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1. The translesion polymerase Pol Y1 is a constitutive component of the B. subtilis replication machinery

   https://doi.org/10.1093/nar/gkae637  

   Marrin, McKayla E.; Foster, Michael R.; Santana, Chloe M.; Choi, Yoonhee; Jassal, Avtar S.; Rancic, Sarah J.; Greenwald, Carolyn R.; Drucker, Madeline N.; Feldman, Denholm T.; Thrall, Elizabeth S. (July 2024, Nucleic Acids Research)

   Abstract Unrepaired DNA damage encountered by the cellular replication machinery can stall DNA replication, ultimately leading to cell death. In the DNA damage tolerance pathway translesion synthesis (TLS), replication stalling is alleviated by the recruitment of specialized polymerases to synthesize short stretches of DNA near a lesion. Although TLS promotes cell survival, most TLS polymerases are low-fidelity and must be tightly regulated to avoid harmful mutagenesis. The gram-negative bacterium Escherichia coli has served as the model organism for studies of the molecular mechanisms of bacterial TLS. However, it is poorly understood whether these same mechanisms apply to other bacteria. Here, we use in vivo single-molecule fluorescence microscopy to investigate the TLS polymerase Pol Y1 in the model gram-positive bacterium Bacillus subtilis. We find significant differences in the localization and dynamics of Pol Y1 in comparison to its E. coli homolog, Pol IV. Notably, Pol Y1 is constitutively enriched at or near sites of replication in the absence of DNA damage through interactions with the DnaN clamp; in contrast, Pol IV has been shown to be selectively enriched only upon replication stalling. These results suggest key differences in the roles and mechanisms of regulation of TLS polymerases across different bacterial species. 

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2. The tardigrade Hypsibius exemplaris dramatically upregulates DNA repair pathway genes in response to ionizing radiation

   https://doi.org/10.1016/j.cub.2024.03.019  

   Clark-Hachtel, Courtney M.; Hibshman, Jonathan D.; De Buysscher, Tristan; Stair, Evan R.; Hicks, Leslie M.; Goldstein, Bob (May 2024, Current Biology)

   Tardigrades can survive remarkable doses of ionizing radiation, up to about 1,000 times the lethal dose for humans. How they do so is incompletely understood. We found that the tardigrade Hypsibius exemplaris suffers DNA damage upon gamma irradiation, but the damage is repaired. We show that this species has a specific and robust response to ionizing radiation: irradiation induces a rapid upregulation of many DNA repair genes. This upregulation is unexpectedly extreme—making some DNA repair transcripts among the most abundant transcripts in the animal. By expressing tardigrade genes in bacteria, we validate that increased expression of some repair genes can suffice to increase radiation tolerance. We show that at least one such gene is important in vivo for tardigrade radiation tolerance. We hypothesize that the tardigrades’ ability to sense ionizing radiation and massively upregulate specific DNA repair pathway genes may represent an evolved solution for maintaining DNA integrity. 

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3. A conserved bacterial genetic basis for commensal-host specificity

   https://doi.org/10.1126/science.adp7748  

   Gutiérrez-García, Karina; Aumiller, Kevin; Dodge, Ren; Obadia, Benjamin; Deng, Ann; Agrawal, Sneha; Yuan, Xincheng; Wolff, Richard; Zhu, Haolong; Hsia, Ru-Ching; et al (December 2024, Science)

   Animals selectively acquire specific symbiotic gut bacteria from their environments that aid host fitness. To colonize, a symbiont must locate its niche and sustain growth within the gut. Adhesins are bacterial cell surface proteins that facilitate attachment to host tissues and are often virulence factors for opportunistic pathogens. However, the attachments are often transient and nonspecific, and additional mechanisms are required to sustain infection. In this work, we use live imaging of individual symbiotic bacterial cells colonizing the gut of livingDrosophila melanogasterto show thatLactiplantibacillus plantarumspecifically recognizes the fruit fly foregut as a distinct physical niche.L. plantarumestablishes stably within its niche through host-specific adhesins encoded by genes carried on a colonization island. The adhesin binding domains are conserved throughout the Lactobacillales, and the island also encodes a secretion system widely conserved among commensal and pathogenic bacteria. 

