Human RAD51 Protein Forms Amyloid-like Aggregates In Vitro .
Daniel V KachkinKirill V VolkovJulia V SopovaAlexander G BobylevSergey A FedotovSergei G Inge-VechtomovOxana V GalzitskayaYury O ChernoffAleksandr A RubelAnna Y AksenovaPublished in: International journal of molecular sciences (2022)
RAD51 is a central protein of homologous recombination and DNA repair processes that maintains genome stability and ensures the accurate repair of double-stranded breaks (DSBs). In this work, we assessed amyloid properties of RAD51 in vitro and in the bacterial curli-dependent amyloid generator (C-DAG) system. Resistance to ionic detergents, staining with amyloid-specific dyes, polarized microscopy, transmission electron microscopy (TEM), X-ray diffraction and other methods were used to evaluate the properties and structure of RAD51 aggregates. The purified human RAD51 protein formed detergent-resistant aggregates in vitro that had an unbranched cross-β fibrillar structure, which is typical for amyloids, and were stained with amyloid-specific dyes. Congo-red-stained RAD51 aggregates demonstrated birefringence under polarized light. RAD51 fibrils produced sharp circular X-ray reflections at 4.7 Å and 10 Å, demonstrating that they had a cross-β structure. Cytoplasmic aggregates of RAD51 were observed in cell cultures overexpressing RAD51 . We demonstrated that a key protein that maintains genome stability, RAD51, has amyloid properties in vitro and in the C-DAG system and discussed the possible biological relevance of this observation.
Keyphrases
- dna repair
- dna damage
- dna damage response
- endothelial cells
- electron microscopy
- high resolution
- amino acid
- oxidative stress
- stem cells
- magnetic resonance imaging
- genome wide
- single cell
- mass spectrometry
- bone marrow
- dna methylation
- optical coherence tomography
- ionic liquid
- induced pluripotent stem cells
- gene expression
- mesenchymal stem cells
- high speed