Magnetosome-inspired synthesis of soft ferrimagnetic nanoparticles for magnetic tumor targeting.

Magnetic targeting is one of the most promising approaches for improving the targeting efficiency by which magnetic drug carriers are directed using external magnetic fields to reach their targets. As a natural magnetic nanoparticle (MNP) of biological origin, the magnetosome is a special "organelle" formed by biomineralization in magnetotactic bacteria (MTB) and is essential for MTB magnetic navigation to respond to geomagnetic fields. The magnetic targeting of magnetosomes, however, can be hindered by the aggregation and precipitation of magnetosomes in water and biological fluid environments due to the strong magnetic attraction between particles. In this study, we constructed a magnetosome-like nanoreactor by introducing MTB Mms6 protein into a reverse micelle system. MNPs synthesized by thermal decomposition exhibit the same crystal morphology and magnetism (high saturation magnetization and low coercivity) as natural magnetosomes but have a smaller particle size. The DSPE-mPEG-coated magnetosome-like MNPs exhibit good monodispersion, penetrating the lesion area of a tumor mouse model to achieve magnetic enrichment by an order of magnitude more than in the control groups, demonstrating great prospects for biomedical magnetic targeting applications.

Sesbanimide R, a Novel Cytotoxic Polyketide Produced by Magnetotactic Bacteria.

Genomic information from various magnetotactic bacteria suggested that besides their common ability to form magnetosomes, they potentially also represent a source of bioactive natural products. By using targeted deletion and transcriptional activation, we connected a large biosynthetic gene cluster (BGC) of the trans-acyltransferase polyketide synthase (trans-AT PKS) type to the biosynthesis of a novel polyketide in the alphaproteobacterium Magnetospirillum gryphiswaldense Structure elucidation by mass spectrometry and nuclear magnetic resonance spectroscopy (NMR) revealed that this secondary metabolite resembles sesbanimides, which were very recently reported from other taxa. However, sesbanimide R exhibits an additional arginine moiety the presence of which reconciles inconsistencies in the previously proposed sesbanimide biosynthesis pathway observed when comparing the chemical structure and the potential biochemistry encoded in the BGC. In contrast to the case with sesbanimides D, E, and F, we were able to assign the stereocenter of the arginine moiety experimentally and two of the remaining three stereocenters by predictive biosynthetic tools. Sesbanimide R displayed strong cytotoxic activity against several carcinoma cell lines.IMPORTANCE The findings of this study contribute a new secondary metabolite member to the glutarimide-containing polyketides. The determined structure of sesbanimide R correlates with its cytotoxic bioactivity, characteristic for members of this family. Sesbanimide R represents the first natural product isolated from magnetotactic bacteria and identifies this highly diverse group as a so-far-untapped source for the future discovery of novel secondary metabolites.

Magnetospirillum magneticum as a Living Iron Chelator Induces TfR1 Upregulation and Decreases Cell Viability in Cancer Cells.

Interest has grown in harnessing biological agents for cancer treatment as dynamic vectors with enhanced tumor targeting. While bacterial traits such as proliferation in tumors, modulation of an immune response, and local secretion of toxins have been well studied, less is known about bacteria as competitors for nutrients. Here, we investigated the use of a bacterial strain as a living iron chelator, competing for this nutrient vital to tumor growth and progression. We established an in vitro co-culture system consisting of the magnetotactic strain Magnetospirillum magneticum AMB-1 incubated under hypoxic conditions with human melanoma cells. Siderophore production by 108 AMB-1/mL in human transferrin (Tf)-supplemented media was quantified and found to be equivalent to a concentration of 3.78 µM ± 0.117 µM deferoxamine (DFO), a potent drug used in iron chelation therapy. Our experiments revealed an increased expression of transferrin receptor 1 (TfR1) and a significant decrease of cancer cell viability, indicating the bacteria's ability to alter iron homeostasis in human melanoma cells. Our results show the potential of a bacterial strain acting as a self-replicating iron-chelating agent, which could serve as an additional mechanism reinforcing current bacterial cancer therapies.

Fusion expression of nanobodies specific for the insecticide fipronil on magnetosomes in Magnetospirillum gryphiswaldense MSR-1.

