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Expression of human uncoupling protein-3 in Drosophila insulin-producing cells increases insulin-like peptide (DILP) levels and shortens lifespan.

Humphrey DM, Toivonen JM, Giannakou M, Partridge L, Brand MD - Exp. Gerontol. (2009)

Bottom Line: Low, ubiquitous expression of hUCP3 at levels found in rodent skeletal muscle mitochondria did not affect proton conductance in mitochondria isolated from whole flies, but high pan-neuronal expression of hUCP3 increased the proton conductance of mitochondria isolated from fly heads.Expression of hUCP3 in the mNSC did not alter expression of dilp2, dilp3 or dilp5 mRNA, but led to increased amounts of DILP2 in fly heads.These data suggest that lowering mitochondrial coupling by high expression of hUCP3 alters mNSC function in a way that appears to increase DILP-levels in fly heads and lead to a concomitant decrease in lifespan.

View Article: PubMed Central - PubMed

Affiliation: MRC Dunn Human Nutrition Unit, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK. dickon.humphrey@iop.kcl.ac.uk

ABSTRACT
Uncoupling proteins (UCPs) can dissipate mitochondrial protonmotive force by increasing the proton conductance of the inner membrane and through this effect could decrease ROS production, ameliorate oxidative stress and extend lifespan. We investigated whether ubiquitous, pan-neuronal or neurosecretory cell-specific expression of human UCP3 (hUCP3) in adult Drosophila melanogaster affected lifespan. Low, ubiquitous expression of hUCP3 at levels found in rodent skeletal muscle mitochondria did not affect proton conductance in mitochondria isolated from whole flies, but high pan-neuronal expression of hUCP3 increased the proton conductance of mitochondria isolated from fly heads. Expression of hUCP3 at moderate levels in adult neurons led to a marginal lifespan-extension in males. However, high expression of hUCP3 in neuronal tissue shortened lifespan. The life-shortening effect was replicated when hUCP3 was expressed specifically in median neurosecretory cells (mNSC), which express three of the Drosophila insulin-like peptides (DILPs). Expression of hUCP3 in the mNSC did not alter expression of dilp2, dilp3 or dilp5 mRNA, but led to increased amounts of DILP2 in fly heads. These data suggest that lowering mitochondrial coupling by high expression of hUCP3 alters mNSC function in a way that appears to increase DILP-levels in fly heads and lead to a concomitant decrease in lifespan.

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Kinetics of proton leak in head mitochondria from a high neuronal hUCP3 expresser. (A) The dependence of proton leak on membrane potential was determined as described in Section 2. Mitochondria were isolated from heads from UAS-hUCP3-high/elav-GS flies (line J) and +/elav-GS driver control flies with (+) and without (-) RU486. The vertical dotted line indicates the highest potential common to all individual data points (110.3 mV). (B) Respiration rate driving proton leak at 110.3 mV for the four groups. Significance was determined using one-way ANOVA. Data from three separate mitochondrial preparations performed singly or in duplicate.
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fig3: Kinetics of proton leak in head mitochondria from a high neuronal hUCP3 expresser. (A) The dependence of proton leak on membrane potential was determined as described in Section 2. Mitochondria were isolated from heads from UAS-hUCP3-high/elav-GS flies (line J) and +/elav-GS driver control flies with (+) and without (-) RU486. The vertical dotted line indicates the highest potential common to all individual data points (110.3 mV). (B) Respiration rate driving proton leak at 110.3 mV for the four groups. Significance was determined using one-way ANOVA. Data from three separate mitochondrial preparations performed singly or in duplicate.

Mentions: Proton leak calculations were performed in Microsoft Excel. When comparing proton-leak curves at a fixed membrane potential, the FORECAST function was used to calculate respiration rates between data points (vertical dotted line in Fig. 3A). Mean and standard errors in Fig. 3B were calculated from individual repeats. Student’s t-test was used to determine significance of differences between two groups of data. When more than two groups of data were compared, variance was measured using one-way ANOVA followed by Tukey’s multiple comparison post tests. The significance confidence limit was set at 95%. Statistical analysis was performed using GraphPad Prism version 4.0a for Mac OS X (GraphPad Software, San Diego California USA). Log-rank tests of survivorship curves were performed using JMP™ IN statistical software (SAS Institute Inc.).


