, controlling neuropeptide gene expression, feeding and metabolism in accordance with nutrient availability.

Keywords: Brain; Drosophila; Feeding; Insulin-like peptide; Metabolism.

Fig. 1. Mio levels are reduced when Mio-RNAi is targeted to the nervous system

(A) nSyb-Gal4>GFP shows fluorescence expression in neurons. Arrows show GFP fluorescence in the lobes of the brain and the ventral nerve chord. (B) Quantitative PCR was performed for Mio using cDNA from batches of 30 heads from 5–8 day old nSyb-Gal4>Mio-IR (n=12) and nSyb-Gal4>Mio^dsRNA (n=6) animals compared to nSyb-Gal4>GFP^dsRNA controls (n=6). Values represent means ± SEM. *P < 0.05 by one-way independent weighted ANOVA and Tukey post-hoc test comparing each experimental genotype to the GFP^dsRNA control.

Fig. 2. Mio acts in the brain to regulate macromolecule storage, but not food consumption

(A) Triglyceride/protein and (B) glycogen/protein ratios from 5–8 day old nSyb-Gal4>Mio-IR (n=20) and nSyb-Gal4>Mio^dsRNA (n=45) flies compared to nSyb-Gal4>GFP^dsRNA controls (n=34). (C) Total food consumption over 24hrs was measured using the CAFE assay in 5–8 day old nSyb-Gal4>Mio-IR (n=13) and nSyb-Gal4>Mio^dsRNA (n=27) females compared to nSyb-Gal4>GFP^dsRNA controls (n=17). Values represent means ± SEM. *P < 0.05 by one-way, independent, weighted ANOVA and Tukey post-hoc test comparing each experimental genotype to the GFP^dsRNA control.

Fig. 3. Mio functions specifically in the IPCs to control feeding, but not nutrient storage

(A) Triglyceride/protein and (B) glycogen/protein ratios from 5–8 day old dilp2-Gal4>Mio-IR (n=56) and dilp2-Gal4>Mio^dsRNA (n=40) flies compared to dilp2-Gal4>GFP^dsRNA controls (n=36). (C) Total food consumption over 24hrs was measured using the CAFE assay on 5–8 day old dilp2-Gal4>Mio-IR (n=22) and dilp2-Gal4>Mio^dsRNA (n=15) flies compared to dilp2-Gal4>GFP^dsRNA controls (n=12). Values represent means ± SEM. *P < 0.05 by one-way, independent, weighted ANOVA and Tukey post-hoc test comparing each experimental genotype to its respective GFP^dsRNA control.

Fig. 4. Mio regulates dilp expression in the IPCs

Quantitative PCR was performed for (A) dilp2, (B) dilp3, (C) dilp5, and (D) dsk using cDNA from batches of 30 heads from 5–8 day old dilp2-Gal4>Mio-IR (n=4–8) and dilp2-Gal4>Mio^dsRNA (n=4–8) animals compared to dilp2-Gal4>GFP^dsRNA controls (n=6). Values represent means ± SEM. *P < 0.05 by one-way independent, weighted, ANOVA and Tukey post-hoc test comparing each experimental genotype to the GFP^dsRNA control.