Effects of high arsenic and fluoride soil concentrations on soybean plants

Arsenic (As) and Fluoride (F) are present in many soils, affecting crops and posing risks in the food chain. We performed pot experiments on spiked soils enriched in these elements either individually or simultaneously, over a wide range of concentrations. Soybean biomass production, grain yield, As and F accumulation and distribution within the plant, and the antioxidant response to these stresses were analyzed. Arsenic was more toxic than F. At As levels >35 mg/kg and F levels >375 mg/kg, yield loss reached 60% and 30%, respectively. At the highest dose of As plants died within 2 weeks, whereas F showed no lethality. When they were applied simultaneously, detrimental effects were more important. As and F in plants increased in all soybean organs although grains presented the lowest concentrations. Antioxidant enzymes were enhanced in plants but this increase was not high enough to cope with the oxidative damage.     

Taxol from fungal endophytes and the issue of biodiversity

Fungi represent one of the most understudied and diverse group of organisms. Commonly, these organisms make associations with higher life forms and may proceed to biochemically mimic the host organism. An excellent example of this is the anticancer drug, taxol, which had been previously supposed to occur only in the plant genusTaxus (yew). However, taxol has been reported in a novel endophytic fungus—Taxomyces andreanae, but also has been demonstrated to occur in a number of unrelated fungal endophytes includingPestalotia, Pestalotiopsis, Fusarium, Alternaria, Pithomyces, Monochaetia and others. Thus, this report presents information on the presence of taxol among disparate fungal genera, and uses these observations as an additional argument to support efforts to study fungal endophytes and preserve their associated host plants.     

Molecular Dynamics Simulation on the Connection Machine

An algorithm for the efficient calculation of macromolecular force fields on the Connection Machine is described. The full force field is separated into bond interactions and non-bonding interactions. Only the latter are implemented on the Connection Machine, the former, less computationally intensive tasks are performed by an existing, conventional molecular dynamics code on the front end. Parallelization of the evaluation of non-bonding interactions is achieved by the Replicated Systolic Loop algorithm introduced in this paper. The algorithm is a variant of the Systolic Loop scheme often used for the computation of 2-particle forces for the classical N-particle problem.     

PKA and Epac cooperate to augment bradykinin-induced interleukin-8 release from human airway smooth muscle cells

Background

Platelet-derived growth factor A (PDGF-A) signals solely through PDGF-Rα, and is required for fibroblast proliferation and transdifferentiation (fibroblast to myofibroblast conversion) during alveolar development, because pdgfa-null mice lack both myofibroblasts and alveoli. However, these PDGF-A-mediated mechanisms remain incompletely defined. At postnatal days 4 and 12 (P4 and P12), using mouse lung fibroblasts, we examined (a) how PDGF-Rα correlates with ki67 (proliferation marker) or alpha-smooth muscle actin (αSMA, myofibroblast marker) expression, and (b) whether PDGF-A directly affects αSMA or modifies stimulation by transforming growth factor beta (TGFβ).

Methods

Using flow cytometry we examined PDGF-Rα, αSMA and Ki67 in mice which express green fluorescent protein (GFP) as a marker for PDGF-Rα expression. Using real-time RT-PCR we quantified αSMA mRNA in cultured Mlg neonatal mouse lung fibroblasts after treatment with PDGF-A, and/or TGFβ.

Results

The intensity of GFP-fluorescence enabled us to distinguish three groups of fibroblasts which exhibited absent, lower, or higher levels of PDGF-Rα. At P4, more of the higher than lower PDGF-Rα + fibroblasts contained Ki67 (Ki67+), and Ki67+ fibroblasts predominated in the αSMA + but not the αSMA- population. By P12, Ki67+ fibroblasts comprised a minority in both the PDGF-Rα + and αSMA+ populations. At P4, most Ki67+ fibroblasts were PDGF-Rα + and αSMA- whereas at P12, most Ki67+ fibroblasts were PDGF-Rα- and αSMA-. More of the PDGF-Rα + than - fibroblasts contained αSMA at both P4 and P12. In the lung, proximate αSMA was more abundant around nuclei in cells expressing high than low levels of PDGF-Rα at both P4 and P12. Nuclear SMAD 2/3 declined from P4 to P12 in PDGF-Rα-, but not in PDGF-Rα + cells. In Mlg fibroblasts, αSMA mRNA increased after exposure to TGFβ, but declined after treatment with PDGF-A.

Conclusion

During both septal eruption (P4) and elongation (P12), alveolar PDGF-Rα may enhance the propensity of fibroblasts to transdifferentiate rather than directly stimulate αSMA, which preferentially localizes to non-proliferating fibroblasts. In accordance, PDGF-Rα more dominantly influences fibroblast proliferation at P4 than at P12. In the lung, TGFβ may overshadow the antagonistic effects of PDGF-A/PDGF-Rα signaling, enhancing αSMA-abundance in PDGF-Rα-expressing fibroblasts.     

