Comparative analysis of gene expression in Streptococcus pneumoniae and Lactococcus lactis

The pFL10 plasmid vector for translational fusions was constructed. pFL10 is based in the promiscuous pLS1 replicon and contains the pC194 cat gene deprived of its transcriptional promoter and Shine-Dalgarno (SD) sequence. Three promoters (Pcit, PpolA and PtetL) from Gram-positive bacteria, inserted in pFL10, were tested for their ability to drive transcription in Lactococcus lactis and Streptococcus pneumoniae. These promoters were coupled to the SD sequence of the lactococcal citP gene fused to the cat gene. Determination of the 5' ends of the three mRNAs by primer extension revealed the same start sites in both bacterial systems. However, it was observed a differential efficiency of promoter utilization by the RNA polymerases from the two hosts. The transcriptional behavior correlates with expression of the cat gene measured by determinations of chloramphenicol acetyltransferase (CAT) activity. Substitution of the SD of citP by that of the T7 phi 10 gene rendered a similar decrease of the CAT production in both bacterial systems.