Condensed chromatin and cell inactivation by single-hit kinetics

Mammalian cells are extremely sensitive to gamma rays at mitosis, the time at which their chromatin is maximally condensed. The radiation-induced killing of mitotic cells is well described by single-hit inactivation kinetics. To investigate if radiation hypersensitivity by single-hit inactivation correlated with chromatin condensation, Chinese hamster ovary (CHO) K1 (wild-type) and xrs-5 (radiosensitive mutant) cells were synchronized by mitotic shake-off procedures and the densities of their chromatin cross sections and their radiosensitivities were measured immediately and 2 h into G1 phase. The chromatin of G1-phase CHO K1 cells was dispersed uniformly throughout their nuclei, and its average density was at least three times less than in the chromosomes of mitotic CHO K1 cells. The alpha-inactivation co-efficient of mitotic CHO K1 cells was approximately 2.0 Gy(-1) and decreased approximately 10-fold when cells entered G1 phase. The density of chromatin in CHO xrs-5 cell chromosomes at mitosis was greater than in CHO K1 cell chromosomes, and the radiosensitivity of mitotic CHO xrs-5 cells was the greatest with alpha = 5.1 Gy(-1). In G1 phase, CHO xrs-5 cells were slightly more resistant to radiation than when in mitosis, but a significant proportion of their chromatin was found to remain in condensed form adjacent to the nuclear membrane. These studies indicate that in addition to their known defects in DNA repair and V(D)J recombination, CHO xrs-5 cells may also be defective in some process associated with the condensation and/or dispersion of chromatin at mitosis. Their radiation hypersensitivity could result, in part, from their DNA remaining in compacted form during interphase. The condensation status of DNA in other mammalian cells could define their intrinsic radiosensitivity by single-hit inactivation, the mechanism of cell killing which dominates at the dose fraction size (1.8-2.0 Gy) most commonly used in radiotherapy.     

PHOTOSYNTHESIS AND PRODUCTION OF HYDROGEN PEROXIDE BY SYMBIODINIUM (PYRRHOPHYTA) PHYLOTYPES WITH DIFFERENT THERMAL TOLERANCES1

Occurrences whereby cnidaria lose their symbiotic dinoflagellate microalgae (Symbiodinium spp.) are increasing in frequency and intensity. These so‐called bleaching events are most often related to an increase in water temperature, which is thought to limit certain Symbiodinium phylotypes from effectively dissipating absorbed excitation energy that is otherwise used for photochemistry. Here, we examined photosynthetic characteristics and hydrogen peroxide (H₂O₂) production, a possible signal involved in bleaching, from two Symbiodinium types (a thermally “tolerant” A1 and “sensitive” B1) representative of cnidaria–Symbiodinium symbioses of reef‐building Caribbean corals. Under steady‐state growth at 26°C, a higher efficiency of PSII photochemistry, rate of electron turnover, and rate of O₂ production were observed for type A1 than for B1. The two types responded very differently to a period of elevated temperature (32°C): type A1 increased light‐driven O₂ consumption but not the amount of H₂O₂ produced; in contrast, type B1 increased the amount of H₂O₂ produced without an increase in light‐driven O₂ consumption. Therefore, our results are consistent with previous suggestions that the thermal tolerance of Symbiodinium is related to adaptive constraints associated with photosynthesis and that sensitive phylotypes are more prone to H₂O₂ production. Understanding these adaptive differences in the genus Symbiodinium will be crucial if we are to interpret the response of symbiotic associations, including reef‐building corals, to environmental change.     

Improved tolerance to bile salts of aggregated Bifidobacterium longum produced during continuous culture with immobilized cells

The effect of cell immobilization and continuous culture was studied on selected physiological and technological characteristics of Bifidobacterium longum NCC2705 cultivated for 20 days in a two stage continuous fermentation system. Continuous immobilized cell (IC) cultures with and without glucose limitation exhibited formation of macroscopic cell aggregates after 12 and 9 days, respectively. Auto-aggregation resulted in underestimation of viable cell counts by plate counts by more than 2 log units CFU/ml compared with qPCR method. Modifications of cell membrane composition might partially explain aggregate formation in IC cultures. Decreases in the ratio of unsaturated to saturated fatty acid content from 1.74 to 0.58 might also contribute to the enhanced tolerance of IC cells to porcine bile salts and aminoglycosidic antibiotics compared with free cells from batch cultures.The enhanced resistance against bile salts in combination with auto-aggregation may confer an advantage to probiotic bacteria produced by IC technology.     

