Characterization of a spontaneous mutant of Azotobacter vinelandii in which vanadium-dependent nitrogen fixation is not inhibited by molybdenum

A spontaneous mutant derivative of Azotobacter vinelandii CA12 (ΔnifHDK), in which vanadium-dependent nitrogen fixation is not inhibited by molybdenum (A. vinelandii CARR), grows profusely on BNF-agar containing 1 μM Na₂MoO₄, alone or supplemented with 1 μM V₂O₅. The expression of A. vinelandii vnfH::lacZ and vnfA::lacZ fusions in A. vinelandii CARR was not inhibited by 1 mM Na₂MoO₄, whereas molybdenum at much lower concentration inhibited the expression of vnfH::lacZ and vnfA::lacZ fusions in A. vinelandii CA12. The mutant also exhibited normal acetylene reduction activity in the presence of 1 μM Na₂MoO₄. The expression of A. vinelandii nifH::lacZ fusion in A. vinelandii CARR was low even though the cells were cultured under non-repressing conditions with urea as nitrogen source in the presence of Na₂MoO₄. The molybdenum content of A. vinelandii CARR cells was found to be about one-fourth that of A. vinelandii CA12. No nitrate reductase activity could be detected in A. vinelandii CARR when the cells were cultured in the presence of 10 μM Na₂MoO₄, whereas A. vinelandii CA12 exhibited some activity even with 100 pM Na₂MoO₄.     

Effects of drought on nitrogen fixation in soybean root nodules

Soybean plants [Glycine max (L.) Merr.] were grown in silica sand and were drought stressed for a 4 week period during reproductive development and without any mineral N supply in order to maximize demand for fixed nitrogen. A strain of Bradyrhizobium japonicum that forms large quantities of polysaccharide in nodules was used to determine whether or not the supply of reduced carbon to bacteroids limits nitrogenase activity. A depression of 30–40% in nitrogen content in leaves and pods of stressed plants indicated a marked decline in nitrogen fixation activity during the drought period. A 50% increase in the accumulation of bacterial polysaccharide in nodules accompanied this major decrease in nitrogen fixation activity and this result indicates that the negative impact of drought on nodules was not due to a depression of carbon supply to bacteroids. The drought treatment resulted in a statistically significant increase in N concentration in leaves and pods. Because N concentration and chlorophyll concentration in leaves were not depressed, there was no evidence of nitrogen deficiency in drought‐stressed plants, and this result indicates that the negative impact of drought on nodule function was not the cause of the depression of shoot growth. At the end of the drought period, the concentration of carbohydrates, amino nitrogen, and ureides was significantly increased in nodules on drought‐stressed plants. The overall results support the view that, under drought conditions, nitrogen fixation activity in nodules was depressed because demand for fixed N to support growth was lower.     

Decadally cycling soil carbon is more sensitive to warming than faster‐cycling soil carbon

The response of soil organic carbon (SOC) pools to globally rising surface temperature crucially determines the feedback between climate change and the global carbon cycle. However, there is a lack of studies investigating the temperature sensitivity of decomposition for decadally cycling SOC which is the main component of total soil carbon stock and the most relevant to global change. We tackled this issue using two decadally ¹³C‐labeled soils and a much improved measuring system in a long‐term incubation experiment. Results indicated that the temperature sensitivity of decomposition for decadally cycling SOC (>23 years in one soil and >55 years in the other soil) was significantly greater than that for faster‐cycling SOC (<23 or 55 years) or for the entire SOC stock. Moreover, decadally cycling SOC contributed substantially (35–59%) to the total CO₂ loss during the 360‐day incubation. Overall, these results indicate that the decomposition of decadally cycling SOC is highly sensitive to temperature change, which will likely make this large SOC stock vulnerable to loss by global warming in the 21st century and beyond.     