BACKGROUND: Magnetic nanoparticles such as magnetosomes modified with antibodies allow a high probability of their interaction with targets of interest. Magnetosomes biomineralized by magnetotactic bacteria are in homogeneous nanoscale size and have crystallographic structure, and high thermal and colloidal stability. Camelidae derived nanobodies (Nbs) are small in size, thermal stable, highly water soluble, easy to produce, and fusible with magnetosomes. We aimed to functionalize Nb-magnetosomes for the analysis of the insecticide fipronil. RESULTS: Three recombinant magnetotactic bacteria (CF, CF+ , and CFFF) biomineralizing magnetosomes with different abundance of Nbs displayed on the surface were constructed. Compared to magnetosomes from the wild type Magnetospirillum gryphiswaldense MSR-1, all of the Nb-magnetosomes biosynthesized by strains CF, CF+ , and CFFF showed a detectable level of binding capability to fipronil-horseradish peroxidase (H2-HRP), but none of them recognized free fipronil. The Nb-magnetosomes from CFFF were oxidized with H2O2 or a glutathione mixture consisting of reduced glutathione and oxidized glutathione in vitro and their binding affinity to H2-HRP was decreased, whereas that to free fipronil was enhanced. The magnetosomes treated with the glutathione mixture were employed to develop an enzyme-linked immunosorbent assay for the detection of fipronil in water samples, with average recoveries in a range of 78-101%. CONCLUSIONS: The economical and environmental-friendly Nb-magnetosomes biomineralized by the bacterial strain MSR-1 can be potentially applied to nanobody-based immunoassays for the detection of fipronil or nanobody-based assays in general.

OxyR controls magnetosome formation by regulating magnetosome island (MAI) genes, iron metabolism, and redox state.

Magnetospirillum gryphiswaldense MSR-1 uses chains of magnetosomes, membrane-enveloped magnetite (Fe(II)Fe(III)2O4) nanocrystals, to align along magnetic field. The process of magnetosome biomineralization requires a precise biological control of redox conditions to maintain a balanced amounts of ferric and ferrous iron. Here, we identified functions of the global regulator OxyR (MGMSRv2_4250, OxyR-4250) in MSR-1 during magnetosome formation. OxyR deletion mutant ΔoxyR-4250 displayed reduced magnetic response, and increased levels of intracellular ROS (reactive oxygen species). OxyR-4250 protein upregulated expression of six antioxidant genes (ahpC1, ahpC2, katE, katG, sodB, trxA), four iron metabolism-related regulator genes (fur, irrA, irrB, irrC), a bacterioferritin gene (bfr), and a DNA protection gene (dps). OxyR-4250 was shown, for the first time, to directly regulate magnetosome island (MAI) genes mamGFDC, mamXY, and feoAB1 operons. Taken together, our findings indicate that OxyR-4250 helps maintain a proper redox environment for magnetosome formation by eliminating excess ROS, regulating iron homeostasis and participating in regulation of Fe2+/Fe3+ ratio within the magnetosome vesicle through regulating MAI genes.

Magnetotactic Bacteria Accumulate a Large Pool of Iron Distinct from Their Magnetite Crystals.

Magnetotactic bacteria (MTB) are ubiquitous aquatic microorganisms that form intracellular nanoparticles of magnetite (Fe3O4) or greigite (Fe3S4) in a genetically controlled manner. Magnetite and greigite synthesis requires MTB to transport a large amount of iron from the environment. Most intracellular iron was proposed to be contained within the crystals. However, recent mass spectrometry studies suggest that MTB may contain a large amount of iron that is not precipitated in crystals. Here, we attempted to resolve these discrepancies by performing chemical and magnetic assays to quantify the different iron pools in the magnetite-forming strain Magnetospirillum magneticum AMB-1, as well as in mutant strains showing defects in crystal precipitation, cultivated at various iron concentrations. All results show that magnetite represents at most 30% of the total intracellular iron under our experimental conditions and even less in the mutant strains. We further examined the iron speciation and subcellular localization in AMB-1 using the fluorescent indicator FIP-1, which was designed for the detection of labile Fe(II). Staining with this probe suggests that unmineralized reduced iron is found in the cytoplasm and associated with magnetosomes. Our results demonstrate that, under our experimental conditions, AMB-1 is able to accumulate a large pool of iron distinct from magnetite. Finally, we discuss the biochemical and geochemical implications of these results.IMPORTANCE Magnetotactic bacteria (MTB) produce iron-based intracellular magnetic crystals. They represent a model system for studying iron homeostasis and biomineralization in microorganisms. MTB sequester a large amount of iron in their crystals and have thus been proposed to significantly impact the iron biogeochemical cycle. Several studies proposed that MTB could also accumulate iron in a reservoir distinct from their crystals. Here, we present a chemical and magnetic methodology for quantifying the iron pools in the magnetotactic strain AMB-1. Results showed that most iron is not contained in crystals. We then adapted protocols for the fluorescent Fe(II) detection in bacteria and showed that iron could be detected outside crystals using fluorescence assays. This work suggests a more complex picture for iron homeostasis in MTB than previously thought. Because iron speciation controls its fate in the environment, our results also provide important insights into the geochemical impact of MTB.

Light regulation of resistance to oxidative damage and magnetic crystal biogenesis in Magnetospirillum magneticum mediated by a Cys-less LOV-like protein.

Light-oxygen-voltage (LOV) proteins are ubiquitous photoreceptors that can interact with other regulatory proteins and then mediate their activities, which results in cellular adaptation and subsequent physiological changes. Upon blue-light irradiation, a conserved cysteine (Cys) residue in LOV covalently binds to flavin to form a flavin-Cys adduct, which triggers a subsequent cascade of signal transduction and reactions. We found a group of natural Cys-less LOV-like proteins in magnetotactic bacteria (MTB) and investigated its physiological functions by conducting research on one of these unusual LOV-like proteins, Amb2291, in Magnetospirillum magneticum. In-frame deletion of amb2291 or site-directive substitution of alanine-399 for Cys mutants impaired the protective responses against hydrogen peroxide, thereby causing stress and growth impairment. Consequently, gene expression and magnetosome formation were affected, which led to high sensitivity to oxidative damage and defective phototactic behaviour. The purified wild-type and A399C-mutated LOV-like proteins had similar LOV blue-light response spectra, but Amb2291A399C exhibited a faster reaction to blue light. We especially showed that LOV-like protein Amb2291 plays a role in magnetosome synthesis and resistance to oxidative stress of AMB-1 when this bacterium was exposed to red light and hydrogen peroxide. This finding expands our knowledge of the physiological function of this widely distributed group of photoreceptors and deepens our understanding of the photoresponse of MTB. KEY POINTS: • We found a group of Cys-less light-oxygen-voltage (LOV) photoreceptors in magnetotactic bacteria, which prompted us to study the light-response and biological roles of these proteins in these non-photosynthetic bacteria. • The Cys-less LOV-like protein participates in the light-regulated signalling pathway and improves resistance to oxidative damage and magnetic crystal biogenesis in Magnetospirillum magneticum. • This result will contribute to our understanding of the structural and functional diversity of the LOV-like photoreceptor and help us understand the complexity of light-regulated model organisms.

Single-cell determination of iron content in magnetotactic bacteria: implications for the iron biogeochemical cycle.

Magnetotactic bacteria (MTB) are ubiquitous aquatic microorganisms that mineralize dissolved iron into intracellular magnetic crystals. After cell death, these crystals are trapped into sediments that remove iron from the soluble pool. MTB may significantly impact the iron biogeochemical cycle, especially in the ocean where dissolved iron limits nitrogen fixation and primary productivity. A thorough assessment of their impact has been hampered by a lack of methodology to measure the amount of, and variability in, their intracellular iron content. We quantified the iron mass contained in single MTB cells of Magnetospirillum magneticum strain AMB-1 using a time-resolved inductively coupled plasma-mass spectrometry methodology. Bacterial iron content depends on the external iron concentration, and reaches a maximum value of ~10-6 ng of iron per cell. From these results, we calculated the flux of dissolved iron incorporation into environmental MTB populations and conclude that MTB may mineralize a significant fraction of dissolved iron into crystals.

A gradient-forming MipZ protein mediating the control of cell division in the magnetotactic bacterium Magnetospirillum gryphiswaldense.

