Effects of colchicine on anther and microspore culture of bread wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

The aim of this work was to study the effects of colchicine application on chromosome doubling and androgenic response in anther and microspore culture of different bread wheat genotypes. Colchicine was applied during a mannitol stress pretreatment or during the first 48 h of culture at concentrations of 0, 150 and 300 mg l⁻¹. When colchicine was applied during stress pretreatment, the percentage of doubling depended on genotype and concentration. A significant increase in doubling was observed with 300 mg l⁻¹ in the low androgenic responding cv. Caramba. Colchicine incorporation during the first hours of culture improved percentage of doubling in all genotypes, in both anther and microspore culture. Application of 300 mg l⁻¹ colchicine improved the percentage of doubling in the two low responding genotypes, to 118% of control in DH24033, and 75% in Caramba in microspore and anther culture, respectively. Concerning the androgenic response, the effect of colchicine on embryo formation and percentage of green plants depended on the genotype and on the culture method. In cv. Pavon, a 2- and a 3-fold increase in percentage of embryogenesis and green plants, respectively, were obtained with 300 mg l⁻¹ colchicine in microspore culture. However, no significant differences in these two variables were observed in anther culture. The number of green doubled haploid (DH) plants reflects the index of success of the procedure. Regardless of the culture method, when colchicine was incorporated during the first hours of culture, the number of green DH plants increased significantly in three of four genotypes. These results confirm the usefulness of colchicine application during the first hours of culture in wheat breeding programs.     

Familial breast/ovarian cancer and BRCA1/2 genetic screening: the role of immunohistochemistry as an additional method in the selection of patients.

Only 20-25% of families screened for BRCA1/2 mutations are found positive. Because only a positive result is informative, we studied the role of BRCA1/2 immunohistochemistry as an additional method for patient selection. From 53 high-risk-affected probands, 18 (34%) had available paraffin blocks of their tumors and were selected for this study. Mutation screening was done by conformation-sensitive gel electrophoresis and multiplex ligation-dependent probe amplification. For immunohistochemistry, 21 neoplastic specimens (15 breast carcinomas, 5 ovary neoplasms, and 1 rectal adenocarcinoma) were analyzed with BRCA1 (monoclonal antibody, Ab-1, oncogene) and BRCA2 (polyclonal antibody, Ab-2, oncogene) antibodies. Absence of the BRCA1 protein was confirmed in negative tumors by Western blotting. Seven patients were positive for BRCA1/2 mutations: 5 for BRCA1 and 2 for BRCA2. Four out of five positive patients had tumors negative for BRCA1 immunostaining, and the remaining 13 BRCA1-negative patients had positive BRCA1 immunostaining in all tumor samples. Sensitivity to predict for BRCA1 mutation carriers was 80%, and specificity was 100%, with a positive predictive value of 100% and a negative predictive value of 93%. This correlation was statistically significant (p=0.001). No correlation was observed for BRCA2. If larger studies confirm these results, high-risk patients with BRCA1-negative tumors should be screened first for this gene.     

Use of textile waste water along with liquid NPK fertilizer for production of wheat on saline sodic soils

A field experiment in collaboration with a private textile industry (Noor Fatima Fabrics Private (Ltd.), Faisalabad) was conducted to evaluate the effect of disposed water from bleaching unit, printing unit and end drain for improving growth and yield of wheat under saline sodic soil. Textile waste water along with canal water (control) was applied with and without liquid NPK fertilizer. The application of liquid NPK fertilizer with end drain waste water increased plant height, spike length, flag leaf length, root length, number of tillers (m^?2), number of fertile tillers (m^?2), 1000 grain weight, grain yield, straw yield and biological yield up to 21, 20, 20, 44, 17, 20, 14, 44, 40 and 41%, respectively compared to canal water (control). Similarly, the NPK uptake in grain was increased up to 15, 30 and 28%, respectively by liquid fertilizer treated end drain water as compare to canal water with liquid fertilizer. Moreover, concentration of different heavy metals particularly Cu, Cr, Pb and Cd was decreased in grains by application of waste water along with liquid NPK. The result may imply that waste water application along with liquid-NPK could be a novel approach for improving growth and yield of wheat in saline sodic soils.     

A conserved sequence in caveolin-1 is both necessary and sufficient for caveolin polarity and cell directional migration

Caveolin-1 (Cav-1) plays an important role in the organization of signaling molecules involved in a variety of signaling pathways, including those mediating cell motility. Here we show that amino acids K47-K57 of Cav-1 are a highly conserved sequence in Cav-1 and Cav-3 proteins, and that expression of either K47-K57 deletion Cav-1 mutant or wild-type Cav-2 that lacks this sequence exhibits a non-polarized distribution pattern. Expression of K47-K57 in Cav-2 leads to Cav-2 polarity, suggesting that expression of K47-K57 is sufficient to direct caveolin polarity. Importantly, we show that expression of this sequence is both necessary and sufficient to promote cell directional migration. Thus, our results support the conclusion that Cav-1 polarity is critical for cell directional migration.     

