Complex Iron Uptake by the Putrebactin-Mediated and Feo Systems in Shewanella oneidensis

Abstract

is an extensively studied bacterium capable of respiring minerals, including a variety of iron ores, as terminal electron acceptors (EAs). Although iron plays an essential and special role in iron respiration of , little has been done to date to investigate the characteristics of iron transport in this bacterium. In this study, we found that all proteins encoded by the cluster for putrebactin ( native siderophore) synthesis (PubABC), recognition-transport of Fe-putrebactin across the outer membrane (PutA), and reduction of ferric putrebactin (PutB) are essential to putrebactin-mediated iron uptake. Although homologs of PutA are many, none can function as its replacement, but some are able to work with other bacterial siderophores. We then showed that Fe-specific Feo is the other primary iron uptake system, based on the synthetical lethal phenotype resulting from the loss of both iron uptake routes. The role of the Feo system in iron uptake appears to be more critical, as growth is significantly impaired by the absence of the system but not of putrebactin. Furthermore, we demonstrate that hydroxyl acids, especially α-types such as lactate, promote iron uptake in a Feo-dependent manner. Overall, our findings underscore the importance of the ferrous iron uptake system in metal-reducing bacteria, providing an insight into iron homeostasis by linking these two biological processes. is among the first- and the best-studied metal-reducing bacteria, with great potential in bioremediation and biotechnology. However, many questions regarding mechanisms closely associated with those applications, such as iron homeostasis, including iron uptake, export, and regulation, remain to be addressed. Here we show that Feo is a primary player in iron uptake in addition to the siderophore-dependent route. The investigation also resolved a few puzzles regarding the unexpected phenotypes of the mutant and lactate-dependent iron uptake. By elucidating the physiological roles of these two important iron uptake systems, this work revealed the breadth of the impacts of iron uptake systems on the biological processes.

Free Rather Than Total Iron Content Is Critically Linked to the Fur Physiology in Shewanella oneidensis

“…A large majority of TonB-dependent siderophore receptors (TDSRs) were produced in substantially increased amounts, consistent with the findings that they are induced under iron starvation conditions as well as in the fur mutant (Andrews et al, 2003;Fu et al, 2018). These receptors have been proposed to have roles in iron uptake via various siderophores produced by other bacteria in the environment (Liu et al, 2018). In line with this, proteins associated with siderophore-iron uptake, such as FbpABC, were also increasingly produced in the fur mutant.…”

“…and FeoB polypeptides) is a transport system essential for ferrous iron uptake (Dong et al, 2017;Liu et al, 2018;Wang et al, 2020). As a result, a strain lacking putA and feoAB genes ( putA feo) could survive and grow only when hemin is supplemented as an iron source.…”

“…We then turned to iron uptake. In S. oneidensis, iron uptake is predominantly mediated by the ferrous iron uptake system (Feo) (Liu et al, 2018; Figure 1). According to the proteomic data, in the fur mutant the Feo system was modestly up-regulated, FeoA and FeoB, 1.27-and 1.89-fold, respectively, suggesting that this system may not be functionally compromised by the Fur loss.…”

“…Next to the Feo system is the siderophore-dependent system for iron uptake in S. oneidensis (Dong et al, 2017). This system is composed of enzymes for biosynthesis of siderophores (PubABC) and proteins for their transport (PutA) and reduction (PutB, reductase), all of which are functionally essential (Liu et al, 2018;Wang et al, 2020; Figure 1). Among them, enzymes for siderophore synthesis (PubABC, 0.96, 0.36, and 0.48-fold, respectively) were less abundant than in the wildtype, as revealed by the proteomic analysis (Supplementary Table S1).…”

“…To test this idea, effects of iron chelators, to which the cell membrane is permeable and impermeable, on the fur mutant were compared. We reasoned that the influence of desferrioxamine (DFO), a commercially available siderophore that cannot be imported into S. oneidensis cells and thus functions to abruptly terminate iron uptake (Liu et al, 2018), should be different from that of cell-permeant 2-2-dipyridyl, which is potentially able to lower free iron in the cytoplasm. At 2 and 5 µM, DFO decreased cyt c biosynthesis (whitened cell pellets, Supplementary Figure S5) and iron content of the wild-type significantly while the effect was not evident at lower concentrations (Figures 7A,B).…”