Global Gene Expression Analysis of Iron-Inducible Genes in Magnetospirillum magneticum AMB-1

Abstract

Iron uptake systems were identified by global expression profiling of Magnetospirillum magneticum AMB-1. feo, tpd, and ftr, which encode ferrous iron transporters, were up-regulated under iron-rich conditions. The concomitant rapid iron uptake and magnetite formation suggest that these uptake systems serve as iron supply lines for magnetosome synthesis.Iron is crucial in microbial metabolism. It exists in two redox states: the reduced Fe 2ϩ soluble ferrous state and the oxidized Fe 3ϩ ferric form, which is extremely insoluble (1). In magnetotactic bacteria, the formation of highly organized membranebound intracellular magnetites (Fe 3 O 4 ) or greigites (Fe 3 S 4 ) requires the acquisition of a large amount of iron several orders of magnitude greater than that required by Escherichia coli (5). Because of the large amount of iron to be transported, complex iron uptake systems that differ from known transport mechanisms in other microorganisms are expected to be operating in magnetotactic bacteria. Exceptional progress has been made in the isolation and characterization of functional genes and proteins involved in magnetosome synthesis (2,11,13,21,24), but at present, specific iron transport systems remain unidentified in magnetotactic bacteria.Here we report the identification of iron uptake systems by global expression profiling of the magnetotactic bacterium Magnetospirillum magneticum AMB-1 grown under iron-rich and iron-deficient conditions. Our results indicate that despite the unusual high-iron requirement of M. magneticum AMB-1, it utilizes robust but simple iron uptake systems similar to those of other gram-negative bacteria. This robust ferrous iron uptake suggests a significant contribution to magnetite synthesis. This study is the first to identify specific iron uptake systems in the complex iron metabolism of magnetotactic bacteria. The data presented here may facilitate future studies on the mechanism of magnetosome formation.To monitor iron uptake and magnetite formation, M. magneticum AMB-1 (ATCC 700264) was grown at 25°C under microaerobic conditions by sparging argon gas for 10 min into 500 ml of MSGM medium as previously described (5), with various iron concentrations of 0.1 to 300 M. All iron measurements were performed by atomic absorbance spectrophotometry. Extracellular iron concentrations were measured at different time points in cell-free culture supernatants. For intracellular iron measurements, cells were disrupted by lysozyme treatment (20) and ultracentrifuged at 100,000 ϫ g to separate the insoluble (magnetites) and soluble iron fractions.Iron was rapidly taken up in iron-rich cultures, and a corresponding increase of intracellular iron was observed within 10 min (Fig. 1A). Up to 70% of the initial iron concentration of the medium was taken up, and intracellular iron increased to 5,000 nmol/10 9 cells after 60 min. Insoluble iron in the cytoplasm, which mostly included magnetites, also increased within 10 min (Fig. 1B). These data indicate that the external iron was rapidly assimilate...

Rapid magnetosome formation shown by real-time x-ray magnetic circular dichroism

“…Of the total iron taken up by the cells, 75% is consumed in the first 15 min and 86% in the first 30 min. This result is supported by recent iron-uptake gene studies that also show significant cellular iron uptake within 10 min of induction, which is similar to the results presented here (19). The magnetic optical density (C mag ) is a qualitative measure of the cells' magnetization (20).…”

“…Rapid magnetosome formation appears correlated with respiratory growth under microaerobic conditions at low pO2; the slower magnetosome formation reported by Komeili was for bacteria grown anaerobically with nitrate as the terminal electron acceptor, and hence energy production would be lower. However, the secondary phase of maturation from small, immature magnetite crystals of 5-20 nm into perfect cubooctahedral particles, previously observed for both M. gryphiswaldense MSR-1 and M. magneticum AMB1 (17,19), appears much shorter in our studies.…”

“…Study of magnetosome formation in the same bacterium, M. magneticum AMB1, by Suzuki et al (19) showed that iron uptake was extremely rapid, with 60% of total uptake occurring within 10 min, with an intracellular concentration of iron Ͼ3,000 nmol/10 9 cells. By contrast, in the work by Komeili, the first rapid phase of iron uptake was only completed after 5 h. In both this and related work, including that of , magnetosome formation occurs concomitantly with iron uptake, and there is no evidence for a prior iron storage stage.…”

Formation of magnetite by bacteria and its application

“…To identify global gene expression profiles, especially those of iron uptake genes under different iron concentrations, transcriptome analysis through microarray imprinted with DNA from the whole genome sequence of AMB-1 was investigated (Suzuki et al 2006). Three ferrous and two ferric iron uptake systems encoded by specified genes were identified from the genome sequence, and two distinct patterns were observed in relation to the iron concentration in the growth medium and gene expression.…”

Whole genome sequence of Desulfovibrio magneticus strain RS-1 revealed common gene clusters in magnetotactic bacteria

“…The various crystal morphologies and compositions observed are species or strain dependent, implying a high degree of biological control (Spring and Schleifer 1995). Recent comprehensive molecular studies, including proteomic (Okamura et al 2000;Grünberg et al 2001Grünberg et al , 2004Arakaki et al 2003;Tanaka et al 2006), transcriptomic Suzuki et al 2006;Würdemann et al 2006), and whole-genomic analyses (Matsunaga et al 2005;Schübbe et al 2009), have revealed that the proteins associated with these particles play key roles in magnetosome biomineralization (Nakamura et al 1995;Arakaki et al 2003;Komeili et al 2006;Scheffel et al 2006). On the basis of the results of these studies, it has been described that the mechanism underlying magnetosome formation involves multiple processes including vesicle formation and alignment (Komeili et al 2006;Scheffel et al 2006), vesicular iron accumulation (Nakamura et al 1995), and iron-oxide crystallization (Arakaki et al 2003).…”