Promotion of presynaptic filament assembly by the ensemble of S. cerevisiae Rad51 paraloques with Rad52 - Bernstein - Pitt

Promotion of presynaptic filament assembly by the ensemble of S. cerevisiae Rad51 paraloques with Rad52

Rad55 bridges Csm2-Psy3 in the Shu complex with Rad51 in order to help loading Rad51 onto the ssDNA coated with RPA (presynaptic filament) => if the Rad55-Rad57-Csm2 interaction is disrupted, presynaptic filament cannot occur as well as HR.

Rad51 and Rad52 individually interact strongly with Rad55-Rad57 but weakly with the Shu complex. The role of Rad52 (F. 3e) is to enhance the Rad51 loading onto RPA-coated ssDNA in the presence of Shu complex and Rad55-Rad57. The RPA protein hinders the loading of Rad51 onto ssDNA when there are only Rad55-Rad57 and Rad52; but if the Shu complex is present, it enhances the Rad51 loading even though the Shu complex does not really interact with RPA. However, Csm2-Psy2 heterodimer possesses the DNA binding domain that may help the loading of Rad51 onto ssDNA(?). Then Rad54 promotes homology search and strand invasion and D-loop/DNA joint is resolved by different pathways to generate crossover or non-crossover product.

The Shu1 and Shu2 are not directly involved in presynaptic filament formation even though they are crucial in stalled DNA replication repair; Csm2-Psy2 can help this process as well as the whole Shu complex.

So even though Rad55 in the Rad55-Rad57 heterodimer was found to strongly interact with Csm2-Psy3, does Rad57 directly interact with the Csm2-Psy3 or Shu1 or Shu2  component of the Shu complex? Does Rad57 also interact with either or both Rad51 and Rad52? What happens to the presynaptic filament formation if Rad57 is absent and only Rad55 is present?

The RPA is loaded on ssDNA in order to promote the DNA replication, recombination, and repair in eukaryotes. What happens if  ssDNA is not coated by RPA because of a mutation in RPA, but the Rad51 is still able to be loaded on ssDNA in the presence of other factors except the Shu complex because there is no RPA, so is the Rad51 loading to ssDNA in this case able to rescue with the DNA repair in eukaryotes?

1. Even though Rad55 in the Rad55-Rad57 heterodimer was found to strongly interact with Csm2-Psy3, does Rad57 directly interact with the Csm2-Psy3 or Shu1 or Shu2  component of the Shu complex? Does Rad57 also interact with either or both Rad51 and Rad52? What happens to the presynaptic filament formation if Rad57 is absent and only Rad55 is present?

It is possible that Rad57 also directly interacts with Csm2.  We cannot rule out that possibility although I think it is likely only Rad55.  Shu1, Shu2, and Psy3 are not mediating interactions with Rad51, Rad55-Rad57, or Rad52.  I’m not sure if Rad57 interacts with Rad51 or Rad52 but I would imagine that it does.  Expression of Rad55 is depends on Rad57 and visa versa and so it is impossible in vivo or in vitro to determine what would happen to the presynaptic filament without either of them.

2. The RPA is loaded on ssDNA in order to promote the DNA replication, recombination, and repair in eukaryotes. What happens if  ssDNA is not coated by RPA because of a mutation in RPA, but the Rad51 is still able to be loaded on ssDNA in the presence of other factors except the Shu complex because there is no RPA, so is the Rad51 loading to ssDNA in this case able to rescue with the DNA repair in eukaryotes?

RPA is essential for viability and so it is impossible to eliminate RPA.  There are some temperature sensitive conditional mutants that exist (at least in yeast).  Rad51 readily loads onto ssDNA in the absence of RPA and does not require mediators.