A programmed cell division delay preserves genome integrity during natural genetic transformation in Streptococcus pneumoniae.

Competence for genetic transformation is a differentiation program during which exogenous DNA is imported into the cell and integrated into the chromosome. In Streptococcus pneumoniae, competence develops transiently and synchronously in all cells during exponential phase, and is accompanied by a pause in growth. Here, we reveal that this pause is linked to the cell cycle. At least two parallel pathways impair peptidoglycan synthesis in competent cells. Single-cell analyses demonstrate that ComM, a membrane protein induced during competence, inhibits both initiation of cell division and final constriction of the cytokinetic ring. Competence also interferes with the activity of the serine/threonine kinase StkP, the central regulator of pneumococcal cell division. We further present evidence that the ComM-mediated delay in division preserves genomic integrity during transformation. We propose that cell division arrest is programmed in competent pneumococcal cells to ensure that transformation is complete before resumption of cell division, to provide this pathogen with the maximum potential for genetic diversity and adaptation.

Streptococcus pneumoniae, le transformiste.

Streptococcus pneumoniae (the pneumococcus) is an important human pathogen. Natural genetic transformation, which was discovered in this species, involves internalization of exogenous single-stranded DNA and its incorporation into the chromosome. It allows acquisition of pathogenicity islands and antibiotic resistance and promotes vaccine escape via capsule switching. This opinion article discusses how recent advances regarding several facets of pneumococcal transformation support the view that the process has evolved to maximize plasticity potential in this species, making the pneumococcus le transformiste of the bacterial kingdom and providing an advantage in the constant struggle between this pathogen and its host.

Midcell recruitment of the DNA uptake and virulence nuclease, EndA, for pneumococcal transformation.

Genetic transformation, in which cells internalize exogenous DNA and integrate it into their chromosome, is widespread in the bacterial kingdom. It involves a specialized membrane-associated machinery for binding double-stranded (ds) DNA and uptake of single-stranded (ss) fragments. In the human pathogen Streptococcus pneumoniae, this machinery is specifically assembled at competence. The EndA nuclease, a constitutively expressed virulence factor, is recruited during competence to play the key role of converting dsDNA into ssDNA for uptake. Here we use fluorescence microscopy to show that EndA is uniformly distributed in the membrane of noncompetent cells and relocalizes at midcell during competence. This recruitment requires the dsDNA receptor ComEA. We also show that under 'static' binding conditions, i.e., in cells impaired for uptake, EndA and ComEA colocalize at midcell, together with fluorescent end-labelled dsDNA (Cy3-dsDNA). We conclude that midcell clustering of EndA reflects its recruitment to the DNA uptake machinery rather than its sequestration away from this machinery to protect transforming DNA from extensive degradation. In contrast, a fraction of ComEA molecules were located at cell poles post-competence, suggesting the pole as the site of degradation of the dsDNA receptor. In uptake-proficient cells, we used Cy3-dsDNA molecules enabling expression of a GFP fusion upon chromosomal integration to identify transformed cells as GFP producers 60-70 min after initial contact between DNA and competent cells. Recording of images since initial cell-DNA contact allowed us to look back to the uptake period for these transformed cells. Cy3-DNA foci were thus detected at the cell surface 10-11 min post-initial contact, all exclusively found at midcell, strongly suggesting that active uptake of transforming DNA takes place at this position in pneumococci. We discuss how midcell uptake could influence homology search, and the likelihood that midcell uptake is characteristic of cocci and/or the growth phase-dependency of competence.