A conserved lysine residue of plant Whirly proteins is necessary for higher order protein assembly and protection against DNA damage.

All organisms have evolved specialized DNA repair mechanisms in order to protect their genome against detrimental lesions such as DNA double-strand breaks. In plant organelles, these damages are repaired either through recombination or through a microhomology-mediated break-induced replication pathway. Whirly proteins are modulators of this second pathway in both chloroplasts and mitochondria. In this precise pathway, tetrameric Whirly proteins are believed to bind single-stranded DNA and prevent spurious annealing of resected DNA molecules with other regions in the genome. In this study, we add a new layer of complexity to this model by showing through atomic force microscopy that tetramers of the potato Whirly protein WHY2 further assemble into hexamers of tetramers, or 24-mers, upon binding long DNA molecules. This process depends on tetramer-tetramer interactions mediated by K67, a highly conserved residue among plant Whirly proteins. Mutation of this residue abolishes the formation of 24-mers without affecting the protein structure or the binding to short DNA molecules. Importantly, we show that an Arabidopsis Whirly protein mutated for this lysine is unable to rescue the sensitivity of a Whirly-less mutant plant to a DNA double-strand break inducing agent.

Figure: Structural comparison of the K67A variant with the non-mutated WHY2 protein. (A) Comparison of the two crystal forms of WHY2 K67A with the non-mutated protein. The RMSD for superimposing the crystal forms I and II of WHY2 K67A onto the non-mutated protein are 0.7Å and 1.1Å, respectively. (B) Comparison of the dT32-bound form of WHY2 K67A with the dT32-bound non-mutated protein. The RMSD for superimposing WHY2 K67A onto the non-mutated protein is 0.2Å.

Results from:
Cappadocia L, Parent JS, Zampini E, Lepage E, Sygusch J, Brisson N.
Nucleic Acids Res. 2012 Jan;40(1):258-69. doi: 10.1093/nar/gkr740. Epub 2011 Sep 12.
PMID: 21911368 [PubMed - indexed for MEDLINE]
PMCID: PMC3245945