PUBLICATION
Rad54 Phosphorylation Promotes Homologous Recombination by Balancing Rad54 Mobility and DNA Binding
- Authors
- Lengert, N., Spies, J., Drossel, B.
- ID
- ZDB-PUB-200611-8
- Date
- 2019
- Source
- Biophysical journal 116: 1406-1419 (Journal)
- Registered Authors
- Keywords
- none
- MeSH Terms
-
- Protein Structure, Quaternary
- Adenosine Triphosphatases/metabolism
- DNA-Binding Proteins/chemistry
- DNA-Binding Proteins/metabolism*
- Protein Domains
- Homologous Recombination
- Animals
- Rad51 Recombinase/metabolism
- Movement*
- Protein Binding
- Phosphorylation
- Zebrafish
- Models, Molecular
- Humans
- DNA Helicases/chemistry
- DNA Helicases/metabolism*
- Protein Multimerization
- DNA/genetics*
- DNA/metabolism*
- PubMed
- 30961891 Full text @ Biophys. J.
Citation
Lengert, N., Spies, J., Drossel, B. (2019) Rad54 Phosphorylation Promotes Homologous Recombination by Balancing Rad54 Mobility and DNA Binding. Biophysical journal. 116:1406-1419.
Abstract
The repair of DNA double-strand breaks by homologous recombination is of crucial importance for maintaining genomic stability. Two major players during this repair pathway are Rad51 and Rad54. Previously, it was shown that Rad54 exists as a monomer or oligomer when bound to DNA and drives the displacement of Rad51 by translocating along the DNA. Moreover, phosphorylation of Rad54 was reported to stimulate this clearance of Rad51 from DNA. However, it is currently unclear how phosphorylation of Rad54 modulates its molecular-structural function and how it affects the activity of monomeric or oligomeric Rad54 during the removal of Rad51. To examine the impact of Rad54 phosphorylation on a molecular-structural level, we applied molecular dynamics simulations of Rad54 monomers and hexamers in the absence or presence of DNA. Our results suggest that 1) phosphorylation of Rad54 stabilizes the monomeric form by reducing the interlobe movement of Rad54 monomers and might therefore facilitate multimer formation around DNA and 2) phosphorylation of Rad54 in a higher-order hexamer reduces its binding strength to DNA, which is a requirement for efficient mobility on DNA. To further address the relationship between the mobility of Rad54 and its phosphorylation state, we performed fluorescence recovery after photobleaching experiments in living cells, which expressed different versions of the Rad54 protein. Here, we could measure that the phosphomimetic version of Rad54 was highly mobile on DNA, whereas a nonphosphorylatable mutant displayed a mobility defect. Taken together, these data show that the phosphorylation of Rad54 is a critical event in balancing the DNA binding strength and mobility of Rad54 and might therefore provide optimal conditions for DNA translocation and subsequent removal of Rad51 during homologous recombination repair.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping