A simple quantitative method for the estimation of free ecto-sulfhydryl groups of spermatozoa

A simple rapid quantitative method has been developed for the estimation of sperm ecto-SH groups on the basis of their high affinity binding to the mercurial: [203Hg]p-chloromercuriphenylsulfonic acid (PCMPS) used as a surface probe. The thiol reagent did not penetrate the sperm plasma membrane, as evidenced by the extremely rapid time course of the binding reaction and undetectable uptake of [203Hg]PCMPS by intact goat spermatozoa. The binding reaction was not due to contaminating broken or damaged cells, if any. The method consists of incubating of highly motile goat spermatozoa with PCMPS in a modified Ringer solution at 37 degrees C for 5 min, agglutination of the labelled cells with polyethyleneimine (100 micrograms/ml) and filtration and washing of the cell suspension through Whatman No. 1 filter discs under mild vacuum. The binding interaction is proportional to cell concentration, specific and saturable at 50 microM PCMPS. The method is capable of estimating free ecto-SH as low as 25 pmoles. Spermatozoa possess 286 +/- 61 pmoles of free ecto-SH groups/10(6) cells. Scatchard analysis showed the presence in goat spermatozoa of multiple classes of ecto-SH groups differing in their affinity for PCMPS.     

Prioritizing Tiger Conservation through Landscape Genetics and Habitat Linkages

Even with global support for tiger (Panthera tigris) conservation their survival is threatened by poaching, habitat loss and isolation. Currently about 3,000 wild tigers persist in small fragmented populations within seven percent of their historic range. Identifying and securing habitat linkages that connect source populations for maintaining landscape-level gene flow is an important long-term conservation strategy for endangered carnivores. However, habitat corridors that link regional tiger populations are often lost to development projects due to lack of objective evidence on their importance. Here, we use individual based genetic analysis in combination with landscape permeability models to identify and prioritize movement corridors across seven tiger populations within the Central Indian Landscape. By using a panel of 11 microsatellites we identified 169 individual tigers from 587 scat and 17 tissue samples. We detected four genetic clusters within Central India with limited gene flow among three of them. Bayesian and likelihood analyses identified 17 tigers as having recent immigrant ancestry. Spatially explicit tiger occupancy obtained from extensive landscape-scale surveys across 76,913 km² of forest habitat was found to be only 21,290 km². After accounting for detection bias, the covariates that best explained tiger occupancy were large, remote, dense forest patches; large ungulate abundance, and low human footprint. We used tiger occupancy probability to parameterize habitat permeability for modeling habitat linkages using least-cost and circuit theory pathway analyses. Pairwise genetic differences (F _ST) between populations were better explained by modeled linkage costs (r>0.5, p<0.05) compared to Euclidean distances, which was in consonance with observed habitat fragmentation. The results of our study highlight that many corridors may still be functional as there is evidence of contemporary migration. Conservation efforts should provide legal status to corridors, use smart green infrastructure to mitigate development impacts, and restore habitats where connectivity has been lost.     

Dsi-1, a region with frequent proviral insertions in Moloney murine leukemia virus-induced rat thymomas.

Dsi-1 is a region of chromosomal DNA that underwent proviral insertion in 3 of 24 Moloney murine leukemia virus-induced rat thymomas. In one of these tumors, a provirus is also integrated adjacent to the proto-oncogene c-myc. The proviruses in Dsi-1 have been characterized and appear to be complete. The proviruses were located within a 2-kilobase region that contained four prominent DNase I-hypersensitive sites. These hypersensitive sites were observed in Moloney murine leukemia virus-induced thymomas but not in NRK cells. The region of Dsi-1 immediately 3' to the insertions cross-hybridized with human and chicken DNA, indicating that it contains highly conserved sequences. No evidence could be found for the expression of this highly conserved region. Dsi-1 was mapped to mouse chromosome 4. This location demonstrates that Dsi-1 is different from 16 of the known proto-oncogenes (c-abl, c-erbA c-erbB, c-ets-1, c-ets-2, c-fes, c-fos, c-myb, c-myc, c-raf, A-raf, c-Ha-ras, c-Ki-ras, N-ras, c-sis, and c-src) and 12 cellular regions of tumor-associated integrations in retrovirus-induced tumors (c-erbB, Fis-1, int-1, int-2, Mis-1/pvt-1, Mlvi-1, Mlvi-2, c-mos, c-myb, c-myc, Pim-1, and c-Ha-ras). Hybridization experiments indicated that Dsi-1 is probably different from five additional proto-oncogenes (c-fgr, c-fms, c-mos, neu, and c-yes) and from two additional frequent integration regions (lck and Mlvi-3).     

Bioenergetic consequences of protein overexpression in Saccharomyces cerevisiae

Summary Some bioenergetic consequences of overexpression of plasmid-encoded homologous (phosphoglycerate kinase), and heterologous (prochymosin), protein in S. cerevisiae strains grown in chemostat culture have been investigated. Both overexpressing strains were found to exhibit similar fermentation patterns despite a 10-fold difference in product expression levels. Biomass yields were lower than those for a control strain, and the onset of oxido-fermentative metabolism occurred at a lower dilution rate. A marked rise in cellular ATP content with increasing dilution rate during oxidative growth was observed in the strain overexpressing yeast phosphoglycerate kinase (PGK); this at present cannot be adequately explained. The inorganic phosphate content of the overexpressing strains was higher than that of the control and the phosphorylation potential of the prochymosin expressing strain was up to 10-fold lower than both the control and PGK overexpressing strains. It is proposed that expression of heterologous prochymosin imposes a greater energy drain on the host than overexpression of homologous PGK. This energetic drain may be a limiting factor in heterologous gene expression.     

