Restriction-like phenomena in transformation of Bacillus subtilis recA

Abstract

Genetic transformation in recAl strains of Bacillus subtilis was studied to test the hypothesis that, in these strains, a major pathway of recombination is missing, leaving only residual transformation via a pathway specific for transduction. The two putative recombinational pathways have been hypothesized to differ in either length of synapsed regions or specificity for nucleotide sequence homology. It was found that the efficiency of transformation of recAl cells by deoxyribonucleic acid (DNA) from the heterologous strain W23 was much lower than when a homologous donor DNA was used, the relative efficiency being different for different genetic markers. Because the frequency of recombination between linked markers is only slightly changed in recAl recipients, and because markers of heterologous origin in DNA from intergenotic strains are not discriminated against strongly by recAl recipients, it is concluded that neither a difference in length of synapsed DNA nor a difference in specificity for nucleotide sequence homology accounts for reduced transformation in recAl cells. It is proposed that at some time between uptake and integration, heterologous DNA is inactivated by restriction, and that aberrant restriction of repaired regions may account for reduced transformation by homologous DNA.

Repair and subsequent fragmentation of deoxyribonucleic acid in ultraviolet-irradiated Bacillus subtilis recA

“…The experiments described above indicate that the pleiotropic effects of the recAl mutation in B. subtilis are probably due to a defect in DNA repair that leads to fragmentation of the genome. That defect must also account for the reduction in transformation and transduction in recAl strains (9,11,13,14). It is not likely that a defect in post-replication repair causes the fragmentation, because neither star- 16,509 26,244 a Values are total 3H counts per minute recovered in sucrose gradients used to calculate strand-break induction for Fig.…”

“…Perhaps the restriction system acting after irradiation or transformation in recA cells is a similarly induced function, explaining the apparent lack of faulty restriction in normally growing recA cells. However, restriction enzymes make double-strand breaks in DNA (1, 5), which are generally lethal, and there is no evidence to date that restriction-like effects in transformation of recA cells are lethal, although restrictable sequences seem to be integrated (11). It may be that in the recA cells only a single strand is attacked and that a modified complementary sequence does not protect a restrictable site.…”

Capacity for postreplication repair correlated with transducibility in Rec- mutants of Bacillus subtilis

“…Cells were frozen in SGA medium containing glycerol (final concentration, 15%) and stored at -60°C. The preparation of cultures for studies of repair resynthesis has been described (19,20).…”

Effect of 6-(p-hydroxyphenylazo)-uracil on the homologous and heterologous transduction processes in Bacillus subtilis

“…One could thus surnise that the homologous transduction process enters the rec pathway at some stage after that catalyzed by the products of genes A and G. No mutant with the opposite behavior (altered in homologous transduction and not in heterologous) has ever been observed. Also the hypothesis that a restriction-like phenomenon is at the basis of the phenotype of recA mutants has been suggested (9).…”