Calpastatin is upregulated in non-immune neuronal cells via toll-like receptor 2 (TLR2) pathways by lipid-containing agonists

Calpain (intracellular Ca^2 +-dependent protease) and calpastatin (calpain specific endogenous inhibitor) are widely distributed in biological systems, and have been implicated in many cellular physiological and pathological processes. Calpastatin level is of central importance to the control of calpain activity. We demonstrated for the first time that calpastatin is overexpressed in mycoplasma-contaminated cultured cells (SH-SY5Y cells that are infected by a strain of Mycoplasma hyorhinis (NDMh)). We have found that the calpastatin-upregulating activity resides in the mycoplasmal membrane lipoproteins, and is associated with NF-κB activation. Calpain-promoted proteolysis is attenuated in the NDMh lipoprotein-treated cells. Here we show that the NDMh lipoproteins promoted an increase in calpastatin in SH-SY5Y cells via the TLR2/TAK1/NF-κB pathway. The synthetic mycoplasmal lipopeptide MALP-2 and the bacterial lipopeptide PAM3CSK4 (TLR2 agonists) also promoted calpastatin upregulation. LPS (TLR4 agonist) activated NF-κB without calpastatin increase in the cell. In contrast, lipoteichoic acid (TLR2 agonist) upregulated calpastatin not via NF-κB activation, but via the MEK1/ELK1 pathway. Zymosan and peptidoglycan, TLR2 agonists that lack lipids, did not induce calpastatin upregulation. Cell treatment with a calpastatin-upregulating agonist (lipoteichoic acid) led to the attenuation of Ca^2 +-promoted calpain activity, whereas agonists that do not upregulate calpastatin (LPS, Zymosan) were ineffective. Overall, the results indicate that in these non-immune cells, calpastatin is upregulated by TLR2-agonists containing lipids, with more than one downstream pathway involved. Such agonists may be useful for studying mechanisms and factors involved in calpastatin regulation. In addition, suitable TLR2 agonists may be of interest in devising treatments for pathological processes involving excessive calpain activation.     

Eumycetoma caused by Curvularia lunata in a dog

Curvularia lunata was cultured from black granules found in granulomatous tumefactions excised from the subcutis of a three year old Medium Schnauzer dog. Draining sinuses were present in some of the tumefactions. Accordingly the diagnosis of eumycotic mycetoma was made. This diagnosis was confirmed by histopathological examination. During the four years following the first surgical intervention, several more similar tumefactions were excised on three different occasions. The dog died of chronic renal failure at the age of 8 years. There was no bone involvement or visceral diffusion of the fungus. The granules were examined by scanning electron microscopy. Immunoglobulins in the dog's serum, assessed by a qualitative test, proved to be equal to immunoglobulins in the serum of a control dog. Precipitating antibodies against C. lunata were not found. The dog was treated for 150 days with itraconazole. In spite of good initial results, recurrence of the fungal lesions were observed after the treatment's interruption. Further treatment with itraconazole for 45 days proved ineffective. No side effects of the drug were observed. This is, to the best of our knowledge, the first case in which C. lunata is identified as the causative agent of an animal eumycetoma.     

The role of the thymus in the establishment of suppressor cells in liver chimeras.

The role of the thymus in the establishment of specific suppression was studied in an experimental model in which lethally irradiated F1 mice were reconstituted with parental C57BL neonatal liver cells and then challenged with immunocompetent spleen cells syngeneic to the donor. In this model, inhibition of a graft-versus-host response by these spleen cells served as an indication of the establishment of a suppressive state towards syngeneic antigens in the liver chimeras. It was demonstrated that since thymectomy of the chimeras could not prevent the elicitation of a graft-versus-host response by spleen cells, the thymus is essential for the establishment of this specific suppression. Reconstitution of such chimeras with intact thymus grafts of either donor (C57BL) or host (F1) origin led to the inhibition of the graft-versus-host response and to the reappearance of the suppressive state. Removal of the thymus in intact liver chimeras after establishment of a suppressive state did not affect suppression. Thus, it was concluded that the thymus is needed in the chimeras during a critical period in the development of suppression. Once suppression is established, the presence of the thymus is no longer required.     

