Non-canonical regulation of Fructose metabolism by FruR in Vibrio cholerae

Abstract

In most bacterial species, fructose uptake is mediated by the fructose-specific phosphoenolpyruvate (PEP)-dependent carbohydrate phosphotransferase (PTS Fru ).The PTS Fru consists of FruA, membrane-spanning protein that is responsible for fructose transport, and FruB that delivers a phosphate group from PEP to FruA, which finally phosphorylates the fructose during its transportation across the membrane. While most PTS sugars are phosphorylated at their 6-position, fructose is phosphorylated at 1-position. Fructose 1-phosphate (F1P) is then converted to fructose 1,6-bisphosphate (FBP) by 1-phosphofructokinase (FruK). In bacteria belonging to Gammaproteobacteria including Escherichia coli, Pseudomonas putida and Vibrio cholerae, FruA, FruB and FruK are encoded in a single operon (fruBKA). It has been known that, in most bacteria possessing the fruBKA operon such as E. coli and P. putida, transcription of the fruBKA operon is negatively regulated by the fructose repressor (FruR) which binds to the cognate operator(s) located downstream of the transcription start site (TSS) of the operon and F1P is the inducer of this operon. However, this study reveals that the FruR-mediated transcriptional regulation of the fruBKA operon in V. cholerae is completely different and more sophisticated than that of E. coli and P. putida. In V. cholerae, FruR is essential for the transcription of the fruBKA operon and the growth on fructose. FruR directly binds to three cognate operators located at the intergenic region between the gene coding for FruR itself and the fruBKA operon. Among these operators, the operator centered at 20.5 bp upstream of the TSS of the fruBKA operon is essential but the other two operators are dispensable for the activation of the fruBKA transcription and thus the growth of V. cholerae on fructose. Furthermore, the FruR-mediated transcriptional activation requires an intracellular F1P. The exogenous expression of FruR mutants defective in F1P recognition could not complement the growth defect of a fruR mutant on fructose. The binding properties of the FruR to the operators are altered by F1P so that FruR enables the RNA polymerase to activate the transcription of the fruBKA operon. Taken together, in this study shows the non-canonical regulation of the fruBKA expression by FruR in V. cholerae.