FurC Regulates Expression of zupT for the Central Zinc Importer ZupT of Cupriavidus metallidurans

Abstract

The zinc importer ZupT is required for the efficient allocation of zinc to zinc-dependent proteins in the metal-resistant bacterium Cupriavidus metallidurans but not for zinc import per se. The expression of zupT is upregulated under conditions of zinc starvation. C. metallidurans contains three members of the Fur family of regulators that qualify as candidates for the zupT regulator. The expression of a zupT-lacZ reporter gene fusion was strongly upregulated in a ⌬furC mutant but not in a ⌬furA or ⌬furB mutant. Expression of the genes for transition-metal importers (pitA, corA1, corA2, and corA3) was not changed in this pattern in all three ⌬fur mutants, but they were still downregulated under conditions of elevated zinc concentrations, indicating the presence of another zinc-dependent regulator. FurA was a central regulator of the iron metabolism in C. metallidurans, and furA was constitutively expressed under the conditions tested. Expression of furB was upregulated under conditions of iron starvation, and FurB could be an iron starvation Fur connecting general metal and iron homeostasis, as indicated by the phenotype of a ⌬furB ⌬furC double mutant. FurC was purified as a Strep-tagged protein and retarded the electrophoretic mobility of a DNA fragment upstream of zupT. Binding of FurC to this operator region was influenced by the presence of zinc ions and EDTA. Thus, FurC is the main zinc uptake regulator (Zur) of C. metallidurans and represses synthesis of the central zinc importer ZupT when sufficient zinc is present. Cupriavidus metallidurans is able to sustain its cellular transition-metal homeostasis even in the presence of mM metal ion concentrations and even in the presence of a mixture of them. This is achieved by highly redundant metal uptake systems, which have only minimal cation selectivity. These uptake systems are in combination with metal efflux systems, which remove surplus cations (1). The most sophisticated efflux systems and batteries of helper proteins are encoded by metal resistance determinants on the two native plasmids of wild-type strain CH34, pMOL28 and pMOL30, but a variety of resistance determinants also are located on the two chromosomes of this betaproteobacterium of the order Burkholderiales (2, 3).

Interplay between seven secondary metal uptake systems is required for full metal resistance of Cupriavidus metallidurans

“…by up-regulation of iron import pathways via FurA or FurB. 17 Third, while deletion of zntB from D5 (leading to D6) had no significant effect on the metal content ( Fig. 2), deletion of hoxN in D6 (leading to D7) increased the magnesium content, restored the iron and copper content to the higher and the zinc content to the lower level of the D4 strain.…”

Interplay between the Zur Regulon Components and Metal Resistance in Cupriavidus metallidurans

“…This means that CobW3 may decrease zinc uptake by interfering with the minimally nine other zinc uptake systems that C. metallidurans possesses in addition to ZupT (24,25). While expression of genes for efflux systems is not strongly influenced by CobW3 (see results in the supplemental material) and those encoding uptake systems, with the exception of zupT, are influenced by zinc only at a low level (28,29), CobW3 may act upon the activity of metal transport systems, e.g., ZntB, CorA1, and CorA3. CobW3-like proteins are also widespread in Burkholderiales, here mostly in Cupriavidus species; however, most of the 2,951 entries found still contained the GCXCC motif, and many have no histidine-rich stretch at the carboxy terminus and retain the amino acid stretch following the switch region ( Fig.…”

Zinc Homeostasis at the Bacteria/Host Interface—From Coordination Chemistry to Nutritional Immunity

“…It sustains proper zinc homeostasis even in the presence of m m concentrations of this metal and may also ensure the proper metal homeostasis of all other ions . The efflux systems of C. metallidurans include, besides the mentioned ZneB, CzcD, and ZntA specific mainly for Zn II , also, for example, PbrA, specific for Pb II , or CnrR, specific for Ni II …”