Differential Roles of the Pseudomonas aeruginosa PA14 rpoN Gene in Pathogenicity in Plants, Nematodes, Insects, and Mice

We cloned the rpoN (ntrA, glnF) gene encoding the alternate sigma factor sigma(54) from the opportunistic multihost pathogen Pseudomonas aeruginosa strain PA14. A marker exchange protocol was used to construct the PA14 rpoN insertional mutation rpoN::Gen(r). PA14 rpoN::Gen(r) synthesized reduced levels of pyocyanin and displayed a variety of phenotypes typical of rpoN mutants, including a lack of motility and the failure to grow on nitrate, glutamate, or histidine as the sole nitrogen source. Compared to wild-type PA14, rpoN::Gen(r) was ca. 100-fold less virulent in a mouse thermal injury model and was significantly impaired in its ability to kill the nematode Caenorhabditis elegans. In an Arabidopsis thaliana leaf infectivity assay, although rpoN::Gen(r) exhibited significantly reduced attachment to trichomes, stomata, and the epidermal cell surface, did not attach perpendicularly to or perforate mesophyll cell walls, and proliferated less rapidly in Arabidopsis leaves, it nevertheless elicited similar disease symptoms to wild-type P. aeruginosa PA14 at later stages of infection. rpoN::Gen(r) was not impaired in virulence in a Galleria mellonella (greater wax moth) pathogenicity model. These data indicate that rpoN does not regulate the expression of any genes that encode virulence factors universally required for P. aeruginosa pathogenicity in diverse hosts.     

Degeneration of Axotomized Projection Neurons in the Rat dLGN: Temporal Progression of Events and Their Mitigation by a Single Administration of FGF2

Removal of visual cortex in the rat axotomizes projection neurons in the dorsal lateral geniculate nucleus (dLGN), leading to cytological and structural changes and apoptosis. Biotinylated dextran amine was injected into the visual cortex to label dLGN projection neurons retrogradely prior to removing the cortex in order to quantify the changes in the dendritic morphology of these neurons that precede cell death. At 12 hours after axotomy we observed a loss of appendages and the formation of varicosities in the dendrites of projection neurons. During the next 7 days, the total number of dendrites and the cross-sectional areas of the dendritic arbors of projection neurons declined to about 40% and 20% of normal, respectively. The response of dLGN projection neurons to axotomy was asynchronous, but the sequence of structural changes in individual neurons was similar; namely, disruption of dendrites began within hours followed by cell soma atrophy and nuclear condensation that commenced after the loss of secondary dendrites had occurred. However, a single administration of fibroblast growth factor-2 (FGF2), which mitigates injury-induced neuronal cell death in the dLGN when given at the time of axotomy, markedly reduced the dendritic degeneration of projection neurons. At 3 and 7 days after axotomy the number of surviving dendrites of dLGN projection neurons in FGF-2 treated rats was approximately 50% greater than in untreated rats, and the cross-sectional areas of dendritic arbors were approximately 60% and 50% larger. Caspase-3 activity in axotomized dLGN projection neurons was determined by immunostaining for fractin (fractin-IR), an actin cleavage product produced exclusively by activated caspase-3. Fractin-IR was seen in some dLGN projection neurons at 36 hours survival, and it increased slightly by 3 days. A marked increase in reactivity was seen by 7 days, with the entire dLGN filled with dense fractin-IR in neuronal cell somas and dendrites.     

Phenotype dictates the growth response of vascular smooth muscle cells to pulse pressure in vitro.

The objective of this study was to determine the effect of phenotype on pulse pressure-induced signaling and growth of vascular smooth muscle cells in vitro. Using a perfused transcapillary culture system, cells were exposed to increases in pulsatile flow and hence pulse pressure and maintained for 72 h before cells were harvested. Cell proliferation was determined by cell number, DNA synthesis, and proliferating cell nuclear antigen expression. Mitogen-activated protein kinase (MAPK) levels were determined by immunoblot and kinase activity by phosphorylation of myelin basic protein. Cell phenotype was determined by immunoblot and immunocytofluorescence using antisera specific for the differentiation markers alpha-actin, myosin, calponin, osteopontin, and phospholamban. In cells that highly expressed these differentiation markers, there was a significant increase in cell growth in response to chronic increases in pulse pressure without a significant change in MAPK activity in these cells. In contrast, in cells that weakly expressed SMC differentiation markers, there was a significant decrease in cell growth concomitant with a significant decrease in MAPK signaling in these cells. We conclude that SMC phenotype dictates the growth response of SMC to mechanical force in vitro.     

Host genomic influences on HIV/AIDS

The AIDS era has seen multiple advances in the power of genetics research; scores of host genetic protective factors have been nominated and several have translated to the bedside. We discuss how genomics may inform HIV/AIDS prevention, treatment and eradication.     

Crystals of myohemerythrin

Crystals have been grown of metazide myohemerythrin from the sipunculid Dendrostomum pyroides. The crystals are in space group P2₁2₁2₁ and have unit cell dimensions of $\text{a = 41.6}\text{A}\text{?}\text{, b = 80.0}\text{A}\text{?}\text{and}\text{c = 37.8}\text{A}\text{?}$. An asymmetric unit contains a single myohemerythrin molecule.     

Regional grassland productivity responses to precipitation during multiyear above‐ and below‐average rainfall periods

There is considerable uncertainty in the magnitude and direction of changes in precipitation associated with climate change, and ecosystem responses are also uncertain. Multiyear periods of above‐ and below‐average rainfall may foretell consequences of changes in rainfall regime. We compiled long‐term aboveground net primary productivity (ANPP) and precipitation (PPT) data for eight North American grasslands, and quantified relationships between ANPP and PPT at each site, and in 1–3 year periods of above‐ and below‐average rainfall for mesic, semiarid cool, and semiarid warm grassland types. Our objective was to improve understanding of ANPP dynamics associated with changing climatic conditions by contrasting PPT–ANPP relationships in above‐ and below‐average PPT years to those that occurred during sequences of multiple above‐ and below‐average years. We found differences in PPT–ANPP relationships in above‐ and below‐average years compared to long‐term site averages, and variation in ANPP not explained by PPT totals that likely are attributed to legacy effects. The correlation between ANPP and current‐ and prior‐year conditions changed from year to year throughout multiyear periods, with some legacy effects declining, and new responses emerging. Thus, ANPP in a given year was influenced by sequences of conditions that varied across grassland types and climates. Most importantly, the influence of prior‐year ANPP often increased with the length of multiyear periods, whereas the influence of the amount of current‐year PPT declined. Although the mechanisms by which a directional change in the frequency of above‐ and below‐average years imposes a persistent change in grassland ANPP require further investigation, our results emphasize the importance of legacy effects on productivity for sequences of above‐ vs. below‐average years, and illustrate the utility of long‐term data to examine these patterns.