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4. Elimination of transforming activity and gene degradation during UV and UV/H ₂ O ₂ treatment of plasmid-encoded antibiotic resistance genes

   https://doi.org/10.1039/c8ew00200b  

   Yoon, Younggun; Dodd, Michael C.; Lee, Yunho (January 2018, Environmental Science: Water Research & Technology)

   null (Ed.)

   To better understand the elimination of transforming activity of antibiotic resistance genes (ARGs), this study investigated the deactivation of transforming activity of an ARG (in Escherichia coli as a host) and ARG degradation (according to quantitative PCR [qPCR] with different amplicon sizes) during UV (254 nm) and UV/H 2 O 2 treatments of plasmid pUC19 containing an ampicillin resistance gene ( amp R ). The required UV fluence for each log 10 reduction of the transforming activity during UV treatment was ∼37 mJ cm −2 for both extra- and intra-cellular pUC19 (the latter within E. coli ). The resulting fluence-based rate constant ( k ) of ∼6.2 × 10 −2 cm 2 mJ −1 was comparable to the k determined previously for transforming activity loss of plasmids using host cells capable of DNA repair, but much lower (∼10-fold) than that for DNA repair-deficient cells. The k value for pUC19 transforming activity loss was similarly much lower than the k calculated for cyclobutane-pyrimidine dimer (CPD) formation in the entire plasmid. These results indicate the significant role of CPD repair in the host cells. The degradation rate constants ( k ) of amp R measured by qPCR increased with increasing target amplicon size (192–851 bp) and were close to the k calculated for the CPD formation in the given amplicons. Further analysis of the degradation kinetics of plasmid-encoded genes from this study and from the literature revealed that qPCR detected most UV-induced DNA damage. In the extracellular plasmid, DNA damage mechanisms other than CPD formation ( e.g. , base oxidation) were detectable by qPCR and gel electrophoresis, especially during UV/H 2 O 2 treatment. Nevertheless, the enhanced DNA damage for the extracellular plasmids did not result in faster elimination of the transforming activity. Our results indicate that calculated CPD formation rates and qPCR analyses are useful for predicting and/or measuring the rate of DNA damage and predicting the efficiency of transforming activity elimination for plasmid-encoded ARGs during UV-based water disinfection and oxidation processes. 

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5. RnhP is a plasmid‐borne RNase HI that contributes to genome maintenance in the ancestral strain Bacillus subtilis NCIB 3610

   https://doi.org/10.1111/mmi.14601  

   Nye, Taylor M.; McLean, Emma K.; Burrage, Andrew M.; Dennison, Devon D.; Kearns, Daniel B.; Simmons, Lyle A. (September 2020, Molecular Microbiology)

   Abstract RNA‐DNA hybrids form throughout the chromosome during normal growth and under stress conditions. When left unresolved, RNA‐DNA hybrids can slow replication fork progression, cause DNA breaks, and increase mutagenesis. To remove hybrids, all organisms use ribonuclease H (RNase H) to specifically degrade the RNA portion. Here we show that, in addition to chromosomally encoded RNase HII and RNase HIII,Bacillus subtilisNCIB 3610 encodes a previously uncharacterized RNase HI protein, RnhP, on the endogenous plasmid pBS32. Like other RNase HI enzymes, RnhP incises Okazaki fragments, ribopatches, and a complementary RNA‐DNA hybrid. We show that while chromosomally encoded RNase HIII is required for pBS32 hyper‐replication, RnhP compensates for the loss of RNase HIII activity on the chromosome. Consequently, loss of RnhP and RNase HIII impairs bacterial growth. We show that the decreased growth rate can be explained by laggard replication fork progression near the terminus region of the right replichore, resulting in SOS induction and inhibition of cell division. We conclude that all three functional RNase H enzymes are present inB. subtilisNCIB 3610 and that the plasmid‐encoded RNase HI contributes to chromosome stability, while the chromosomally encoded RNase HIII is important for chromosome stability and plasmid hyper‐replication. 

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