Mutagenic effects of carbosulfan,a carbamate pesticide

The genotoxic effects of carbosulfan were evaluated using chromosome aberration (CA), bone marrow micronucleus (MN) and sperm abnormality assays in mice. All the three acute doses (1.25, 2.5 and 5mg/kg) of carbosulfan induced significant dose-dependent increase in the frequency of CA (P<0.02), micronucleated polychromatic erythrocytes (PCEs) (P<0.05) and sperm head abnormalities (P<0.05) but did not affect the total sperm count. The highest acute dose of carbosulfan induced >7-fold increase in the frequency of CA, >3.5-fold increase in the frequency of micronucleated PCEs and >4.6-fold increase in the frequency of sperms with abnormal head morphology following intraperitoneal exposure as compared to the untreated controls. The present findings suggest that carbosulfan is a potent genotoxic agent and may be regarded as a potential germ cell mutagen also.     

Structure and reorientational dynamics of angiotensin I and II: a microscopic physical insight

We present a study of structural analysis and reorientational dynamics of Angiotensin I (AngI) and Angiotensin II (AngII) in aqueous solution. AngI is a decapeptide that acts as a precursor to the octapeptide AngII in the Renin-Angiotensin-Aldosterone system for blood pressure regulation. Experimental structural characterization of these peptides, carried out with circular dichroism and infrared spectroscopy, showed that the angiotensins are mostly disordered but exhibit a measurable population of ordered structures at room temperature. Interestingly, these change from the unordered polyproline-like conformation for AngI to a more compact and ordered conformation for AngII as the length of the peptide is decreased. Anisotropy decay measurements with picosecond time resolution indicate slower overall tumbling and a greater amplitude of internal motion in AngI compared to AngII, consistent with more compact and less flexible structure of the active form of the peptide. To model the microscopic behavior of the peptides, 2-μs molecular dynamics simulation trajectories were generated for AngI and AngII, at 300?K using the OPLS-AA potential and SPC water. The structures sampled in the simulations mostly agree with the experimental results, showing the prevalence of disordered structures, turns, and polyproline helices. Additionally, the computational results predict fewer sampled conformations, tighter side-chain packing and marked increase of Phe8 solvent accessibility upon AngI truncation to AngII. Our combined approach of experiment and extensive computer simulation thus yields new information on the conformational dynamics of the angiotensins, helping provide insight into the structural basis for the potency of AngI relative to AngII.     

ReducedS-adenosylmethionine: Protein-lysineN-methyltransferase activity (protein methylase III) in shiverer mutant mouse brain

Mice with the dysmyelinating mutation shiverer were studied by measuring the activity of two protein methylases and myelin marker enzymes in the brain. It was observed thatS-adenosylmethionine: protein-lysineN-methyltransferase (protein methylase III, EC. 2.1.1.43) activity is significantly reduced in phenotypically affected homozygous shiverer (shi/shi) mutant mouse brain compared to the unaffected heterozygous littermate brain. This reduction in enzyme activity is manifested mainly by reduced formation of trimethyllysine during the in vitro methylation of histone. In contrast, myelin marker enzymes such as 2,3-cyclic nucleotide 3-phosphohydrolase and 5-nucleotidase as well asS-adenosyl-methionine: protein-carboxylO-methyltransferase (protein methylase II, EC. 2.1.1.24) activities were not significantly affected in these strains of mice.     

Binding to syntenin-1 protein defines a new mode of ubiquitin-based interactions regulated by phosphorylation

Syntenin-1 is a PDZ domain-containing adaptor that controls trafficking of transmembrane proteins including those associated with tetraspanin-enriched microdomains. We describe the interaction of syntenin-1 with ubiquitin through a novel binding site spanning the C terminus of ubiquitin, centered on Arg(72), Leu(73), and Arg(74). A conserved LYPSL sequence in the N terminus, as well as the C-terminal region of syntenin-1, are essential for binding to ubiquitin. We present evidence for the regulation of this interaction through syntenin-1 dimerization. We have also established that syntenin-1 is phosphorylated downstream of Ulk1, a serine/threonine kinase that plays a critical role in autophagy and regulates endocytic trafficking. Importantly, Ulk1-dependent phosphorylation of Ser(6) in the LYPSL prevents the interaction of syntenin-1 with ubiquitin. These results define an unprecedented ubiquitin-dependent pathway involving syntenin-1 that is regulated by Ulk1.     