Cell division needs to be tightly regulated and closely coordinated with other cellular processes to ensure the generation of fully viable offspring. Here, we investigate division site placement by the cell division regulator MipZ in the alphaproteobacterium Magnetospirillum gryphiswaldense, a species that forms linear chains of magnetosomes to navigate within the geomagnetic field. We show that M. gryphiswaldense contains two MipZ homologs, termed MipZ1 and MipZ2. MipZ2 localizes to the division site, but its absence does not cause any obvious phenotype. MipZ1, by contrast, forms a dynamic bipolar gradient, and its deletion or overproduction cause cell filamentation, suggesting an important role in cell division. The monomeric form of MipZ1 interacts with the chromosome partitioning protein ParB, whereas its ATP-dependent dimeric form shows non-specific DNA-binding activity. Notably, both the dimeric and, to a lesser extent, the monomeric form inhibit FtsZ polymerization in vitro. MipZ1 thus represents a canonical gradient-forming MipZ homolog that critically contributes to the spatiotemporal control of FtsZ ring formation. Collectively, our findings add to the view that the regulatory role of MipZ proteins in cell division is conserved among many alphaproteobacteria. However, their number and biochemical properties may have adapted to the specific needs of the host organism.

Effects of Environmental Conditions on High-Yield Magnetosome Production by Magnetospirillum gryphiswaldense MSR-1

Background: Magnetotactic bacteria are a heterogeneous group of Gram-negative prokaryote cells that produce linear chains of magnetic particles called magnetosomes, intracellular organelles composed of magnetic iron particles. Many important applications have been defined for magnetic nanoparticles in biotechnology, such as cell separation applications, as well as acting as carriers of enzymes, antibodies, or anti-cancer drugs. Since the bacterial growth is difficult and the yield of magnetosome production is low, the application of magnetosome has not been developed on a commercial scale. Methods: Magnetospirillum gryphiswaldense strain MSR-1 was used in a modified current culture medium supplemented by different concentrations of oxygen, iron, carbon, and nitrogen, to increase the yield of magnetosomes. Results: Our improved MSR-1 culture medium increased magnetosome yield, magnetosome number per bacterial cell, magnetic response, and bacterial cell growth yield significantly. The yield of magnetosome increased approximately four times. The optimized culture medium containing 25 mM of Na-pyruvate, 40 mM of NaNO3, 200 µM of ferrous sulfate, and 5-10 ppm of dissolved oxygen (DO) resulted in 186.67 mg of magnetosome per liter of culture medium. The iron uptake increased significantly, and the magnetic response of the bacteria to the magnetic field was higher than threefold as compared to the previously reported procedures. Conclusion: This technique not only decreases the cultivation time but also reduces the production cost. In this modified method, the iron and DO are the major factors affecting the production of magnetosome by M. gryphiswaldense strain MSR-1. However, refining this technique will enable a further yield of magnetosome and cell density.

The Potential of Intrinsically Magnetic Mesenchymal Stem Cells for Tissue Engineering.

The magnetization of mesenchymal stem cells (MSC) has the potential to aid tissue engineering approaches by allowing tracking, targeting, and local retention of cells at the site of tissue damage. Commonly used methods for magnetizing cells include optimizing uptake and retention of superparamagnetic iron oxide nanoparticles (SPIONs). These appear to have minimal detrimental effects on the use of MSC function as assessed by in vitro assays. The cellular content of magnetic nanoparticles (MNPs) will, however, decrease with cell proliferation and the longer-term effects on MSC function are not entirely clear. An alternative approach to magnetizing MSCs involves genetic modification by transfection with one or more genes derived from Magnetospirillum magneticum AMB-1, a magnetotactic bacterium that synthesizes single-magnetic domain crystals which are incorporated into magnetosomes. MSCs with either or mms6 and mmsF genes are followed by bio-assimilated synthesis of intracytoplasmic magnetic nanoparticles which can be imaged by magnetic resonance (MR) and which have no deleterious effects on MSC proliferation, migration, or differentiation. The stable transfection of magnetosome-associated genes in MSCs promotes assimilation of magnetic nanoparticle synthesis into mammalian cells with the potential to allow MR-based cell tracking and, through external or internal magnetic targeting approaches, enhanced site-specific retention of cells for tissue engineering.

Biosynthesis of Thermoresponsive Magnetic Nanoparticles by Magnetosome Display System.

