# DisA Limits RecG Activities at Stalled or Reversed Replication Forks
## 4 Discussion
coli_ but present in the Firmicutes, Actinobacteria, Fusobacteria and Termotogae phyla, is implicated in controlling the processing of a perturbed replication fork and contributes to genome integrity by stabilizing stalled or reversed replication forks. Why is relevant to limit fork processing? It seems that repair-by-recombination of a stalled fork is a two-edged sword. If a damaged template base stalls the replisome, and then the stalled fork is reversed, the RuvAB-RecU complex could catalyze HJ cleavage (Figure 5A,B). This would lead to the formation of a one-ended DSB, that should be lethal if an intact homologous template is absent and end resection function have not been synthesized yet, as at the early period of spore outgrowth [15; 16]. We propose that by limiting RecG-mediated fork reversal, DisA provides the timeframe to facilitate the removal of the DNA lesion. We show, however, that if RecG is pre-bound to a stalled or reversed fork, it becomes insensitive to DisA action (Figure 3), suggesting that DisA competes with RecG for binding to stalled or reversed forks. Earlier, we have proposed that DisA also limits RecA-mediated DNA strand exchange and template switching (Figure 5C) [24]. By doing so, DisA might facilitate that RecA protects the extruded strands from degradation, keeping chromosomal stability. Similarly, eukaryotic Rad51 promotes replication fork reversal and protects the extruded nascent strands from degradation [62]. In addition, Rad51 mediators and/or Rad51 itself in eukaryotes modulate fork stability by limiting SMARCAL1 [63; 64], in a way similar as here DisA does with RecG. This way, DisA might contribute to the election of the DDT sub pathway (translesion synthesis, template switching, fork reversal) to overcome replicative stress (Figure 5C). In summary, we illustrate how the DisA checkpoint may help to direct the flow of protein-DNA intermediates from a stalled fork to replication restart preventing fork nucleolytic degradation (Figure 5). Furthermore, by doing so, DisA might also help to control low error rate DDT pathways to maintain the balance between genetic stability and diversity. Some key questions that remain open for future studies will help us to define which branch migration translocase is involved in replication restart in _B. subtilis_. First, it will relevant to determine whether RecA, and perhaps DisA and RecG, contributes to the recruitment of the PriA preprimosomal protein at stalled forks, leading to the resumption of DNA replication. Finally, it will be relevant to determine how DisA coordinates other fork remodeling enzymes (e.g., RuvAB); and determine the interplay of RecG with other DNA helicases at replication-transcription conflicts. The presence of DisA also in non-spore forming bacteria indicates that it has a broader role in genome stability than just during spore revival. **Supplementary Materials:** The following are available online at [[https://www.mdpi.com/article/10.3390/cells10061357/s1](https://www.mdpi.com/article/10.3390/cells10061357/s1)]([https://www.mdpi.com/article/10.3390/cells10061357/s1](https://www.mdpi.com/article/10.3390/cells10061357/s1)), Table S1. Oligonucleotidetides used, Figure S1: DisA DAC activity at different ATP concentrations, Figure S2: DisA does not form a stable complex with RecG, Figure S3: The activities of the PrcA helicase are not regulated by DisA. **Author Contributions:** Conceptualization, R.T., S.A. and J.C.A. ; methodology, R.T., C.G., B.C. and I.B. ; performed the experiments, R.T., C.G., B.C., I.B. and S.A.; analyzed and interpreted the data, R.T., C.G., B.C., I.B, S.A. and J.C.A. ; supervision and funding, S.A. and J.C.A. ; drafted the manuscript, R.T., C.G., B.C., S.A. and J.C.A. ; and wrote the manuscript, J.C.A. All authors have read and agreed to the published version of the manuscript. **Funding:** This work was supported by the Ministerio de Ciencia e Innovacion, Agencia Estatal de Investigacion (MCIU/AEI)/FEDER PGC2018-097054-B-I00 to S.A. and J.C.A. **Institutional Review Board Statement:** Not applicable.