Costs and benefits of natural transformation in Acinetobacter baylyi

Abstract

Background: Natural transformation enables acquisition of adaptive traits and drives genome evolution in prokaryotes. Yet, the selective forces responsible for the evolution and maintenance of natural transformation remain elusive since taken-up DNA has also been hypothesized to provide benefits such as nutrients or templates for DNA repair to individual cells.

Results: We investigated the immediate effects of DNA uptake and recombination on the naturally competent bacterium Acinetobacter baylyi in both benign and genotoxic conditions. In head-to-head competition experiments between DNA uptake-proficient and -deficient strains, we observed a fitness benefit of DNA uptake independent of UV stress. This benefit was found with both homologous and heterologous DNA and was independent of recombination. Recombination with taken-up DNA reduced survival of transformed cells with increasing levels of UV-stress through interference with nucleotide excision repair, suggesting that DNA strand breaks occur during recombination attempts with taken-up DNA. Consistent with this, we show that absence of RecBCD and RecFOR recombinational DNA repair pathways strongly decrease natural transformation.

Conclusions: Our data show a physiological benefit of DNA uptake unrelated to recombination. In contrast, recombination during transformation is a strand break inducing process that represents a previously unrecognized cost of natural transformation.

Keywords: Bacterial evolution; Competence; DNA repair; DprA; Horizontal gene transfer; Natural transformation.

Fig. 1

Relative fitness of wildtype strain (a) and its ∆dprA derivative (b) grown in head-to-head competitions against the DNA uptake-deficient ∆comB-F mutant. The competitions were treated with a combination of an initial UV pulse of 216 J m⁻² or no UV irradiation with a subsequent amendment of no DNA (DNase), homologous DNA or heterologous DNA. In each group n = 13. In the control competitions with no UV and no DNA both the wildtype and its ∆dprA derivative displayed a reduced fitness relative to KOM130 of w = 0.88 (0.83–0.92, 95% CI) and w = 0.86 (0.82–0.91, 95% CI), respectively, due to the biological cost of type IV pilus expression (Additional file 1: Fig. S1). Error bars indicate 95% confidence intervals. Significant differences between the treatment groups (pooled results of both UV- and UV+ competitions) are indicated by asterisks (** P < 0.001, *** P < 0.0001). A two-way ANOVA of all mean values is provided in (Additional file 1: Table S4)

Fig. 2

Relative survival of the pooled total population of the wildtype (closed triangles and bold solid line, n = 16) and the transformant fractions obtained with either pSBP1 DNA (0.1 μg ml⁻¹; open circles with dotted line, n = 10) or with homologous genomic DNA (2 μg ml⁻¹; open squares with solid line, n = 6) with increasing levels of UV irradiation. We also determined the relative survival of the total population (closed triangles and bold dashed line, n = 10) and the transformant fraction in the ∆uvrA mutant obtained with homologous DNA (2 μg ml⁻¹; open squares and dashed line, n = 10). Error bars denote the 95% confidence intervals. The transformation frequencies without UV irradiation were (7.7 ± 1.4) × 10⁻⁴ (pSBP1 DNA) and (2.5 ± 2.8) × 10⁻² (genomic DNA) for the wildtype; and (1.9 ± 1.0) × 10⁻² (genomic DNA) for the ∆uvrA mutant. Mean initial titers (CFU/ml) for the wildtype and the ∆uvrA mutant were 1.8 × 10⁷ and 4.2 × 10⁷, respectively

Fig. 3

Formation of DNA double strand-breaks conferred by nucleotide excision repair during natural transformation. a: Grey lines: genomic DNA; grey spikes: UV-induced lesions. Black line: taken-up DNA single-strand with genetic marker (oval). b: Generation of DNA single-strand-breaks by UvrAB-directed, UvrBC-initiated cleavage of the damage-containing strand upstream and downstream of the lesions, and by RecA-mediated strand invasion of the taken-up DNA and cleavage of the displaced DNA strand. c: Partial repair of single-strand breaks: 1. by removal of the lesion-containing single-strand fragment (left lesion), fill-in synthesis, and covalent ligation (dark grey; catalysed by UvrD, DNA polymerase I, and DNA ligase, respectively); and 2. by covalent joining of the invaded taken-up strand with the genomic DNA at one side (downstream of the marker). Single-strand breaks persist when the UV lesion-containing strands are not removed (right lesion) and when an invaded DNA strand remains unligated at one end (upstream of the marker). One-sided ligation of the invaded DNA strand is common (see text). Following DNA replication (indicated by a replication fork approaching from the left end), single-strand breaks are converted into potentially lethal double-strand breaks

Fig. 4

Natural transformation frequencies of A. baylyi wildtype and DNA recombination-impaired mutants. Transformation frequencies were obtained in liquid transformation experiments with genomic homologous DNA (0.1 μg ml⁻¹) containing a kanamycin resistance marker gene and are given as means with 95% confidence intervals. The initial recipient titers are listed in (Additional file 1: Table S4)