Diverse Mn(II)-Oxidizing Bacteria Isolated from Submarine Basalts at Loihi Seamount

Abstract

Metal-oxidizing bacteria may play a key role in the submarine weathering of volcanic rocks and the formation of ferromanganese crusts. Putative fossil microbes encrusted in Mn oxide phases are commonly observed on volcanic glasses recovered from the deep ocean; however, no known Mn(II)-oxidizing bacteria have been directly identified or cultured from natural weathered basalts. To isolate epilithic Mn(II) oxidizing bacteria, we collected young, oxidized pillow basalts from the cold, outer portions of Loihi Seamount, and from nearby exposures of pillow basalts at South Point and Kealakekua Bay, HI. SEM imaging, EDS spectra and X-ray absorption spectroscopy data show that microbial biofilms and associated Mn oxides were abundant on the basalt surfaces. Using a series of seawater-based media that range from highly oligotrophic to organic-rich, we have obtained 26 mesophilic, heterotrophic Mn(II)-oxidizing isolates dominated by α- and γ-Proteobacteria, such as Sulfitobacter, Methylarcula and Pseudoalteromonas spp. Additional isolates include Microbulbifer, Alteromonas, Marinobacter, and Halomonas spp. None of the isolates, nor their closest relatives, were previously recognized as Mn(II) oxidizing bacteria. The physiological function of Mn(II) oxidation is clearly spread amongst many phylogenetically diverse organisms colonizing basalt surfaces. Our findings support a biological catalysis of Mn(II) oxidation during basalt-weathering, and suggest heterotrophic Mn(II) oxidizing bacteria may be ubiquitously associated with submarine glasses within epilithic and endolithic biofilms.     

A transient outward current dependent on external calcium in rat cerebellar granule cells

Summary The outward potassium current of rat cerebellar granule cells in culture was studied with the whole-cell patch-clamp method. Two voltage-dependent components were identified: a slow current, resembling the classical delayed rectifier current, and a fast component, similar to anI _A-type current. The slow current was insensitive to 4-aminopyridine and independent of external Ca²⁺, but significantly inhibited by 3mM tetraethylammonium. The fast current was depressed by external 4-aminopyridine, with an ED₅₀=0.7mM, and it was abolished by removal of divalent cations from the external medium. The sensitivity of the transient outward current to different divalent cations was investigated by equimolar substitution of Ca²⁺, Mn²⁺ and Mg²⁺. In 2.8mM Mn²⁺, the transient potassium conductance was comparable to that in 2.8mM Ca²⁺, while in 2.8mM Mg²⁺ the transient component was drastically reduced, as in the absence of any divalent cations. However, when Ca²⁺ was present, Mg²⁺ up to 5mM had no effect. The transient current increased with increasing concentrations of external Ca²⁺, [Ca²⁺] _o , and the maximum conductancevs. [Ca²⁺] _o curve could be approximated by a one-site model. In addition, the current recorded with 5.5mM BAPTA in the intracellular solution was not different from that recorded in the absence of any Ca²⁺ buffer. These results suggest that divalent cations modulate the potassium channel interacting with a site on the external side of the cell membrane.     

Effect of heat treatment in vacuum on photoluminescence of anodic alumina

The results are presented of a study of the photoluminescent (PL) properties of an undoped porous anodic alumina (PAA) and PAA doped with manganese ions. The PAA samples were prepared by anodization of aluminum. The effect of annealing conditions in vacuum on the PL spectra was studied for the first time and a comparative analysis was made with the spectra of the PAA annealed in air. Vacuum annealing was used to obtain oxygen‐deficient alumina. A strong dependence of the PAA PL intensity on the annealing temperature in vacuum has been found: for the samples annealed at 600°С, the PL intensity is 15 times higher than that measured on the initial samples, whereas for the samples annealed in air it increases only 4.5‐fold with excitation at the wavelengths of 275 nm. This is the result of the formation of a high concentration of oxygen vacancies during annealing in vacuum under conditions of oxygen deficiency as compared with the samples annealed in air, where diffusion of oxygen from air leads to a decrease in vacancies. A significant increase in the PL intensity permits consideration of the vacuum‐annealed PAA as a promising material for dosimetry.     