Expression of human uncoupling protein-3 in Drosophila insulin-producing cells increases insulin-like peptide (DILP) levels and shortens lifespan.

Humphrey DM, Toivonen JM, Giannakou M, Partridge L, Brand MD - Exp. Gerontol. (2009)

Kinetics of proton leak in head mitochondria from a high neuronal hUCP3 expresser. (A) The dependence of proton leak on membrane potential was determined as described in Section 2. Mitochondria were isolated from heads from UAS-hUCP3-high/elav-GS flies (line J) and +/elav-GS driver control flies with (+) and without (-) RU486. The vertical dotted line indicates the highest potential common to all individual data points (110.3 mV). (B) Respiration rate driving proton leak at 110.3 mV for the four groups. Significance was determined using one-way ANOVA. Data from three separate mitochondrial preparations performed singly or in duplicate.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2698063&req=5

fig3: Kinetics of proton leak in head mitochondria from a high neuronal hUCP3 expresser. (A) The dependence of proton leak on membrane potential was determined as described in Section 2. Mitochondria were isolated from heads from UAS-hUCP3-high/elav-GS flies (line J) and +/elav-GS driver control flies with (+) and without (-) RU486. The vertical dotted line indicates the highest potential common to all individual data points (110.3 mV). (B) Respiration rate driving proton leak at 110.3 mV for the four groups. Significance was determined using one-way ANOVA. Data from three separate mitochondrial preparations performed singly or in duplicate.
Mentions: Proton leak calculations were performed in Microsoft Excel. When comparing proton-leak curves at a fixed membrane potential, the FORECAST function was used to calculate respiration rates between data points (vertical dotted line in Fig. 3A). Mean and standard errors in Fig. 3B were calculated from individual repeats. Student’s t-test was used to determine significance of differences between two groups of data. When more than two groups of data were compared, variance was measured using one-way ANOVA followed by Tukey’s multiple comparison post tests. The significance confidence limit was set at 95%. Statistical analysis was performed using GraphPad Prism version 4.0a for Mac OS X (GraphPad Software, San Diego California USA). Log-rank tests of survivorship curves were performed using JMP™ IN statistical software (SAS Institute Inc.).

Bottom Line: Low, ubiquitous expression of hUCP3 at levels found in rodent skeletal muscle mitochondria did not affect proton conductance in mitochondria isolated from whole flies, but high pan-neuronal expression of hUCP3 increased the proton conductance of mitochondria isolated from fly heads.Expression of hUCP3 in the mNSC did not alter expression of dilp2, dilp3 or dilp5 mRNA, but led to increased amounts of DILP2 in fly heads.These data suggest that lowering mitochondrial coupling by high expression of hUCP3 alters mNSC function in a way that appears to increase DILP-levels in fly heads and lead to a concomitant decrease in lifespan.

View Article: PubMed Central - PubMed

Affiliation: MRC Dunn Human Nutrition Unit, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK. dickon.humphrey@iop.kcl.ac.uk

ABSTRACT
Uncoupling proteins (UCPs) can dissipate mitochondrial protonmotive force by increasing the proton conductance of the inner membrane and through this effect could decrease ROS production, ameliorate oxidative stress and extend lifespan. We investigated whether ubiquitous, pan-neuronal or neurosecretory cell-specific expression of human UCP3 (hUCP3) in adult Drosophila melanogaster affected lifespan. Low, ubiquitous expression of hUCP3 at levels found in rodent skeletal muscle mitochondria did not affect proton conductance in mitochondria isolated from whole flies, but high pan-neuronal expression of hUCP3 increased the proton conductance of mitochondria isolated from fly heads. Expression of hUCP3 at moderate levels in adult neurons led to a marginal lifespan-extension in males. However, high expression of hUCP3 in neuronal tissue shortened lifespan. The life-shortening effect was replicated when hUCP3 was expressed specifically in median neurosecretory cells (mNSC), which express three of the Drosophila insulin-like peptides (DILPs). Expression of hUCP3 in the mNSC did not alter expression of dilp2, dilp3 or dilp5 mRNA, but led to increased amounts of DILP2 in fly heads. These data suggest that lowering mitochondrial coupling by high expression of hUCP3 alters mNSC function in a way that appears to increase DILP-levels in fly heads and lead to a concomitant decrease in lifespan.

Show MeSH
Related in: MedlinePlus