Comprehensive mapping of cystic fibrosis mutations to CFTR protein identifies mutation clusters and molecular docking predicts corrector binding site

Cystic Fibrosis (CF) is caused by mutations in the CFTR gene, of which over 2000 have been reported to date. Mutations have yet to be analyzed in aggregate to assess their distribution across the tertiary structure of the CFTR protein, an approach that could provide valuable insights into the structure‐function relationship of CFTR. In addition, the binding site of Class I correctors (VX‐809, VX‐661, and C18) is not well understood. In this study, exonic CFTR mutations and mutant allele frequencies described in 3 curated databases (ABCMdb, CFTR1, and CFTR2, comprising >130 000 data points) were mapped to 2 different structural models: a homology model of full‐length CFTR protein in the open‐channel state, and a cryo‐electron microscopy core‐structure of CFTR in the closed‐channel state. Accordingly, residue positions of 6 high‐frequency mutant CFTR alleles were found to spatially co‐localize in CFTR protein, and a significant cluster was identified at the NBD1:ICL4 interdomain interface. In addition, immunoblotting confirmed the approximate binding site of Class I correctors, demonstrating that these small molecules act via a similar mechanism in vitro, and in silico molecular docking generated binding poses for their complex with the cryo‐electron microscopy structure to suggest the putative corrector binding site is a multi‐domain pocket near residues F374‐L375. These results confirm the significance of interdomain interfaces as susceptible to disruptive mutation, and identify a putative corrector binding site. The structural pharmacogenomics approach of mapping mutation databases to protein models shows promise for facilitating drug discovery and personalized medicine for monogenetic diseases.     

Protein-based phylogenies support a chimeric origin for the eukaryotic genome

The phylogenetic position of the archaebacteria and the place of eukaryotes in the history of life remain a question of debate. Recent studies based on some protein-sequence data have obtained unusual phylogenies for these organisms. We therefore collected the protein sequences that were available with representatives from each of the major forms of life: the gram-negative bacteria, gram-positive bacteria, archaebacteria, and eukaryotes. Monophyletic, unrooted phylogenies based on these sequence data show that seven of 24 proteins yield a significant gram-positive-archaebacteria clade/gram-negative- eukaryotic clade. The phylogenies for these seven proteins cannot be explained by the traditional three-way split of the eukaryotes, archaebacteria, and eubacteria. Nine of the 24 proteins yield the traditional gram-positive-gram-negative clade/archaebacteria-eukaryotic clade. The remaining eight proteins give phylogenies that cannot be statistically distinguished. These results support the hypothesis of a chimeric origin for the eukaryotic cell nucleus formed from the fusion of an archaebacteria and a gram-negative bacteria.      

Phylogenetic analysis of the 90 kD heat shock family of protein sequences and an examination of the relationship among animals, plants, and fungi species

The heat shock protein (Hsp) sequences, because of their ubiquity and high degree of conservation, provide useful models for phylogenetic analysis. In this paper I have carried out a global alignment of all available sequences (a total of 31) for the 90-kD heat shock protein (Hsp90) family. The minimum amino acid identity that is seen between presently known Hsp90 homologs is about 40% over the entire length, indicating that it is a highly conserved protein. Based on the alignment, a number of signature sequences that either are distinctive of the Hsp90 family or that distinguish between the cytosolic and the endoplasmic reticular forms of Hsp90 have been identified. Detailed phylogenetic analyses based on Hsp90 sequences reported here strongly indicate that the cytosolic and the endoplasmic reticulum (ER) resident forms of Hsp90 constitute paralogous gene families which arose by a gene duplication event that took place very early in the evolution of eukaryotic cells. A minimum of two additional gene duplication events, which took place at a later time, are required to explain the presence of two different forms of Hsp90 that are found in fungi and vertebrate species. In a consensus neighbor-joining bootstrap tree based on Hsp90 sequences, plants and animals species grouped together 989 times of 1,000 (a highly significant score), indicating a closer relationship between them as compared to fungi. A closer affiliation of plant and animal species was also observed in the maximum-parsimony tree, although the relationship was not significantly supported by this method. A survey of the recent literature on this subject indicates that depending on the protein sequence and the methods of phylogenetic analysis, the animal species are indicated as closer relatives to either plants or fungi with significant statistical support for both topologies. Thus the relationship among the animal, plant, and fungi kingdoms remains an unresolved issue at the present time.      