Rhinovirus infection induces extracellular matrix protein deposition in asthmatic and nonasthmatic airway smooth muscle cells

Airway remodeling, which includes increases in the extracellular matrix (ECM), is a characteristic feature of asthma and is correlated to disease severity. Rhinovirus (RV) infections are associated with increased risk of asthma development in young children and are the most common cause of asthma exacerbations. We examined whether viral infections can increase ECM deposition and whether this increased ECM modulates cell proliferation and migration. RV infection of nonasthmatic airway smooth muscle (ASM) cells significantly increased the deposition of fibronectin (40% increase, n = 12) and perlecan (80% increase, n = 14), while infection of asthmatic ASM cells significantly increased fibronectin (75% increase, n = 9) and collagen IV (15% increase, n = 9). We then treated the ASM cells with the Toll-like receptor (TLR) agonists polyinosinic:polycytidylic acid, imiquimod, and pure RV RNA and were able to show that the mechanism through which RV induced ECM deposition was via the activation of TLR3 and TLR7/8. Finally, we assessed whether the virus-induced ECM was bioactive by measuring the amount of migration and proliferation of virus-naive cells that seeded onto the ECM. Basically, ECM from asthmatic ASM cells induced twofold greater migration of virus-naive ASM cells than ECM from nonasthmatic ASM cells, and these rates of migration were further increased on RV-modulated ECM. Increased migration on the RV-modulated ECM was not due to increased cell proliferation, as RV-modulated ECM decreased the proliferation of virus-naive cells. Our results suggest that viruses may contribute to airway remodeling through increased ECM deposition, which in turn may contribute to increased ASM mass via increased cell migration.     

Using Affordable LED Arrays for Photo-Stimulation of Neurons

Standard slice electrophysiology has allowed researchers to probe individual components of neural circuitry by recording electrical responses of single cells in response to electrical or pharmacological manipulations^1,2. With the invention of methods to optically control genetically targeted neurons (optogenetics), researchers now have an unprecedented level of control over specific groups of neurons in the standard slice preparation. In particular, photosensitive channelrhodopsin-2 (ChR2) allows researchers to activate neurons with light^3,4. By combining careful calibration of LED-based photostimulation of ChR2 with standard slice electrophysiology, we are able to probe with greater detail the role of adult-born interneurons in the olfactory bulb, the first central relay of the olfactory system. Using viral expression of ChR2-YFP specifically in adult-born neurons, we can selectively control young adult-born neurons in a milieu of older and mature neurons. Our optical control uses a simple and inexpensive LED system, and we show how this system can be calibrated to understand how much light is needed to evoke spiking activity in single neurons. Hence, brief flashes of blue light can remotely control the firing pattern of ChR2-transduced newborn cells.Download video file.^{(48M, mov)}     

Hell Is Other People? Gender and Interactions with Strangers in the Workplace Influence a Person’s Risk of Depression

We suggest that interactions with strangers at work influence the likelihood of depressive disorders, as they serve as an environmental stressor, which are a necessary condition for the onset of depression according to diathesis-stress models of depression. We examined a large dataset (N = 76,563 in K = 196 occupations) from the German pension insurance program and the Occupational Information Network dataset on occupational characteristics. We used a multilevel framework with individuals and occupations as levels of analysis. We found that occupational environments influence employees’ risks of depression. In line with the quotation that ‘hell is other people’ frequent conflictual contacts were related to greater likelihoods of depression in both males and females (OR = 1.14, p<.05). However, interactions with the public were related to greater likelihoods of depression for males but lower likelihoods of depression for females (OR_intercation = 1.21, p<.01). We theorize that some occupations may involve interpersonal experiences with negative emotional tones that make functional coping difficult and increase the risk of depression. In other occupations, these experiences have neutral tones and allow for functional coping strategies. Functional strategies are more often found in women than in men.     

