COORDINATED REGULATION OF THE EIIMAN AND FRURKI OPERONS OF STREPTOCOCCUS MUTANS BY GLOBAL AND FRUCTOSE-SPECIFIC PATHWAYS

The glucose/mannose-PTS permease EIIMan encoded by manLMN in the dental caries pathogen Streptococcus mutans has a dominant influence on sugar-specific, CcpA-independent catabolite repression (CR). Mutations in manL affect energy metabolism and virulence-associated traits, including biofilm formation, acid tolerance and competence. Using promoter:reporter fusions, expression of the manLMN and the fruRKI operons, encoding a transcriptional regulator, a fructose-1-P kinase and a fructose-PTS permease EIIFru, respectively, was monitored in response to carbohydrate source and in mutants lacking CcpA, FruR, and components of EIIMan. Expression of genes for EIIMan and EIIFru was directly regulated by CcpA and CR, as evinced by in vivo and in vitro methods. Unexpectedly, not only was the fruRKI operon negatively regulated by FruR, but so was manLMN. Carbohydrate transport by EIIMan had a negative influence on expression of manLMN, but not fruRKI. In agreement with the proposed role of FruR in regulating these PTS operons, loss of fruR or fruK substantially altered growth on a number of carbohydrates, including fructose. RNA deep sequencing revealed profound changes in gene regulation caused by deletion of fruK or fruR. Collectively, these findings demonstrate intimate interconnection of the regulation of two major PTS permeases in S. mutans and reveal novel and important contributions of fructose metabolism to global regulation of gene expression. Overall design: mRNA profiling of the wild type UA159 and two mutants fruK-13 and fruR, each grown to exponential phase in batch on a rich medium BHI.