Efficient processing of abasic sites by bacterial nonhomologous end-joining Ku proteins

Abstract

Intracellular reactive oxygen species as well as the exposure to harsh environmental conditions can cause, in the single chromosome of Bacillus subtilis spores, the formation of apurinic/apyrimidinic (AP) sites and strand breaks whose repair during outgrowth is crucial to guarantee cell viability. Whereas double-stranded breaks are mended by the nonhomologous end joining (NHEJ) system composed of an ATP-dependent DNA Ligase D (LigD) and the DNA-end-binding protein Ku, repair of AP sites would rely on an AP endonuclease or an AP-lyase, a polymerase and a ligase. Here we show that B. subtilis Ku (BsuKu), along with its pivotal role in allowing joining of two broken ends by B. subtilis LigD (BsuLigD), is endowed with an AP/deoxyribose 5′-phosphate (5′-dRP)-lyase activity that can act on ssDNA, nicked molecules and DNA molecules without ends, suggesting a potential role in BER during spore outgrowth. Coordination with BsuLigD makes possible the efficient joining of DNA ends with near terminal abasic sites. The role of this new enzymatic activity of Ku and its potential importance in the NHEJ pathway is discussed. The presence of an AP-lyase activity also in the homolog protein from the distantly related bacterium Pseudomonas aeruginosa allows us to expand our results to other bacterial Ku proteins.

Multiple and Variable NHEJ-Like Genes Are Involved in Resistance to DNA Damage in Streptomyces ambofaciens

“…However, no clear trend toward the co-occurrence of these two “variable” genes can be noticed in the 38 analysed genomes. In B. subtilis , the recent involvement of LigD Bsub and Ku Bsub in base excision repair along with their major role in NHEJ supports the third hypothesis (de Ory et al, 2014, 2016). …”

NHEJ enzymes LigD and Ku participate in stationary-phase mutagenesis in Pseudomonas putida

“…The idea that bacterial NHEJ enzymes could have more interaction partners is supported by the recent findings that Ku from mycobacteria and B. subtilis can recruit also other proteins than LigD [24,[70][71][72][73]. Moreover, the presence of an AP-lyase activity was recently demonstrated in Ku protein of B. subtilis and P. aeruginosa, suggesting its additional roles in DNA repair [74].…”

Bacterial NHEJ: a never ending story

“…Its function could be to reduce putative negative effects of this threading inside the chromosome on other DNA repair processes. The regulation of the threading of Ku may also be important for the recruitment of LigD near, or at the DSB site, ensuring an efficient NHEJ repair since LigD alone is a poorly efficient ligase without its Ku partner in vitro (Weller et al , ; Della et al , ; Zhu and Shuman, ; de Ory et al , ; McGovern et al , ). The function of Ku in the NHEJ could be more proactive than previously anticipated by allowing the bridging of DNA ends, thanks to its extended C‐ter (McGovern et al , ), an ability shared by the human Ku (Ramsden and Gellert, ).…”

“…Human Ku, besides its function as a scaffolder of the NHEJ machinery, is also an AP‐lyase suggested to be involved in the repair of AP site near DSBs (Roberts et al , ). This activity is conserved in Ku from B. subtilis and P. aeruginosa (de Ory et al , ). As Ku Bsub is able to thread inward DNA molecules (McGovern et al , ), via loading on their broken ends, this could facilitate the repair of abasic sites surrounding a DSB.…”

“…Remarkably, Ku Bsub is also able to linearize a supercoiled plasmid containing AP‐sites (de Ory et al , ). This counterintuitive effect for a protein intended to repair DSB is supposed to be dependent on its AP‐lyase activity and its ability to bind end‐free dsDNA molecules, DSB arising by the nicking of phosphodiester bonds of proximal AP‐sites on opposite strands of a dsDNA molecules containing numerous such lesions (de Ory et al , ).…”