Bacillus subtilis DinG/XPD-like paralogues, DinG and YpvA, have been implicated in overcoming replication stress. DinG possesses a DEDD exonuclease and DNA helicase domains, whereas YpvA lacks the DEDD exonuclease domain. We report that DinG·Mg²⁺ (hereafter referred to as DinG) degrades linear single-stranded (lss) DNA with 3′→5′ polarity and binds lssDNA with higher affinity than its exonuclease-deficient mutant DinG D10A E12A. DinG’s ssDNA-dependent ATPase activity neither stimulates nor inhibits DNA degradation. When bound to the 3′-end of forked DNA, DinG destabilises and degrades the substrate; however, in the presence of ATP, DinG dissociates before reaching the duplex junction. DinG degrades the RNA strand within RNA–DNA hybrids but does not cleave lssRNA unless complexed with Mn²⁺. DinG removes genomic R-loops, as RnhC and PcrA do. DinG physically interacts with RecA and PolA and functions in the same pathway as translesion synthesis (TLS) DNA polymerases (DNAPs) to respond to both spontaneous and methyl methanesulphonate (MMS)-induced mutagenesis. DinG-mGold forms spontaneous foci at or near replication forks, which become enriched following MMS or rifampicin treatment. We propose that DinG contributes to mitigating replication stress by degrading R-loop barriers and facilitating TLS, potentially via RecA-linked mechanisms.

Addressing the environmental impact of laboratory research provides a meaningful opportunity for workplace-level mitigation. However, in Spain, the importance of sustainability in research is only starting to be recognised at a national level. We developed a scalable, visual-oriented communications strategy that was designed with input from scientific staff and included a live seminar and a coherent set of graphic materials. Five pillars were targeted: three key areas of consumption (energy, water, and plastic use), procurement, and colleague engagement. The scheme was piloted in two of the largest biomedical research centres in Spain: the Severo Ochoa Molecular Biology Centre (CBM) and the Spanish National Centre for Biotechnology (CNB). The pilot engaged 50 laboratories and services across both centres, of which 34 were followed up for 5 months. The results indicate engagement with sustainability measures and shifts toward best practice across metrics, including equipment switch-off, water use, plastic item reuse, and purchasing decisions. During the follow-up period, 23% of participants adopted new measures related to freezer maintenance, decluttering, or sample organization, 55% removed old data from storage to save energy, 100% reduced their use of lifts, and 94% discussed research sustainability with their laboratory/group. The findings also revealed key areas for improvement, including the management of shared resources and resistance to change protocols. These results offer a wide range of quantitative and qualitative insights to guide further efforts and scaling at regional and national levels. Sustainability spotlight: This paper describes a scalable, visual-oriented communications strategy that can be used to promote sustainable behaviours in research laboratories. We present pilot results indicating engagement in sustainability and shifts toward best practice across metrics. Our work contributes to UN Sustainable Development Goal (SDG) 9 (industry and infrastructure) and SDG 12 (responsible consumption and production).

Natural chromosomal transformation (NCT) in Bacillus subtilis requires RecA and its accessory proteins including RecX and RecD2. Inactivation of the nucleoid-associated protein (NAP) Hbsu, Rok, or LrpC results in a dual effect: it enhances NCT but exacerbates the NCT defect of ΔrecX cells. Purified EbfC exhibits characteristic features of NAPs: it binds both single- and double-stranded DNA, protects them from degradation, and forms higher-order protein–DNA complexes via its DNA bridging activity. NCT is reduced upon EbfC inactivation but enhanced by ebfC overexpression. hbs55, Δrok, or ΔlrpC mutations suppress the NCT defect of ΔebfC, yet synergistically increase NCT upon ebfC overexpression. The NCT defect in ΔrecD2 cells is worsened by ebfC overexpression or by hbs55, Δrok, or ΔlrpC mutations. The nucleoid was more compacted in ΔebfC cells, an effect counteracted by hbs55, Δrok, or ΔlrpC mutations. EbfC contributes to DNA repair, and ebfC is epistatic to hbs or lrpC in response to DNA damage. We propose that chromosome folding, modulated by NAPs, plays a critical role in NCT and DNA repair. In this context, EbfC, by regulating nucleoid dynamics as a NAP, opposes the functions of Hbsu, Rok, and LrpC in NCT, while their interconnected roles contribute to DNA repair.