Use of growth indices from radiometric culture for quantification of sheep strains of Mycobacterium avium subsp. paratuberculosis

A simple method for using growth indices from radiometric BACTEC cultures was evaluated for the enumeration of Australian sheep strains of Mycobacterium avium subsp. paratuberculosis. The numbers of viable organisms in inocula were determined by end-point titration in BACTEC cultures. Growth indices were measured by using a BACTEC 460 machine. There was a linear relationship between the number of days taken for the cumulative growth index to reach 1,000 (dCGI1000) and log(10) inoculum size. The use of dCGI1000 was shown to be as effective as the use of growth index data from the entire growth cycle for the estimation of inoculum size. For particular isolates characterized by end-point titration, the dCGI1000 of a single BACTEC vial provided estimates of viable numbers within narrow prediction limits. Predictive relationships were also established for the enumeration of M. avium subsp. paratuberculosis from field samples by using the dCGI1000 of a single BACTEC vial, with prediction limits of +/-1 to 2 log units. Organisms from feces or contaminated soil grew more slowly than those from cultures or tissues, and separate equations were developed for enumeration from these sources.     

Transgenic mice expressing lipoprotein lipase in adipose tissue. Absence of the proximal 3'-untranslated region causes translational upregulation

Lipoprotein lipase (LPL) is a key enzyme in lipoprotein and adipocyte metabolism. Defects in LPL can lead to hypertriglyceridemia and the subsequent development of atherosclerosis. The mechanisms of regulation of this enzyme are complex and may occur at multiple levels of gene expression. Because the 3'-untranslated region (UTR) is involved in LPL translational regulation, transgenic mice were generated with adipose tissue expression of an LPL construct either with or without the proximal 3'-UTR and driven by the aP2 promoter. Both transgenic mouse colonies were viable and expressed the transgene, resulting in a 2-fold increase in LPL activity in white adipose tissue. Neither mouse colony exhibited any obvious phenotype in terms of body weight, plasma lipids, glucose, and non-esterified fatty acid levels. In the mice expressing hLPL with an intact 3'-UTR, hLPL mRNA expression approximately paralleled hLPL activity. However in the mice without the proximal 3'-UTR, hLPL mRNA was low in the setting of large amounts of hLPL protein and LPL activity. In previous studies, the 3'-UTR of LPL was critical for the inhibitory effects of constitutively expressed hormones, such as thyroid hormone and catecholamines. Therefore, these data suggest that the absence of the 3'-UTR results in a translationally unrepressed LPL, resulting in a moderate overexpression of adipose LPL activity.     

Reduced kidney lipoprotein lipase and renal tubule triglyceride accumulation in cisplatin-mediated acute kidney injury

Peroxisome proliferator-activated receptor-α (PPARα) activation attenuates cisplatin (CP)-mediated acute kidney injury by increasing fatty acid oxidation, but mechanisms leading to reduced renal triglyceride (TG) accumulation could also contribute. Here, we investigated the effects of PPARα and CP on expression and enzyme activity of kidney lipoprotein lipase (LPL) as well as on expression of angiopoietin protein-like 4 (Angptl4), glycosylphosphatidylinositol-anchored-HDL-binding protein (GPIHBP1), and lipase maturation factor 1 (Lmf1), which are recognized as important proteins that modulate LPL activity. CP caused a 40% reduction in epididymal white adipose tissue (WAT) mass, with a reduction of LPL expression and activity. CP also reduced kidney LPL expression and activity. Angptl4 mRNA levels were increased by ninefold in liver and kidney tissue and by twofold in adipose tissue of CP-treated mice. Western blots of two-dimensional gel electrophoresis identified increased expression of a neutral pI Angptl4 protein in kidney tissue of CP-treated mice. Immunolocalization studies showed reduced staining of LPL and increased staining of Angptl4 primarily in proximal tubules of CP-treated mice. CP also increased TG accumulation in kidney tissue, which was ameliorated by PPARα ligand. In summary, a PPARα ligand ameliorates CP-mediated nephrotoxicity by increasing LPL activity via increased expression of GPHBP1 and Lmf1 and by reducing expression of Angptl4 protein in the proximal tubule.     

Effect of Arsenic on Nodulation and Nitrogen Fixation of Blackgram (<Emphasis Type="Italic">Vigna mungo</Emphasis>)

Rhizobium–legume symbiotic interaction is an efficient model system for soil remediation and reclamation. We earlier isolated an arsenic (As) (2.8 mM arsenate) tolerant and symbiotically effective Rhizobium strain, VMA301 from Vigna mungo and in this study we further characterized its efficacy for arsenic removal from the soil and its nitrogen fixation capacity. Although nodule formation is delayed in plants with As-treated composite when the inoculum was prepared without arsenic in culture medium, whereas it attains the significant number of nodules compare to plant grown in As-free soil when the inoculum was prepared with arsenic supplemented medium. Arsenic accumulation was higher in roots than root nodules. Nitrogenase activity is reduced to almost 2 fold in plants with As-treated soil but not abolished. These results suggest that this strain, VMA301, has been able to establish an effective symbiotic interaction in V. mungo in As-contaminated soil and can perform dual role of arsenic bioremediation as well as soil nitrogen improvement.