Thermoresponsive magnetic nanoparticles (MNPs) were synthesized using a magnetosome display system. An elastin-like polypeptide decamer of VPGVG (ELP10), which is hydrophobic above the transition temperature ( Tt) and can form an insoluble aggregation, was immobilized on biogenic MNPs in the magnetotactic bacterium, Magnetospirillum magneticum AMB-1. It was suggested that hydrophobicity of the MNP surface increased at 60 °C compared with 20 °C by the immobilization of ELP10. Size distribution analysis indicated that the immobilization of ELP10 onto MNPs induced the increased hydrophobicity with increasing temperatures up to 60 °C, promoting aggregation of the particles by hydrophobic and magnetic interactions. These results suggest that the acceleration of magnetic collection at 60 °C was caused by particle aggregation promoted by hydrophobic interaction between ELP-MNPs. Furthermore, the immobilization of ELP on MNPs gave a quick magnetic collection at 60 °C by external magnetic field. The thermoresponsive properties will further expand the utility of biotechnological applications of biogenic MNPs.

In Vivo Coating of Bacterial Magnetic Nanoparticles by Magnetosome Expression of Spider Silk-Inspired Peptides.

Magnetosomes are natural magnetic nanoparticles with exceptional properties that are synthesized in magnetotactic bacteria by a highly regulated biomineralization process. Their usability in many applications could be further improved by encapsulation in biocompatible polymers. In this study, we explored the production of spider silk-inspired peptides on magnetosomes of the alphaproteobacterium Magnetospirillum gryphiswaldense. Genetic fusion of different silk sequence-like variants to abundant magnetosome membrane proteins enhanced magnetite biomineralization and caused the formation of a proteinaceous capsule, which increased the colloidal stability of isolated particles. Furthermore, we show that spider silk peptides fused to a magnetosome membrane protein can be used as seeds for silk fibril growth on the magnetosome surface. In summary, we demonstrate that the combination of two different biogenic materials generates a genetically encoded hybrid composite with engineerable new properties and enhanced potential for various applications.

Flow cytometry as a rapid analytical tool to determine physiological responses to changing O2 and iron concentration by Magnetospirillum gryphiswaldense strain MSR-1.

Magnetotactic bacteria (MTB) are a diverse group of bacteria that synthesise magnetosomes, magnetic membrane-bound nanoparticles that have a variety of diagnostic, clinical and biotechnological applications. We present the development of rapid methods using flow cytometry to characterize several aspects of the physiology of the commonly-used MTB Magnetospirillum gryphiswaldense MSR-1. Flow cytometry is an optical technique that rapidly measures characteristics of individual bacteria within a culture, thereby allowing determination of population heterogeneity and also permitting direct analysis of bacteria. Scatter measurements were used to measure and compare bacterial size, shape and morphology. Membrane permeability and polarization were measured using the dyes propidium iodide and bis-(1,3-dibutylbarbituric acid) trimethine oxonol to determine the viability and 'health' of bacteria. Dyes were also used to determine changes in concentration of intracellular free iron and polyhydroxylakanoate (PHA), a bacterial energy storage polymer. These tools were then used to characterize the responses of MTB to different O2 concentrations and iron-sufficient or iron-limited growth. Rapid analysis of MTB physiology will allow development of bioprocesses for the production of magnetosomes, and will increase understanding of this fascinating and useful group of bacteria.

Chains of magnetosomes with controlled endotoxin release and partial tumor occupation induce full destruction of intracranial U87-Luc glioma in mice under the application of an alternating magnetic field.

Previous studies showed that magnetic hyperthermia could efficiently destroy tumors both preclinically and clinically, especially glioma. However, antitumor efficacy remained suboptimal and therefore required further improvements. Here, we introduce a new type of nanoparticles synthesized by magnetotactic bacteria, called magnetosomes, with improved properties compared with commonly used chemically synthesized nanoparticles. Indeed, mice bearing intracranial U87-Luc glioma tumors injected with 13µg of nanoparticles per mm3 of tumor followed by 12 to 15 of 30min alternating magnetic field applications displayed either full tumor disappearance in 40% of mice or no tumor regression using magnetosomes or chemically synthesized nanoparticles, respectively. Magnetosome superior antitumor activity could be explained both by a larger production of heat and by endotoxins release under alternating magnetic field application. Most interestingly, this behavior was observed when magnetosomes occupied only 10% of the whole tumor volume, which suggests that an indirect mechanism, such as an immune reaction, takes part in tumor regression. This is desired for the treatment of infiltrating tumors, such as glioma, for which whole tumor coverage by nanoparticles can hardly be achieved.