Extracellular Iron Biomineralization by Photoautotrophic Iron-Oxidizing Bacteria

Iron oxidation at neutral pH by the phototrophic anaerobic iron-oxidizing bacterium Rhodobacter sp. strain SW2 leads to the formation of iron-rich minerals. These minerals consist mainly of nano-goethite (α-FeOOH), which precipitates exclusively outside cells, mostly on polymer fibers emerging from the cells. Scanning transmission X-ray microscopy analyses performed at the C K-edge suggest that these fibers are composed of a mixture of lipids and polysaccharides or of lipopolysaccharides. The iron and the organic carbon contents of these fibers are linearly correlated at the 25-nm scale, which in addition to their texture suggests that these fibers act as a template for mineral precipitation, followed by limited crystal growth. Moreover, we evidence a gradient of the iron oxidation state along the mineralized fibers at the submicrometer scale. Fe minerals on these fibers contain a higher proportion of Fe(III) at cell contact, and the proportion of Fe(II) increases at a distance from the cells. All together, these results demonstrate the primordial role of organic polymers in iron biomineralization and provide first evidence for the existence of a redox gradient around these nonencrusting, Fe-oxidizing bacteria.Fe(II) can serve as a source of electrons for phylogenetically diverse microorganisms that precipitate iron minerals as products of their metabolism (see, e.g., references 3, 5, 25, and 30). For example, mixotrophic or autotrophic bacteria can couple the oxidation of Fe(II) to the reduction of nitrate in anoxic and neutral-pH environments. With Fe(III) being highly insoluble at neutral pH, this metabolism leads to the formation of poorly to well-crystallized iron minerals (3, 18, 26, 27) that precipitate partly within the cell periplasm for some strains (22). Similar Fe minerals are also synthesized by autotrophic bacteria that perform anoxygenic photosynthesis, using Fe(II) as an electron donor and light as a source of energy for CO₂ fixation (8, 12, 30), according to the equation HCO₃⁻ + 4 Fe²⁺ + 10 H₂O ⇆ <CH₂O> + 4 Fe(OH)₃ + 7 H⁺.However, the biological mechanisms of iron oxidation in these bacteria and in particular the way they cope with the formation of minerals within their ultrastructures are still not fully understood. Indeed, iron minerals are potentially lethal since their precipitation may alter cellular ultrastructures but also catalyze the production of free radicals (2). Recent genetic studies of the phototrophic, iron-oxidizing bacteria Rhodobacter sp. strain SW2 (6) and Rhodopseudomonas palustris strain TIE-1 (16) have identified genes (fox and pio operons, respectively) encoding proteins specific for iron oxidation. Interestingly, Jiao and Newman (16) suggested that one of these proteins could have a periplasmic localization. However, in contrast to what has been observed in some other phototrophic iron oxidizers (25) and in some nitrate-reducing, iron-oxidizing bacteria (22), no iron-mineral precipitation occurs within the periplasm of the purple nonsulfur iron-oxidizing bacterium Rhodobacter sp. strain SW2 (3). Similarly to some other anaerobic neutrophilic (22, 25) and microaerobic iron-oxidizing bacteria (5, 10), this strain seems indeed to have the ability to localize iron biomineralization at a distance from the cells, leaving large areas of the cells free of precipitates (17, 25). While it has been shown that the Gallionella and Leptothrix genera, for example, produce extracellular polymers that facilitate the nucleation of iron minerals outside cells (see, e.g., references 5 and 9), only a little is known about the existence and function of such polymers in anaerobic, neutrophilic iron-oxidizing bacteria and particularly in the phototrophic strain SW2. In the present study, we investigate iron biomineralization by the photoautotrophic iron-oxidizing bacterium Rhodobacter sp. strain SW2. We use scanning transmission X-ray microscopy (STXM) to map and identify organic polymers produced by the cells as well as the redox state of iron at the 25-nanometer scale regularly during a 2 week-period. These results demonstrate the primordial role of organic polymers in iron biomineralization and provide the first evidence for the existence of a redox gradient around SW2 cells.     

The signals of selective constraints on the mitochondrial non-coding control region: insights from comparative mitogenomics of Clupeoid fishes

Genetica - The vertebrate mitochondrial genome is characterized by an exceptional organization evolving towards a reduced size. However, the persistence of a non-coding and highly variable control...     

The complete genome sequence of Bacillus licheniformis DSM13, an organism with great industrial potential

The genome of Bacillus licheniformis DSM13 consists of a single chromosome that has a size of 4,222,748 base pairs. The average G+C ratio is 46.2%. 4,286 open reading frames, 72 tRNA genes, 7 rRNA operons and 20 transposase genes were identified. The genome shows a marked co-linearity with Bacillus subtilis but contains defined inserted regions that can be identified at the sequence as well as at the functional level. B. licheniformis DSM13 has a well-conserved secretory system, no polyketide biosynthesis, but is able to form the lipopeptide lichenysin. From the further analysis of the genome sequence, we identified conserved regulatory DNA motives, the occurrence of the glyoxylate bypass and the presence of anaerobic ribonucleotide reductase explaining that B. licheniformis is able to grow on acetate and 2,3-butanediol as well as anaerobically on glucose. Many new genes of potential interest for biotechnological applications were found in B. licheniformis; candidates include proteases, pectate lyases, lipases and various polysaccharide degrading enzymes.