GENETIC-REGULATION OF FRUCTOSYLTRANSFERASE IN STREPTOCOCCUS-MUTANS

Streptococcus mutans possesses several extracellular sucrose-metaboliz ing enzymes which have been implicated as important virulence factors in dental caries. This study was initiated to investigate the genetic regulation of one of these enzymes, the extracellular fructosyltransfe rase (Ftf). Fusions were constructed with the region upstream of the S . mutans GS5 Ftf gene (ftf) and a promoterless chloramphenicol acetylt ransferase (CAT) gene. The fusions were integrated at a remote site in the chromosome, and transcriptional activity in response to the addit ion of various carbohydrates to the growth medium was measured. A sign ificant increase in CAT activity was observed when glucose-groan cells were shifted to sucrose-containing medium. Sucrose-induced expression was repressed immediately upon addition of phosphoenolpyruvate phosph otransferase system sugars to the growth media. Deletion analysis of t he ftf upstream region revealed that an inverted repeat structure was involved in the control of ftf expression in response to carbohydrate. However, the control of the level of ftf transcription appeared to in volve a region distinct from that mediating carbohydrate regulation. C AT gene fusions also were constructed with the ftf upstream region fro m S. mutans V403, a fructan-hyperproducing strain which synthesizes in creased levels of Ftf. Sequence analysis of the upstream ftf region in this strain revealed several nucleotide sequence changes which were a ssociated with high-level ftf expression. Comparison of the GS5 and V4 03 ftf expression patterns suggested the presence of a trans-acting fa ctor(s) involved in modulation of ftf expression in response to carboh ydrate. This factor(s) was either absent or altered in V403, resulting in the inability of this organism to respond to the presence of carbo hydrate. The sequences of the ftf regions from three additional fructa n-hyperproducing strains were determined and compared with that of V40 3. Only one strain displayed nucleotide changes similar to those of V4 03. Two additional strains did not have these changes, suggesting that several mechanisms for up-regulation of ftf expression exist.