Elimination of suppressor T-cells in mice undergoing a graft-versus-host reaction expressed by increased response to polyvinylpyrrolidone (PVP).

Mice undergoing a mild GVHR exhibited an enhanced response to PVP considered to be independent of T-helper function and simultaneously, a decreased response to SRBC, known to be T-cell dependent. Such mice had a reduced number of T cells in their spleens expressed by a lower reactivity to T-lectins PHA and Con A and by a decrease in the number of cells killed by anti θ serum. These mice did not show, however, a substantial change in the activity of B lymphocytes contained in their spleens, since the PFC and the mitogenic response to LPS, as well as the number of immunoglobulin bearing cells were similar to that of controls. These results suggest that the enhanced response to PVP in mice undergoing a mild GVHR is a result of the elimination of a certain T-cell population which seems to regulate the immune response to this antigen.     

Dominant plants alter the microclimate along a fog gradient in the Atacama Desert

Plant Ecology - We assessed the impact of fog on microclimate in a poorly understood fog desert under two common nurse plant species, at three sites: (1) foggy coast, (2) intermediate, and (3)...     

Saguaro (Carnegiea gigantea, Cactaceae) age-height relationships and growth: the development of a general growth curve

Because the growth rate of saguaros varies across different environments, past studies on saguaro population structure required extensive data collection (often over many decades) followed by site-specific analysis to estimate age at the sampled locale. However, when height-growth data from different populations are compared, the overall shape of the growth curves is similar. In this study, a formula was developed to establish saguaro age-height relationships (using stepwise regression) that can be applied to any saguaro population and only requires a site-specific factor to adjust the curve to the local growth rate. This adjustment factor can be established more efficiently and requires less data than the full analyses required for previous studies. Saguaro National Park East (SNP-E) was used as the baseline factor, set to 1.0. Results yielded a factor of 0.743 for SNP West. When the formula was applied to 10-yr interval data from Organ Pipe Cactus National Monument (OPCNM) in Arizona, USA, this location had a factor of 0.617 (relative to SNP-E). With this formula and relatively little field sampling, the age of any individual saguaro (whether the individual was sampled or not) in any population can be estimated.     

Normal mode analysis suggests a quaternary twist model for the nicotinic receptor gating mechanism

We present a three-dimensional model of the homopentameric alpha7 nicotinic acetylcholine receptor (nAChR), that includes the extracellular and membrane domains, developed by comparative modeling on the basis of: 1), the x-ray crystal structure of the snail acetylcholine binding protein, an homolog of the extracellular domain of nAChRs; and 2), cryo-electron microscopy data of the membrane domain collected on Torpedo marmorata nAChRs. We performed normal mode analysis on the complete three-dimensional model to explore protein flexibility. Among the first 10 lowest frequency modes, only the first mode produces a structural reorganization compatible with channel gating: a wide opening of the channel pore caused by a concerted symmetrical quaternary twist motion of the protein with opposing rotations of the upper (extracellular) and lower (transmembrane) domains. Still, significant reorganizations are observed within each subunit, that involve their bending at the domain interface, an increase of angle between the two beta-sheets composing the extracellular domain, the internal beta-sheet being significantly correlated to the movement of the M2 alpha-helical segment. This global symmetrical twist motion of the pentameric protein complex, which resembles the opening transition of other multimeric ion channels, reasonably accounts for the available experimental data and thus likely describes the nAChR gating process.     

Protein secondary structure assignment revisited: a detailed analysis of different assignment methods

Background  

A number of methods are now available to perform automatic assignment of periodic secondary structures from atomic coordinates, based on different characteristics of the secondary structures. In general these methods exhibit a broad consensus as to the location of most helix and strand core segments in protein structures. However the termini of the segments are often ill-defined and it is difficult to decide unambiguously which residues at the edge of the segments have to be included. In addition, there is a "twilight zone" where secondary structure segments depart significantly from the idealized models of Pauling and Corey. For these segments, one has to decide whether the observed structural variations are merely distorsions or whether they constitute a break in the secondary structure.