Legume symbiotic nitrogen fixation by beta-proteobacteria is widespread in nature

Following the initial discovery of two legume-nodulating Burkholderia strains (L. Moulin, A. Munive, B. Dreyfus, and C. Boivin-Masson, Nature 411:948-950, 2001), we identified as nitrogen-fixing legume symbionts at least 50 different strains of Burkholderia caribensis and Ralstonia taiwanensis, all belonging to the beta-subclass of proteobacteria, thus extending the phylogenetic diversity of the rhizobia. R. taiwanensis was found to represent 93% of the Mimosa isolates in Taiwan, indicating that beta-proteobacteria can be the specific symbionts of a legume. The nod genes of rhizobial beta-proteobacteria (beta-rhizobia) are very similar to those of rhizobia from the alpha-subclass (alpha-rhizobia), strongly supporting the hypothesis of the unique origin of common nod genes. The beta-rhizobial nod genes are located on a 0.5-Mb plasmid, together with the nifH gene, in R. taiwanensis and Burkholderia phymatum. Phylogenetic analysis of available nodA gene sequences clustered beta-rhizobial sequences in two nodA lineages intertwined with alpha-rhizobial sequences. On the other hand, the beta-rhizobia were grouped with free-living nitrogen-fixing beta-proteobacteria on the basis of the nifH phylogenetic tree. These findings suggest that beta-rhizobia evolved from diazotrophs through multiple lateral nod gene transfers.     

Survivability is more fundamental than evolvability

For a lineage to survive over long time periods, it must sometimes change. This has given rise to the term evolvability, meaning the tendency to produce adaptive variation. One lineage may be superior to another in terms of its current standing variation, or it may tend to produce more adaptive variation. However, evolutionary outcomes depend on more than standing variation and produced adaptive variation: deleterious variation also matters. Evolvability, as most commonly interpreted, is not predictive of evolutionary outcomes. Here, we define a predictive measure of the evolutionary success of a lineage that we call the k-survivability, defined as the probability that the lineage avoids extinction for k generations. We estimate the k-survivability using multiple experimental replicates. Because we measure evolutionary outcomes, the initial standing variation, the full spectrum of generated variation, and the heritability of that variation are all incorporated. Survivability also accounts for the decreased joint likelihood of extinction of sub-lineages when they 1) disperse in space, or 2) diversify in lifestyle. We illustrate measurement of survivability with in silico models, and suggest that it may also be measured in vivo using multiple longitudinal replicates. The k-survivability is a metric that enables the quantitative study of, for example, the evolution of 1) mutation rates, 2) dispersal mechanisms, 3) the genotype-phenotype map, and 4) sexual reproduction, in temporally and spatially fluctuating environments. Although these disparate phenomena evolve by well-understood microevolutionary rules, they are also subject to the macroevolutionary constraint of long-term survivability.     

Why genomics is more than genomes

A report on the 2004 meeting on Molecular Genetics of Bacteria and Bacteriophages, Cold Spring Harbor, USA, 25-29 August 2004.     

Nitrogen fixation by Klebsiella pneumoniae is inhibited by certain multicopy hybrid nif plasmids.

In our studies of nif gene regulation, we have observed that certain hybrid nif plasmids drastically inhibit the expression of the chromosomal nif genes of Klebsiella pneumonia. Wild-type (Nif+) K. pneumoniae strains that acquire certain hybrid nif plasmids also acquire the Nif- phenotype; these strains lose 90 to 99% of all detectable nitrogen fixation activity and grow poorly (or not at all) on solid media with N2 as the sole nitrogen source. We describe experiments which defined this inhibition of the Nif+ phenotype by hybrid nif plasmids and identify and characterize four nif DNA regions associated with this inhibition. We show that plasmids carrying these nif regions could recombine with, but not complement, nif chromosomal mutations. Our results suggest that inhibition of the Nif+ phenotype will provide a useful bioassay for some of the factors that mediate nif gene expression.     

