In the
dental caries pathogen Streptococcus mutans,
phosphotransacetylase (Pta) and
acetate kinase (Ack) convert
pyruvate into
acetate with the concomitant generation of
ATP. The genes for this pathway are tightly regulated by multiple environmental and intracellular inputs, but the basis for differential expression of the genes for Pta and Ack in S. mutans had not been investigated. Here, we show that inactivation in S. mutans of ccpA or codY reduced the activity of the ackA promoter, whereas a ccpA mutant displayed elevated pta promoter activity. The interactions of CcpA with the promoter regions of both genes were observed using electrophoretic mobility shift and
DNase protection assays. CodY bound to the ackA promoter region but only in the presence of
branched-chain amino acids (BCAAs).
DNase footprinting revealed that the upstream region of both genes contains two catabolite-responsive elements (cre1 and cre2) that can be bound by CcpA. Notably, the cre2 site of ackA overlaps with a CodY-binding site. The CcpA- and CodY-binding sites in the promoter region of both genes were further defined by site-directed mutagenesis. Some differences between the reported consensus CodY binding site and the region protected by S. mutans CodY were noted. Transcription of the pta and ackA genes in the ccpA mutant strain was markedly different at low pH relative to transcription at neutral pH. Thus, CcpA and CodY are direct regulators of transcription of ackA and pta in S. mutans that optimize
acetate metabolism in response to
carbohydrate,
amino acid availability, and environmental pH.IMPORTANCE The human
dental caries pathogen Streptococcus mutans is remarkably adept at coping with extended periods of
carbohydrate limitation during fasting periods. The
phosphotransacetylase-
acetate kinase (Pta-Ack) pathway in S. mutans modulates
carbohydrate flux and fine-tunes the ability of the organisms to cope with stressors that are commonly encountered in the oral cavity. Here, we show that CcpA controls transcription of the pta and ackA genes via direct interaction with the promoter regions of both genes and that
branched-chain amino acids (BCAAs), particularly
isoleucine, enhance the ability of CodY to bind to the promoter region of the ackA gene. A working model is proposed to explain how regulation of pta and ackA genes by these allosterically controlled regulatory
proteins facilitates proper
carbon flow and energy production, which are essential functions during
infection and pathogenesis as
carbohydrate and
amino acid availability continually fluctuate.