Immunohistochemical localization of TLR2 and CD14 in gingival tissue of healthy individuals and patients with chronic periodontitis

We used immunohistochemistry to quantify and compare the expression of Toll-like receptor 2 (TLR2) and cluster of differentiation 14 (CD14) in gingival tissues of both healthy individuals and patients with chronic periodontitis. We also correlated the expression of TLR2 and CD14 with the histological grades of chronic periodontitis. We examined 30 gingival specimens from chronic periodontitis patients and 10 from healthy individuals. Tissues from both groups were immunostained with antibodies against TLR2 and CD14. TLR2 and CD14 were expressed by endothelial cells, fibroblasts, lymphocytes and plasma cells. The immunohistochemical expression of TLR2 and CD14 was significantly greater in inflammatory cells of the chronic periodontitis group than in healthy individuals. Expression of these molecules was greater in the inflammatory cells of connective tissue adjacent to pocket epithelium in both groups. The expression of TLR2 and CD14 was greatest in the periodontitis group that was classified as severe grade, followed by moderate and mild grades, which suggests a role of TLR2 and CD14 in the pathogenesis of chronic periodontitis. The positive correlation of TLR2 and CD14 expression levels with the severity grades of chronic periodontitis suggests that they are correlated also with disease severity; therefore, they may be useful for predicting disease progression. Our findings are consistent with the possibility that CD14 acts as a co-receptor for TLR2.     

Dual role of PKC in modulating pharmacomechanical coupling in fetal and adult cerebral arteries

This study tested the hypothesis that protein kinase C (PKC) has dual regulation on norepinephrine (NE)-mediated inositol 1,4, 5-trisphosphate [Ins (1,4,5)P(3)] pathway and vasoconstriction in cerebral arteries from near-term fetal ( approximately 140 gestational days) and adult sheep. Basal PKC activity values (%membrane bound) in fetal and adult cerebral arteries were 38 +/- 4% and 32 +/- 4%, respectively. In vessels of both age groups, the PKC isoforms alpha, beta(I), beta(II), and delta were relatively abundant. In contrast, compared with the adult, cerebral arteries of the fetus had low levels of PKC-epsilon. In response to 10(-4) M phorbol 12,13-dibutyrate (PDBu; PKC agonist), PKC activity in both fetal and adult cerebral arteries increased 40-50%. After NE stimulation, PKC activation with PDBu exerted negative feedback on Ins(1,4,5)P(3) and intracellular Ca(2+) concentration ([Ca(2+)](i)) in arteries of both age groups. In turn, PKC inhibition with staurosporine resulted in augmented NE-induced Ins(1,4,5)P(3) and [Ca(2+)](i) responses in adult, but not fetal, cerebral arteries. In adult tissues, PKC stimulation by PDBu increased vascular tone, but not [Ca(2+)](i). In contrast, in the fetal artery, PKC stimulation was associated with an increase in both tone and [Ca(2+)](i). In the presence of zero extracellular [Ca(2+)], these PDBu-induced responses were absent in the fetal vessel, whereas they remained unchanged in the adult. We conclude that, although basal PKC activity was similar in fetal and adult cerebral arteries, PKC's role in NE-mediated pharmacomechanical coupling differed significantly in the two age groups. In both fetal and adult cerebral arteries, PKC modulation of NE-induced signal transduction responses would appear to play a significant role in the regulation of vascular tone. The mechanisms differ in the two age groups, however, and this probably relates, in part, to the relative lack of PKC-epsilon in fetal vessels.     

Salivary and serum procalcitonin and C-reactive protein as biomarkers of periodontitis in United States veterans with osteoarthritis or rheumatoid arthritis

Serum procalcitonin (ProCT) is elevated in response to bacterial infections, whereas high sensitivity C-reactive protein (hsCRP) is a nonspecific inflammatory marker that is increased by excess adipose tissue. We examined the efficacy of ProCT and hsCRP as biomarkers of periodontitis in the saliva and serum of patients with arthritis, which is characterized by variable levels of systemic inflammation that potentially can confound the interpretation of inflammatory biomarkers. Blood and unstimulated whole saliva were collected from 33 patients with rheumatoid arthritis (RA) and 50 with osteoarthritis (OA). Periodontal status was assessed by full mouth examination and patients were categorized as having no/mild, moderate or severe periodontitis by standard parameters. Salivary and serum ProCT and hsCRP concentrations were compared. BMI, diabetes, anti-inflammatory medications and smoking status were ascertained from the patient records. Differences between OA and RA in proportionate numbers of patients were compared for race, gender, diabetes, adiposity and smoking status. Serum ProCT was significantly higher in arthritis patients with moderate to severe and severe periodontitis compared with no/mild periodontitis patients. There were no significant differences in salivary ProCT or salivary or serum hsCRP in RA patients related to periodontitis category. Most of the OA and RA patients were middle aged or older, 28.9% were diabetic, 78.3% were overweight or obese, and slightly more than half were either current or past smokers. The OA and RA groups differed by race, but not gender; blacks and males were predominant in both groups. The OA and RA groups did not differ in terms of controlled or uncontrolled diabetes, smoking status or BMI. The RA patients had been prescribed more anti-inflammatory medication than the OA patients. Our results demonstrate that circulating ProCT is a more discriminative biomarker for periodontitis than serum hsCRP in patients with underlying arthritis. Any elevation in salivary and serum hsCRP due to periodontitis apparently was overshadowed by differences among these patients in factors that influence CRP, such as the extent of inflammation between RA and OA, the extent of adipose tissue, the use of anti- inflammatory medications and smoking status. Although our study showed no differences in salivary ProCT related to severity of periodontitis, this biomarker also may be useful with further refinement.