Glutaredoxin-2 is required to control proton leak through uncoupling protein-3

Area of Science:

• Mitochondrial bioenergetics
• Redox regulation of protein function
• Post-translational modifications

Background:

• Glutathionylation controls protein function in response to cellular redox changes.
• Uncoupling protein-3 (UCP3) glutathionylation state modulates mitochondrial proton leak and reactive oxygen species production.
• The enzymatic basis for UCP3 glutathionylation remained unknown, with prior studies using chemical agents.

Purpose of the Study:

• To identify the enzyme responsible for UCP3 glutathionylation.
• To investigate the role of this enzyme in regulating UCP3 activity and mitochondrial function.
• To elucidate the impact of Grx2 on energy metabolism.

Main Methods:

• Mitochondrial bioenergetic analysis in wild-type and Grx2 knockout (Grx2(-/-)) mouse skeletal muscle mitochondria.
• Assessment of proton leak and coupled respiration.
• Experimental manipulation of Grx2 levels via knockdown in primary myotubes.
• Utilizing diamide as a glutathionylation catalyst.

Main Results:

• Grx2 knockout (Grx2(-/-)) mitochondria exhibited increased UCP3-dependent proton leak.
• This increased proton leak did not impair coupled respiration.
• Grx2 knockdown in primary myotubes augmented proton leak-dependent respiration, an effect absent in UCP3 knockout cells.
• Diamide treatment reversed the effects observed in Grx2 knockout mitochondria.

Conclusions:

• Glutaredoxin-2 (Grx2) is the first identified enzyme that directly mediates UCP3 glutathionylation, leading to its inhibition.
• Grx2 regulates mitochondrial proton leak and energy metabolism through the glutathionylation of UCP3.
• This study establishes a novel enzymatic mechanism for controlling UCP3 activity and its role in cellular redox homeostasis.