Effects of dissolved oxygen concentration and iron addition on immediate-early gene expression of Magnetospirillum gryphiswaldense MSR-1.

We report the effects of dissolved oxygen (DO) concentration and iron addition on gene expression of Magnetospirillum gryphiswaldense MSR-1 cells during fermentations, focusing on 0.25-24 h after iron addition. The DO was strictly controlled at 0.5% or 5% O2, and compared with aerobic condition. Uptake of iron (and formation of magnetosomes) was only observed in the 0.5% O2 condition where there was little difference in cell growth and carbon consumption compared to the 5% O2 condition. Quantitative reverse transcription PCR analysis showed a rapid (within 0.25 h) genetic response of MSR-1 cells after iron addition for all the genes studied, except for MgFnr (oxygen sensor gene) and fur (ferric uptake regulator family gene), and which in some cases was oxygen dependent. In particular, expression of sodB1 (superoxide dismutase gene) and feoB1 (ferrous transport protein B1 gene) was markedly reduced in cultures at 0.5% O2 compared to those at higher oxygen tensions. Moreover, expression of katG (catalase-peroxidase gene) and feoB2 (ferrous transport protein B2 gene) was reduced markedly by iron addition, regardless of oxygen conditions. These data provide a greater understanding of molecular response of MSR-1 cells to environmental conditions associated with oxygen and iron metabolisms, especially relevant to immediate-early stage of fermentation.

Proteins with CHADs (Conserved Histidine α-Helical Domains) Are Attached to Polyphosphate Granules In Vivo and Constitute a Novel Family of Polyphosphate-Associated Proteins (Phosins).

On the basis of bioinformatic evidence, we suspected that proteins with a CYTH (CyaB thiamine triphosphatase) domain and/or a CHAD (conserved histidine α-helical domain) motif might represent polyphosphate (polyP) granule-associated proteins. We found no evidence of polyP targeting by proteins with CYTH domains. In contrast, two CHAD motif-containing proteins from Ralstonia eutropha H16 (A0104 and B1017) that were expressed as fusions with enhanced yellow fluorescent protein (eYFP) colocalized with polyP granules. While the expression of B1017 was not detectable, the A0104 protein was specifically identified in an isolated polyP granule fraction by proteome analysis. Moreover, eYFP fusions with the CHAD motif-containing proteins MGMSRV2-1987 from Magnetospirillum gryphiswaldense and PP2307 from Pseudomonas putida also colocalized with polyP granules in a transspecies-specific manner. These data indicated that CHAD-containing proteins are generally attached to polyP granules. Together with the findings from four previously polyP-attached proteins (polyP kinases), the results of this study raised the number of polyP-associated proteins in R. eutropha to six. We suggest designating polyP granule-bound proteins with CHAD motifs as phosins (phosphate), analogous to phasins and oleosins that are specifically bound to the surface of polyhydroxyalkanoate (PHA) granules in PHA-accumulating bacteria and to oil droplets in oil seed plants, respectively.IMPORTANCE The importance of polyphosphate (polyP) for life is evident from the ubiquitous presence of polyP in all species on earth. In unicellular eukaryotic microorganisms, polyP is located in specific membrane-enclosed organelles, called acidocalcisomes. However, in most prokaryotes, polyP is present as insoluble granules that have been designated previously as volutin granules. Almost nothing is known regarding the macromolecular composition of polyP granules. Particularly, the absence or presence of cellular compounds on the surface of polyP granules has not yet been investigated. In this study, we identified a novel class of proteins that are attached to the surface of polyP granules in three model species of Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria These proteins are characterized by the presence of a CHAD (conserved histidine α-helical domain) motif that functions as a polyP granule-targeting signal. We suggest designating CHAD motif-containing proteins as phosins [analogous to phasins for poly(3-hydroxybutyrate)-associated proteins and to oleosins for oil droplet-associated proteins in oil seed plants]. The expression of phosins in different species confirmed their polyP-targeting function in a transspecies-specific manner. We postulate that polyP granules in prokaryotic species generally have a complex surface structure that consists of one to several polyP kinases and phosin proteins. We suggest differentiating polyP granules from acidocalcisomes by designating them as polyphosphatosomes.

Bacterial magnetic nanoparticles for photothermal therapy of cancer under the guidance of MRI.