DNA-PKcs is activated under nutrient starvation and activates Akt,MST1, FoxO3a,and NDR1

BackgroundPresence of unperfused regions containing cells under hypoxia and nutrient starvation; contributes to radioresistance in solid human tumors. We have previously reported that cultured cells; under nutrient starvation show resistance to ionizing radiation compare with cells under normal; condition, and that nutrient starvation increases ATM activity, which causes cellular resistance to; ionizing radiation (Murata et al., BBRC2018). For further investigation of molecular mechanisms; underlying radioresistance of cells under nutrient starvation, effects of nutrient starvation on activity; of DNA-PKcs have been investigated because both DNA-PKcs and ATM belong to the PIKK family; and are required for DNA DSBs repair. In addition to DNA-PKcs, effects of nutrient starvation on; activities of FoxO3a and its regulators Akt, MST1 and AMPK have been investigated because FoxO3a; mediates cellular responses to stress and is activated under nutrient starvation.MethodsA human glioblastoma cell line, T98G was used to examine the effects of nutrient starvation on activities and expression of DNA-PKcs, Akt, MST1, FoxO3a, NDR1, and AMPK. To elucidate; signal transduction pathways for FoxO3a activation under nutrient starvation, we examined effects of; specific inhibitors or siRNA for DNA-PKcs or Akt on activities and expression of MST1, FoxO3, NDR1, andAMPK.ResultsUnder nutrient starvation, phosphorylations of DNA-PKcs at Ser2056, Akt at Ser473, MST at Thr183, FoxO3a at Ser413, NDR1 at Ser281 and Thr282, and AMPK at Thr172 were increased, which suggests their activation. Nutrient starvation did not affect expression of DNA-PKcs, Akt, MST1, or NDR1, with decreased expression of FoxO3a and increased expression of AMPK. Inhibition; of DNA-PK suppressed phosphorylation of Akt under nutrient starvation. Inhibition of DNA-PK or; Akt suppressed phosphorylations of MST1, FoxO3a, and NDR1 under nutrient starvation, which; suggests DNA-PKcs and Akt activate MST1, FoxO3a, and NDR1. Inhibition of DNA-PK did not; suppress phosphorylation ofAMPK under nutrient starvation.ConclusionOur data suggest that DN-PKcs is activated under nutrient starvation and activates AktMST1, FoxO3a, and NDR1.     

Highlighting the roles of transition metals and speciation in chemical biology

Transition metal ions play key structural and functional roles, affecting structures of biomolecules and enzyme function. The importance of transition metal ions in chemical biology is, thus, undisputed. However, the aqueous chemistry of metal ions is complicated because they form species in several protonation and redox states. In the presence of metabolites, metal ions can also form coordination complexes. The existence of several species is relevant because enzymes and membrane receptors can distinguish between species even when they are rapidly equilibrating. Thus, metal ions, enzyme cofactors, and therapeutic agents are sensitive to the metal ion speciation chemistry because it affects their interaction with enzymes and other biomolecules. Speciation is also crucial for metal-containing bioorthogonal reactions, since water and metabolites stabilize active catalysts, affect chemoselectivity and reaction yields.     

Metal cation controls myosin and actomyosin kinetics

We have perturbed myosin nucleotide binding site with magnesium‐, manganese‐, or calcium‐nucleotide complexes, using metal cation as a probe to examine the pathways of myosin ATPase in the presence of actin. We have used transient time‐resolved FRET, myosin intrinsic fluorescence, fluorescence of pyrene labeled actin, combined with the steady state myosin ATPase activity measurements of previously characterized D.discoideum myosin construct A639C:K498C. We found that actin activation of myosin ATPase does not depend on metal cation, regardless of the cation‐specific kinetics of nucleotide binding and dissociation. The rate limiting step of myosin ATPase depends on the metal cation. The rate of the recovery stroke and the reverse recovery stroke is directly proportional to the ionic radius of the cation. The rate of nucleotide release from myosin and actomyosin, and ATP binding to actomyosin depends on the cation coordination number.     

