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Substitution of PIM1 protease in mitochondria by Escherichia coli Lon protease
U Teichmann1, L van Dyck, B Guiard
1Institut für Physiologische Chemie der Universität München, Goethestrasse 33, 80336 München, Germany.
Abstract:
PIM1 protease in mitochondria belongs to a conserved family of ATP-dependent proteases, which includes the Escherichia coli Lon protease. Yeast cells lacking PIM1 are largely defective in degrading misfolded proteins in the mitochondrial matrix, are respiratory deficient, and lose integrity of mitochondrial DNA. In order to analyze whether E. coli Lon protease is functionally equivalent to mitochondrial PIM1 protease, yeast cells lacking the PIM1 gene were transformed with a construct consisting of a mitochondrial targeting sequence fused onto the Lon protease. In these cells, the fusion protein was expressed and imported into mitochondria, and the targeting sequence was removed. In the absence of PIM1 protease, the E. coli Lon protease mediated the degradation of misfolded proteins in the matrix space in cooperation with the mitochondrial hsp70 system. These cells maintained the integrity of the mitochondrial genome and the respiratory function at 30 degrees C but not at 37 degrees C. Stabilization of mitochondrial DNA in Deltapim1 cells depended on protein degradation by the E. coli Lon protease, as a proteolytically inactive Lon variant was not capable of substituting for a loss of PIM1 protease. These results demonstrate functional conservation of Lon-like proteases from prokaryotes to eukaryotes and shed new light on the role of Lon-like proteases in mitochondrial biogenesis.
Insights
The E. coli Lon protease can functionally replace the mitochondrial PIM1 protease in yeast, degrading misfolded proteins and maintaining mitochondrial DNA integrity. This highlights the conserved function of Lon-like proteases across species.
Area of Science:
- Mitochondrial biology
- Protease function
- Molecular evolution
Background:
- Mitochondrial PIM1 protease is crucial for degrading misfolded proteins in yeast.
- Loss of PIM1 function leads to respiratory deficiency and mitochondrial DNA instability.
- The Escherichia coli Lon protease is a related ATP-dependent protease.
Purpose of the Study:
- To determine if E. coli Lon protease can functionally substitute for yeast PIM1 protease.
- To investigate the role of Lon-like proteases in mitochondrial protein quality control.
- To assess the impact of Lon protease on mitochondrial genome stability.
Main Methods:
- Yeast cells lacking PIM1 were transformed with a mitochondrial targeting sequence fused to E. coli Lon protease.
- The fusion protein was imported into mitochondria and the targeting sequence was cleaved.
- Misfolded protein degradation, respiratory function, and mitochondrial DNA integrity were assessed.
Main Results:
- E. coli Lon protease, when targeted to mitochondria, mediated degradation of misfolded proteins.
- Cells expressing Lon protease maintained mitochondrial genome integrity and respiratory function at 30°C but not 37°C.
- A proteolytically inactive Lon variant failed to rescue PIM1 loss, confirming the importance of protease activity.
Conclusions:
- Lon-like proteases are functionally conserved from prokaryotes to eukaryotes.
- Mitochondrial Lon-like proteases play a vital role in maintaining mitochondrial biogenesis and genome stability.
- Protein degradation by Lon-like proteases is essential for mitochondrial function.