The adnAB Locus, Encoding a Putative Helicase-Nuclease Activity, Is Essential in Streptomyces

Abstract

bHomologous recombination is a crucial mechanism that repairs a wide range of DNA lesions, including the most deleterious ones, double-strand breaks (DSBs). This multistep process is initiated by the resection of the broken DNA ends by a multisubunit helicase-nuclease complex exemplified by Escherichia coli RecBCD, Bacillus subtilis AddAB, and newly discovered Mycobacterium tuberculosis AdnAB. Here we show that in Streptomyces, neither recBCD nor addAB homologues could be detected. The only putative helicase-nuclease-encoding genes identified were homologous to M. tuberculosis adnAB genes. These genes are conserved as a single copy in all sequenced genomes of Streptomyces. The disruption of adnAB in Streptomyces ambofaciens and Streptomyces coelicolor could not be achieved unless an ectopic copy was provided, indicating that adnAB is essential for growth. Both adnA and adnB genes were shown to be inducible in response to DNA damage (mitomycin C) and to be independently transcribed. Introduction of S. ambofaciens adnAB genes in an E. coli recB mutant restored viability and resistance to UV light, suggesting that Streptomyces AdnAB could be a functional homologue of RecBCD and be involved in DNA damage resistance. Cells are under constant genotoxic pressure from both endogenous and exogenous sources. DNA damage needs to be repaired to avoid the formation of deleterious mutations, abortion of replication, and lethal chromosomal breakage. Homologous recombination (HR) is a crucial mechanism that repairs a variety of DNA lesions, including DNA double-strand breaks (DSBs), single-strand DNA gaps, and interstrand cross-links.DSBs are probably the most deleterious DNA damage that a cell can encounter. They are induced in cells by physical agents such as ionizing radiation or UV light, chemical agents, and natural products such as mitomycin C (MMC) or bleomycin. DSBs also result from replication fork collapse during chromosomal replication (1). The failure to repair DSBs can lead to cell death and, in the case of disrepair, can trigger large-scale chromosome rearrangements, favoring the generation of genetic diversity.In bacteria, DSBs are for the most part processed through HR, which requires a homologous DNA template to carry out faithful repair of the damaged DNA duplex. In intensively replicating cells (vegetative growth phase) or immediately after the passing of the replication fork, the sister chromatid can be used as an intact template. In nonreplicating phases, such as late stationary phase or in spores (which contain a single copy of the chromosome), DSBs are more likely repaired by an illegitimate repair pathway. Indeed, illegitimate recombination (IR) does not require an intact homologous sequence but as a consequence has reduced fidelity.The involvement of DSB repair by HR in a range of fundamental cellular processes (e.g., chromosome integrity, replication, segregation, etc.) reveals that HR is conserved in all living organisms. The initiating step of the repair mechanism consists of the resection of the DSB...

Diversity oriented biosynthesis via accelerated evolution of modular gene clusters

“…Recombination and repair in Streptomyces bacteria are relatively poorly understood 41 , 42 . It is our current hypothesis that an essential component of the AE process is localised genetic instability caused by activation of the pSG5 temperature sensitive replicon while still integrated into the genome.…”

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

“…However, recent studies that we performed suggest that Streptomyces has an atypical HR pathway. In fact, in contrast to bacterial models like Escherichia coli, Bacillus subtilis or Mycobacterium , deletion of genes encoding a potential helicase-nuclease complex supposed to be involved in the initial end resection step during HR is not viable in S. ambofaciens (Zhang et al, 2014). On the other hand, deletion of ruvABC and recG , two loci encoding factors that have a redundant role in migration and resolution of Holliday junction during late steps of HR, has only a mild effect on recombination efficiency (Hoff et al, 2016).…”

Bacillus subtilisRecO and SsbA are crucial for RecA-mediated recombinational DNA repair

“…Indeed, the co-expression of recBCD and recA genes of Serratia marcescens, Proteus mirabilis or Helicobacter pylori was necessary to suppress the DNA repair defect in E. coli recBCD cells ( 18 , 19 ). However, the expression of only B. subtilis AddAB or the Streptomyces ambofaciens AdnAB complex was sufficient to partially restore the recombination deficiency of the distantly related E. coli Δ recB or recBCD strain ( 20 , 21 ). In E. coli , in the absence of the RecBCD complex, end-processing requires the ‘activation’ of the RecJ–RecQ complex to promote end resection in the recBC sbcA or recBC sbcB sbcCD context.…”