Iron-mediated degradation of ribosomes under oxidative stress is attenuated by manganese

Protein biosynthesis is fundamental to cellular life and requires the efficient functioning of the translational machinery. At the center of this machinery is the ribosome, a ribonucleoprotein complex that depends heavily on Mg²⁺ for structure. Recent work has indicated that other metal cations can substitute for Mg²⁺, raising questions about the role different metals may play in the maintenance of the ribosome under oxidative stress conditions. Here, we assess ribosomal integrity following oxidative stress both in vitro and in cells to elucidate details of the interactions between Fe²⁺ and the ribosome and identify Mn²⁺ as a factor capable of attenuating oxidant-induced Fe²⁺-mediated degradation of rRNA. We report that Fe²⁺ promotes degradation of all rRNA species of the yeast ribosome and that it is bound directly to RNA molecules. Furthermore, we demonstrate that Mn²⁺ competes with Fe²⁺ for rRNA-binding sites and that protection of ribosomes from Fe²⁺-mediated rRNA hydrolysis correlates with the restoration of cell viability. Our data, therefore, suggest a relationship between these two transition metals in controlling ribosome stability under oxidative stress.     

Manganese Oxide/Carbon Aerogel Composite: an Outstanding Supercapacitor Electrode Material

Manganese oxide/carbon aerogel composite electrodes are prepared by a self‐limiting anodic‐electrochemical deposition of manganese oxide into a binder‐enriched carbon aerogel layer, drop‐cast on a graphite substrate, using 0.1 M Mn(CH₃COO)₂·4H₂O as the electrolyte. Manganese oxide grows in the form of thin nanofibers along the backbone of the carbon aerogel, leaving adequate working space for the electrolyte and enabling a fuller extent of the utilization of the manganese oxide to make the composite an outstanding supercapacitor electrode material. The manganese oxide is determined to be Mn₃O₄ with the Raman spectroscopy and high‐resolution transmission electron microscopy. The rectangularity of the cyclic‐voltammogram loops of the composite electrode is excellent and remains that way for scan rates up to a very‐high value of 500 mV s^?1, indicating extremely good redox reversibility and cycle efficiency. At a scan rate of 25 mV s^?1, the specific capacitance, as measured in 0.5 M Na₂SO₄ for a potential window of 0.1–0.9 V vs. Ag/AgCl, reaches a maximum value of 503 F g^?1 and experiences only a negligible decay of less than 1% at the 6000th cycle, implying an extraordinary cycling stability. The cycling efficiency is as high as 98% at a current density of 8 A g^?1 cm^?2, showing an almost‐ideal capacitive behavior. The power density reaches 48.5 kW kg^?1 and the energy density 21.6 W h kg^?1 at a scan rate of 500 mV s^?1, well above the specifications of current state‐of‐the‐art supercapacitors.     

Production of melanin bleaching enzyme of fungal origin and its application in cosmetics

We screened wild fungal isolates for melanolytic activity and found thatSporotrichum pruinosum was the most promising of the very limited number of fungi that decolourised synthetic melanin. We used a submerged aerobic process to produce a skin depigmentation enzyme by this strain, and found that in the medium the presence of Mn²⁺ ions was necessary, the limitation of carbon source was beneficial, and Zn²⁺ ions were inhibitory. Cultivation in a stirred bioreactor required immobilization of mycelium and use of low stirring velocity. A partially purified enzyme was prepared and tested for depigmentation of human skin corneocytes and whole epidermis of phototypes III and V. This is the first study demonstrating the effective enzymatic degradation of the skin melanin rather than inhibition of its synthesis. This opens the possibility of using melanolytic enzymes in cosmetic skin lightening.