COMPETENCE-SPECIFIC INDUCTION OF RECA IS REQUIRED FOR FULL RECOMBINATION PROFICIENCY DURING TRANSFORMATION IN STREPTOCOCCUS-PNEUMONIAE

Transcriptional activation of the recA gene of Streptococcus pneumonia e was previously shown to occur at competence. A 5.7 kb recA-specific transcript that contained at least two additional genes, cinA and dinF , was identified. We now report the complete characterization of the r ecA operon and investigation of the role of the competence-specific in duction of recA. The 5.7 kb competence-specific recA transcript is sho wn to include lytA, which encodes the pneumococcal autolysin, a protei n previously shown to contribute to virulence of S. pneumoniae. Uncoup ling (denoted Ind(-)) of recA and/or the downstream genes was achieved through the placement of transcription terminators within the operon, either upstream or downstream of recA. Prevention of the competence-s pecific induction of recA severely affected spontaneous transformation . Transformation efficiencies of recA(+) (Ind(-)) and of wild-type cel ls were compared under various conditions and with different donor DNA . Chromosomal transformation was reduced 17-(chromosomal donor) to 45- fold (recombinant plasmid donor), depending on the donor DNA, and plas mid establishment was reduced 129-fold. Measurement of uptake of radio actively labelled donor DNA in transformed cells in parallel with scor ing for transformants (chromosomal donor) revealed normal uptake, but a 51-fold reduction in recombination in a recA(+) (Ind(-)) strain, ind icating that the transformation defect was primarily in recombination. Strikingly enough, a much larger (460-fold) reduction in recombinatio n was observed for the shortest homologous donor fragment used (878 nu cleotides long). possible interpretations of the observation that basa l RecA appears unable to promote efficient recombination whatever the number and the length of donor fragments taken up are proposed. The ro le of recA induction is discussed in view of the potential contributio n of transformation to genome plasticity in this pathogen.