Finding new functions for the Ugp and PitA transport systems of Escherichia coli K-12
Abstract
Escherichia coli K-12 has two major systems for transporting inorganic phosphate (Pi). The high-affinity phosphate specific transport (Pst) system is an ABC transporter that is required for both P i uptake and Pi regulation of the Pho regulon. The low affinity Pi transporter (PitA) is a single transmembrane protein that does not belong to the Pho regulon. Besides, E. coli can utilize organic phosphate transported by several organic phosphate transporters which include Ugp system for transporting glycerol-6-phosphate. Here we discovered that under certain conditions PitA is required for proper assembly or functioning of the Pst transporter. Whereas a non-polar single-gene deletion of pstS (encoding the periplasmic P i binding protein) can be fully complemented by a single or multi-copy plasmid encoding PstS, a ΔpstS ΔpitA mutant cannot. Rather, a ΔpitA Δ pstS mutant can be restored to normal regulation only when genes for the entire pstSCAB region are expressed together from a single or multi-copy plasmid. These results provide the first evidence for a functional interaction between the PitA and PstSCAB transporters. By use of pitAp- and pitBp-lacZ transcriptional fusions, we show that these promoters are inducible under conditions of P i limitation in a PhoR/PhoB independent manner. Our data contradict the interpretations that PstS is not essential for Pi regulation of the Pho regulon (S. M. Hoffer, J. Tommassen, J. Bacteriol. 183: 5768-5783) and that PhoB negatively controls pitB expression (R. M. Harris, D. C. Webb, S. M. Howitt, G. B. Cox, J. Bacteriol. 183: 5008-5014). Unexpectedly, deleting genes for the PhoR/PhoB two-component system (TCS) resulted in synthetic lethality in a pstSCAB-phoU pitA pitB triple mutant due to an inability to assimilate inorganic or organic phosphate sources. Further experiments showed that growth of a pstSCAB-phoU pitA pitB mutant on organic or inorganic phosphates requires the Ugp system, whose expression is under PhoR/PhoB control. These data provide genetic evidence for uptake of Pi by the Ugp system. The finding that the Ugp system itself is capable of Pi uptake calls into question earlier evidence for control of Pho regulon gene expression, and the ugp operon in particular by internal phosphate.
Degree
Ph.D.
Advisors
Wanner, Purdue University.
Subject Area
Molecular biology
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