Sarcopenia is more than a muscular deficit

Sarcopenia is a complex process that appears in aged muscle associated with a decrease in mass, strength, and velocity of contraction. This process is the result of many molecular, cellular and functional alterations. It has been suggested that sarcopenia may be triggered by reactive oxygen species (ROS) that have accumulated throughout one's lifetime. We found a significant increase in oxidation of DNA and lipids in the elderly muscle, more evident in males, and a reduction in catalase and glutathione transferase activities. Experiments on Ca2+ transport showed an abnormal functional response of aged muscle after exposure to caffeine, which increases the opening of Ca2+ channels, as well a reduced activity of the Ca2+ pump in elderly males. From these results we concluded that oxidative stress play an important role in muscle aging and that oxidative damage is much more evident in elderly males, suggesting a gender difference may be related to hormonal factors. The progression of sarcopenia is directly related to a significant reduction of the regenerative potential of muscle normally due to a type of adult stem cells, known as satellite cells, which lie outside the sarcolemma and remain quiescent until external stimuli trigger as growth factors (IGF-1 or mIGF-1) their re-entry into the cell cycle. One possibility is that the anti oxidative capacity of satellite cells could also be altered and this, in turn, determines the decrease of their regenerative capacity. Data concerning this hypothesis are discussed     

There is more to autophagy than induction

Considerable attention has been paid to the topic of autophagy induction. In part, this is because of the potential for modulating this process for therapeutic purposes. Of course we know that induced autophagy can also be problematic—for example, when trying to eliminate an established tumor that might be relying on autophagy for its own cytoprotective uses. Accordingly, inhibitory mechanisms have been considered; however, the corresponding studies have tended to focus on the pathways that block autophagy under noninducing conditions, such as when nutrients are available. In contrast, relatively little is known about the mechanisms for inhibiting autophagy under inducing conditions. Yet, this type of regulation must be occurring on a routine basis. We know that dysregulation of autophagy, e.g., due to improper activation of Beclin 1 leading to excessive autophagy activity, can cause cell death.¹ Pattingre S, Tassa A, Qu X, Garuti R, Liang XH, Mizushima N, et al. Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy. Cell 2005; 122:927 - 39; http://dx.doi.org/10.1016/j.cell.2005.07.002; PMID: 16179260 [Crossref], [PubMed], [Web of Science ®] , [Google Scholar] Accordingly, we assume that during starvation or other inducing conditions there must be a mechanism to modulate autophagy. That is, once you turn it on, you do not want to let it continue unchecked. But how is autophagy downregulated when the inducing conditions still exist?     

Procollagen is more stable in cellulo than in vitro

The thermal denaturation of both intracellular and freshly secreted chick embryo tendon type I procollagen was investigated using susceptibility to proteolysis by trypsin and chymotrypsin as a probe for triple-helical conformation. Freshly secreted procollagen from the medium of matrix-free tendon cells in suspension or procollagen within the cells and in the pericellular environment melted at 45 degrees C. In contrast, if freshly secreted procollagen was subjected to the melting procedure after dialysis of the medium against 0.4 M NaCl, 0.1 M Tris HCl, pH 7.4 the protein melted at 42 degrees C, the melting temperature of purified procollagen dissolved in the same buffer. In each of these cases, the thermal denaturation profile was narrow, with a width of 1.0-1.5 degrees C. These results demonstrate that, in situ, procollagen is more stable toward thermal denaturation than was previously thought. This extra margin of thermal stability partially resolves the dilemma of how tissues are able to assemble triple-helical procollagen molecules at body temperatures that closely approach the melting temperature of the purified protein.     

Response to drought stress of nitrogen fixation (acetylene reduction) rates by field-grown soybeans

The effects of drought stress on soybean nodule conductance and the maximum rate of acetylene reduction were studied with in situ experiments performed during two seasons and under differing field conditions. In both years drought resulted in decreased nodule conductances which could be detected as early as three days after water was withheld. The maximum rate of acetylene reduction was also decreased by drought and was highly correlated with nodule conductance (r = 0.95). Since nodule conductance is equal to the nodule surface area times the permeability, the relationship of these variables to both whole-plant and unit-nodule nitrogenase activity was explored. Drought stress resulted in a decrease in nodule gas permeability followed by decreases in nodule surface area when drought was prolonged. Under all conditions studied acetylene reduction on a unit-nodule surface area basis was highly correlated with nodule gas permeability (r = 0.92). A short-term oxygen enrichment study demonstrated nodule gas permeability may limit oxygen flux into both drought-stressed and well-watered nodules of these field-grown soybeans.