Mechanistic flexibility as a conserved theme across 3 billion years of nonhomologous DNA end-joining.

DNA double-strand breaks (DSB) represent the most deleterious form of DNA damage. Two mechanistically distinctive repair pathways have evolved to mend these breaks in eukaryotes: homologous recombination (HR) and nonhomologous DNA end-joining (NHEJ) (Lieber et al. 2003). HR is restricted to late S or G2 of the cell cycle, whereas NHEJ can function throughout the cell cycle and is the primary repair pathway for DSBs. NHEJ is also distinctive for the mechanistic flexibility of the nucleases, polymerases, and ligases in eukaryotes (Lieber 2007; Lieber et al. 2007). Initially, NHEJ was thought to be restricted to eukaryotes because the best-studied prokaryote, Escherichia coli, has no ability to join DNA ends (if it did, molecular cloning would have had a different history) (Table 1). Only when bioinformatists discovered a distantly diverged Ku-like gene in diverse prokaryotic genomes did researchers begin to realize the existence of a similar NHEJ pathway in bacteria (Aravind and Koonin 2001; Doherty et al. 2001; d’Adda di Fagagna et al. 2003). The bacterial Ku homolog is suspected to form a homodimer with a conjectured structure similar to the ring-shaped eukaryotic Ku heterodimer (Weller et al. 2002). The gene for an ATP-dependent ligase named LigD was often found to be adjacent to the Ku gene on the bacterial chromosome (Aravind and Koonin 2001; Doherty et al. 2001; Weller and Doherty 2001). This linkage between Ku and an ATP-dependent ligase prompted more extensive studies and later defined a bacterial NHEJ pathway. Table 1. Possible corresponding components between NHEJ in prokaryotes and eukaryotes So why do some but not all bacteria retain this DNA repair pathway? Bacterial NHEJ is nonessential under rapid proliferation conditions because HR is active and a duplicate genome is present to provide homology donors (Pitcher et al. 2007a; Shuman and Glickman 2007). However, those bacteria that retain the NHEJ pathway spend much of their life cycle in stationary phase, at which point HR is not available for DSB repair for lack of homology donors. In addition, in nature, desiccation and dry heat are two naturally occurring physical processes that produce substantial numbers of DSBs in bacteria. Hence, bacterial Ku and LigD are present in bacterial species that often form endospores, because during sporulation, NHEJ is efficient for the DSB repair (Pitcher et al. 2007a). In many bacterial species, unlike the eukaryotic NHEJ ligase IV, LigD is a large, multidomain protein that contains three components within a single polypeptide: a polymerase (POL) domain, a phosphoesterase (PE) domain, and a ligase (LIG) domain (Shuman and Glickman 2007). Therefore, it is tempting to suggest that bacterial Ku and LigD are necessary and sufficient to repair all the DSBs generated in vivo (Della et al. 2004). However, in this issue of Genes & Development, the study by Aniukwu et al. (2008) from the Shuman and Glickman laboratories describes extensive genetic studies combined with in vivo DSB repair analysis in Mycobacterium smegmatis and clearly demonstrates the complexity and flexibility of the NHEJ pathway in bacteria. They uncover a faithful NHEJ pathway specifically for 3′ overhang DSB repair, which is Ku- and LigD-independent. Their results also show that the structure of the broken ends determines the pathway and outcome of the DSB repair.