The bacterial magnetic nanoparticles (BMPs) are biomineralized by the magnetotactic bacteria and naturally covered with a layer of biomembrane. Herein, BMPs were isolated and firstly used for the photothermal therapy (PTT) of cancer under the guidance of magnetic resonance imaging (MRI) in vitro and in vivo. The results showed that BMPs could rapidly convert the energy of 808 nm near-infrared (NIR) light into heat. After internalization by the HepG2 tumor cells, BMPs with good biocompatibility could induce an efficient killing effect after NIR light irradiation, along with a change of mitochondrial membrane potential (ΔΨm) and level of intracellular reactive oxygen species (ROS). The in vivo therapy also confirms that PTT with BMPs could effectively and completely ablate the tumor in mice without inducing observable toxicity. T2-weighted MRI showed a clear tumor boundary and a 25% enhancement of negative contrast enhancement at the tumor site, suggesting that BMPs can act as an effective MRI contrast agent for guiding the PTT. Our results indicate that BMPs could be a potential theranostic agent for simultaneous MRI and PTT of cancer.

In vitro assembly of the bacterial actin protein MamK from ' Candidatus Magnetobacterium casensis' in the phylum Nitrospirae.

Magnetotactic bacteria (MTB), a group of phylogenetically diverse organisms that use their unique intracellular magnetosome organelles to swim along the Earth's magnetic field, play important roles in the biogeochemical cycles of iron and sulfur. Previous studies have revealed that the bacterial actin protein MamK plays essential roles in the linear arrangement of magnetosomes in MTB cells belonging to the Proteobacteria phylum. However, the molecular mechanisms of multiple-magnetosome-chain arrangements in MTB remain largely unknown. Here, we report that the MamK filaments from the uncultivated 'Candidatus Magnetobacterium casensis' (Mcas) within the phylum Nitrospirae polymerized in the presence of ATP alone and were stable without obvious ATP hydrolysis-mediated disassembly. MamK in Mcas can convert NTP to NDP and NDP to NMP, showing the highest preference to ATP. Unlike its Magnetospirillum counterparts, which form a single magnetosome chain, or other bacterial actins such as MreB and ParM, the polymerized MamK from Mcas is independent of metal ions and nucleotides except for ATP, and is assembled into well-ordered filamentous bundles consisted of multiple filaments. Our results suggest a dynamically stable assembly of MamK from the uncultivated Nitrospirae MTB that synthesizes multiple magnetosome chains per cell. These findings further improve the current knowledge of biomineralization and organelle biogenesis in prokaryotic systems.

Iron response regulator protein IrrB in Magnetospirillum gryphiswaldense MSR-1 helps control the iron/oxygen balance, oxidative stress tolerance, and magnetosome formation.

Magnetotactic bacteria are capable of forming nanosized, membrane-enclosed magnetosomes under iron-rich and oxygen-limited conditions. The complete genomic sequence of Magnetospirillum gryphiswaldense strain MSR-1 has been analyzed and found to contain five fur homologue genes whose protein products are predicted to be involved in iron homeostasis and the response to oxidative stress. Of these, only the MGMSRv2_3149 gene (irrB) was significantly downregulated under high-iron and low-oxygen conditions, during the transition of cell growth from the logarithmic to the stationary phase. The encoded protein, IrrB, containing the conserved HHH motif, was identified as an iron response regulator (Irr) protein belonging to the Fur superfamily. To investigate the function of IrrB, we constructed an irrB deletion mutant (ΔirrB). The levels of cell growth and magnetosome formation were lower in the ΔirrB strain than in the wild type (WT) under both high-iron and low-iron conditions. The ΔirrB strain also showed lower levels of iron uptake and H2O2 tolerance than the WT. Quantitative real-time reverse transcription-PCR analysis indicated that the irrB mutation reduced the expression of numerous genes involved in iron transport, iron storage, heme biosynthesis, and Fe-S cluster assembly. Transcription studies of the other fur homologue genes in the ΔirrB strain indicated complementary functions of the Fur proteins in MSR-1. IrrB appears to be directly responsible for iron metabolism and homeostasis and to be indirectly involved in magnetosome formation. We propose two IrrB-regulated networks (under high- and low-iron conditions) in MSR-1 cells that control the balance of iron and oxygen metabolism and account for the coexistence of five Fur homologues.