Transport to the cell surface of a peptide sequence attached to the truncated C terminus of an N-terminally anchored integral membrane protein.

Attempts to construct hybrid proteins that are transported to the plasma membrane are frequently unsuccessful because of perturbations in polypeptide folding. In seeking to minimize this problem, we have used the less common type of integral membrane protein, which has an uncleaved signal-anchor domain and an extracellular carboxyl portion, to transport a peptide sequence of interest to the cell surface. A set of plasmids was constructed that contained the gene encoding respiratory syncytial virus glycoprotein G (RSVG) interrupted immediately after one of several proline codons by a synthetic sequence containing unique restriction endonuclease sites and a stop codon. The shortened RSVG gene was flanked by vaccinia virus DNA to permit cloning and expression in a vaccinia virus vector. An open reading frame encoding four copies of the immunodominant repeating epitope of the circumsporozoite protein of Plasmodium falciparum was inserted into the tails of the truncated RSVG genes. Recombinant vaccinia viruses were isolated and shown to express hybrid proteins that reacted with a monoclonal antibody directed to the repeating circumsporozoite epitope. Moreover, immunofluorescence studies indicated that the peptide was on the external cell surface and available to react with antibodies. Expression of the hybrid protein also occurred in rabbits inoculated with the live recombinant vaccinia virus, as demonstrated by the generation of antibodies that bound to P. falciparum sporozoites in vitro.     

Cytochrome c: ion binding and redox properties. Studies on ferri and ferro forms of horse, bovine, and tuna cytochrome c

The ion binding properties of horse, bovine, and tuna cytochrome c (both oxidized and reduced) have been measured using a combination of ultrafiltration, neutron activation, and ion chromatography. The ions investigated were chloride, phosphate, and Tris-cacodylate. Ion chromatography and neutron activation analysis techniques were employed to determine the concentration of free anions. Binding constants are obtained from modified Scatchard plots (in the range of 10-2000 M-1). The redox potentials for cytochrome c at different ionic strengths, pH 7.0, have been determined. In this paper we report the ionic strength and ion binding effects on the redox properties of horse, bovine, and tuna cytochrome c. Potential versus ionic strength dependence for horse, bovine, and tuna cytochrome c from the experimental data were compared with a theoretical model.     

Abortion in Canada: religious and ideological dimension's of women's attitudes

This paper examines a number of demographic and sociocultural factors (e.g., age, marital status, family size, religion, religious assiduity, sex-role ideology) as predictors of women's attitudes toward abortion, using data from the Canadian Fertility Survey of 1984. The findings suggest that women's abortion attitudes are to a greater extent based on ideological positions. It appears that anti-abortion stance affects those women who are religious, presumably by increasing the relationship between their general sex-role ideological stances and abortion attitudes. Abortion attitudes also vary according to a woman's education, her size, and province/region of residence.     

Physical mapping of two Xp markers DXS16 and DXS143

Summary Lymphocyte karyotyping of an infant girl with the clinical features of microphthalmia, iridoschisis, goiter, hip joint dysplasia, labium synechia and craniotabes revealed an Xp deletion. The lymphocyte karyotypes of the parents were normal. Bromodeoxyuridine incorporation studies showed that, in 42 out of 43 metaphases, the deleted X chromosome was late replicating. In one metaphase, the normal X chromosome was observed to be allocyclic. Using DNA markers from the Xp22 region, the breakpoint was assigned distal to DXS16 (pXUT23) and proximal to DXS143 (dic56). Dosage intensity measurements confirmed that the STS gene and the DNA marker DXS31 were involved in the deleted area. Restriction fragment length polymorphism analysis revealed that the paternally derived X-chromosome was deleted.     

Bimodal oxygen uptake in a freshwater air-breathing fish,Notopterus chitala

Measurements of bimodal oxygen uptake have been made in a freshwater air-breathing fish,Notopterus chitala at 29.0±1(S.D.)°C. xhe mean oxygen uptake from continuously flowing water without any access to air, was found to be 3.58±0.37 (S.E.) ml O₂ · h^?1 and 56.84+4.29 (S.E.) ml O₂ · kg^?1 · h^?1 for a fish weighing 66.92 + 11.27 (S.E.) g body weight. In still water with access to air, the mean oxygen uptake through the gills were recorded to be 2.49 ± 0.31 (S.E.) ml O₂ · h^?1 and 38.78 ± 1.92 (S.E.) ml O₂ · kg^?1 · h^?1 and through the accessory respiratory organs (swim-bladder) 6.04±0.87 (S.E.) ml O₂ · h^?1 and 92.32±2.91 (S.E.) ml O₂ · kg^?1 · h^?1 for a fish averaging 66.92±11.27 (S.E.) g. Out of the total oxygen uptake (131.10 ml O₂ · kg^?1 · h^?1), about 70% was obtained through the aerial route and the remainder 30% through the gills.