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Nucleoid localization of Hsp40 Mdj1 is important for its function in maintenance of mitochondrial DNA
Grzegorz L Ciesielski1, Magdalena Plotka, Mateusz Manicki
1Department of Molecular and Cellular Biology, University of Gdansk, Gdansk, Poland.
Abstract:
Faithful replication and propagation of mitochondrial DNA (mtDNA) is critical for cellular respiration. Molecular chaperones, ubiquitous proteins involved in protein folding and remodeling of protein complexes, have been implicated in mtDNA transactions. In particular, cells lacking Mdj1, an Hsp40 co-chaperone of Hsp70 in the mitochondrial matrix, do not maintain functional mtDNA. Here we report that the great majority of Mdj1 is associated with nucleoids, DNA-protein complexes that are the functional unit of mtDNA transactions. Underscoring the importance of Hsp70 chaperone activity in the maintenance of mtDNA, an Mdj1 variant having an alteration in the Hsp70-interacting J-domain does not maintain mtDNA. However, a J-domain containing fragment expressed at the level that Mdj1 is normally present is not competent to maintain mtDNA, suggesting a function of Mdj1 beyond that carried out by its J-domain. Nevertheless, loss of mtDNA function upon Mdj1 depletion is retarded when the J-domain, is overexpressed. Analysis of Mdj1 variants revealed a correlation between nucleoid association and DNA maintenance activity, suggesting that localization is functionally important. We found that Mdj1 has DNA binding activity and that variants retaining DNA-binding activity also retained nucleoid association. Together, our results are consistent with a model in which Mdj1, tethered to the nucleoid via DNA binding, thus driving a high local concentration of the Hsp70 machinery, is important for faithful DNA maintenance and propagation.
Insights
Mitochondrial DNA (mtDNA) maintenance relies on the Hsp40 co-chaperone Mdj1, which binds to DNA and concentrates Hsp70 machinery at nucleoids for faithful DNA replication and propagation.
Area of Science:
- Mitochondrial biology
- Molecular chaperones
- DNA replication and maintenance
Background:
- Faithful replication and propagation of mitochondrial DNA (mtDNA) are essential for cellular respiration.
- Molecular chaperones, including Hsp40 co-chaperones like Mdj1, are involved in mtDNA transactions.
- Cells lacking Mdj1, an Hsp70 co-chaperone in the mitochondrial matrix, fail to maintain functional mtDNA.
Purpose of the Study:
- To investigate the role of Mdj1 in mtDNA maintenance.
- To determine the functional importance of Mdj1's association with nucleoids and its DNA-binding activity.
- To elucidate the mechanism by which Mdj1 contributes to mtDNA integrity.
Main Methods:
- Localization studies of Mdj1 within mitochondria.
- Analysis of Mdj1 variants with alterations in the J-domain and DNA-binding activity.
- Assessment of mtDNA maintenance in cells with depleted or overexpressed Mdj1 and its variants.
- DNA binding assays for Mdj1.
Main Results:
- The majority of Mdj1 localizes to nucleoids, the functional units of mtDNA transactions.
- An Mdj1 variant lacking Hsp70-interacting J-domain function cannot maintain mtDNA.
- Mdj1 exhibits DNA-binding activity, and variants retaining this activity are associated with nucleoids.
- Overexpression of the J-domain partially rescues mtDNA loss upon Mdj1 depletion.
Conclusions:
- Mdj1's association with nucleoids, mediated by DNA binding, is crucial for its function in mtDNA maintenance.
- Mdj1 likely functions by tethering to nucleoids, concentrating the Hsp70 machinery to ensure faithful mtDNA maintenance and propagation.
- The J-domain alone is insufficient for Mdj1's full function in mtDNA maintenance.
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