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Published on: June 7, 2024
Species-specific and mutant MWFE proteins. Their effect on the assembly of a functional mammalian mitochondrial
Nagendra Yadava1, Prasanth Potluri, Erin N Smith
1Division of Biology and Center for Molecular Genetics, University of California, San Diego, La Jolla, California 92093-0322, USA.
Abstract:
The MWFE protein (70 amino acids) is highly conserved in evolution, but the human protein (80% identical to hamster) does not complement a null mutation in Chinese hamster cells. We have identified a small protein segment where significant differences exist between rodents and primates, illustrating very specifically the need for compatibility of the nuclear and mitochondrial genomes in the assembly of complex I. The segment between amino acids 39 and 46 appears to be critical for species-specific compatibility. Amino acid substitutions in this region were tested that caused a reduction of activity of the hamster protein or converted the inactive human protein into a partially active one. Such mutations could be useful in making mice with partial complex I activity as models for mitochondrial diseases. Their potential as dominant negative mutants was explored. More deleterious mutations in the NDUFA1 gene were also characterized. A conservative substitution, R50K, or a short C-terminal deletion makes the protein completely inactive. In the absence of MWFE, no high molecular weight complex was detectable by Blue Native-gel electrophoresis. The MWFE protein itself is unstable in the absence of assembled mitochondrially encoded integral membrane proteins of complex I.
Insights
The MWFE protein is crucial for complex I assembly, with a specific region (amino acids 39-46) critical for species compatibility between nuclear and mitochondrial genomes. Mutations in this region can model mitochondrial diseases.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- The MWFE protein is highly conserved but shows species-specific functional differences.
- Human MWFE does not complement hamster mutations, indicating a need for genome compatibility.
- Complex I assembly requires coordination between nuclear and mitochondrial genomes.
Purpose of the Study:
- Identify critical regions in MWFE responsible for species-specific compatibility.
- Investigate the role of MWFE in complex I assembly and mitochondrial disease modeling.
- Characterize mutations affecting MWFE function and complex I activity.
Main Methods:
- Comparative sequence analysis of MWFE across species.
- Site-directed mutagenesis to alter amino acids 39-46 in MWFE.
- Functional assays to assess complex I activity in hamster cells.
- Blue Native-gel electrophoresis to analyze complex I assembly.
- Characterization of deleterious mutations in the NDUFA1 gene.
Main Results:
- A critical segment (amino acids 39-46) was identified, crucial for rodent-primate compatibility in complex I assembly.
- Specific amino acid substitutions in this region modulated hamster MWFE activity and partially restored human MWFE function.
- Deleterious mutations, including R50K substitution or C-terminal deletion, rendered MWFE inactive.
- Absence of MWFE prevented high molecular weight complex I detection, and MWFE stability depended on other complex I subunits.
Conclusions:
- The MWFE protein segment (amino acids 39-46) is essential for species-specific compatibility in complex I assembly.
- Mutations in this region offer potential for creating mouse models of mitochondrial diseases with partial complex I activity.
- MWFE stability and function are intrinsically linked to the proper assembly of mitochondrially encoded subunits of complex I.
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