Pycnogenol® and vitamin E inhibit ethanol‐induced apoptosis in rat cerebellar granule cells

Pycnogenol® (PYC), a patented combination of bioflavonoids extracted from the bark of French maritime pine (Pinus maritima), scavenges free radicals and promotes cellular health. The protective capacity of PYC against ethanol toxicity of neurons has not previously been explored. The present study demonstrates that in postnatal day 9 (P9) rat cerebellar granule cells the antioxidants vitamin E (VE) and PYC (1) dose dependently block cell death following 400, 800, and 1600 mg/dL ethanol exposure (2) inhibit the ethanol‐induced activation of caspase‐3 in the same model system; and (3) reduce neuronal membrane disruption as assayed by phosphatidylserine translocation to the cell surface. These results suggest that both PYC and VE have the potential to act as therapeutic agents, antagonizing the induction of neuronal cell death by ethanol exposure. © 2004 Wiley Periodicals, Inc. J Neurobiol 59: 261–271, 2004     

Ghrelin O-acyltransferase (GOAT) has a preference for n-hexanoyl-CoA over n-octanoyl-CoA as an acyl donor

Ghrelin is a peptide hormone in which serine 3 is modified by n-octanoic acid through GOAT (ghrelin O-acyltransferase). However, the enzymological properties of GOAT remain to be elucidated. We analyzed the in vitro activity of GOAT using the recombinant enzyme. Unexpectedly, although the main active form of ghrelin is modified by n-octanoic acid, GOAT had a strong preference for n-hexanoyl-CoA over n-octanoyl-CoA as an acyl donor. Moreover, a four-amino acid peptide derived from the N-terminal sequence of ghrelin can be modified by GOAT, indicating that these four amino acids constitute the core motif for substrate recognition by the enzyme.     

Structural-Thermodynamic Relationships of Interactions in the N-Terminal ATP-Binding Domain of Hsp90

Despite its importance as a target in anti-cancer therapeutics and the numerous rational-based inhibitor design efforts aimed at it, there are only limited data available on structural-thermodynamic relationships of interactions of the N-terminal ATP-binding domain of Hsp90 (N-Hsp90). Here, we redress this by presenting an investigation of binding of nucleotides and ansamycin compounds to this domain. Interactions of nucleotides with N-Hsp90 are relatively weak (> 10 μM) and are strongly enthalpy driven over the temperature range 10-25 °C. Geldanamycin (GA) and its analogues 17-AAG [17-(allylamino)-17-demethoxy-GA] and 17-DMAG (17-N,N-dimethylaminoethylamino-17-demethoxy-GA) bind more strongly and have a dominant favourable enthalpic contribution over the temperature range investigated. We investigated the temperature dependence of the enthalpic contribution to binding. We found that while the ansamycin compounds have the commonly observed negative value, the nucleotides show a negligible or even a positive ΔC_p of binding. These data represent the first observation of a single binding site for which interactions with different ligands result in both negative and positive ΔC_p values. By addressing the likely impact of the potential contributions from protein-ligand interactions, we are able to attribute the anomalous ΔC_p for the nucleotides largely to a change in the conformation of the domain structure and local motion in the lid region of N-Hsp90 with the concomitant exposure of hydrophobic amino acid side chains.     

Effect of thiostrepton and 3′-terminal fragments of aminoacyl-tRNA on EF-Tu and ribosome-dependent GTP hydrolysis

The effect of the antibiotics thiostrepton and micrococcin on EF-T_u-catalyzed (ribosome-dependent) GTP hydrolysis in the presence of A-Phe, C-A-Phe, or C-C-A-Phe (related to the sequence of the 3′-terminus of aminoacyl-tRNA)(System I) or by methanol (‘uncoupled GTPase’, System II) was investigated. In System I, thiostrepton increases the binding affinities of the effectors to the EF-T_u·GTP·70 S ribosome complex, as well as the extent of the GTP hydrolysis, while the K^GTP_m is virtually unchanged. Similarly, in the uncoupled system (System II) and in the absence of effectors, thiostrepton significantly increases V^GTP_max, whereas K^GTP_m remains unaffected. Micrococcin is without any effect in both systems. The ‘uncoupled GTPase’ (in System II) is also strongly inhibited by C-A-Phe. The results indicate the crucial role of the EF-T_u site which binds the aminoacylated C-C-A terminus of aminoacyl-tRNA in promoting GTP hydrolysis. It follows that the binding of the model effectors (such as C-C-A-Phe) to that site is favorably influenced by the interaction of thiostrepton with the 50 S ribosomal subunit, whereas thiostrepton, per se, does not influence the affinity of EF-T_u for GTP.     

Survival of Escherichia coli in the environment: fundamental and public health aspects

In this review, our current understanding of the species Escherichia coli and its persistence in the open environment is examined. E. coli consists of six different subgroups, which are separable by genomic analyses. Strains within each subgroup occupy various ecological niches, and can be broadly characterized by either commensalistic or different pathogenic behaviour. In relevant cases, genomic islands can be pinpointed that underpin the behaviour. Thus, genomic islands of, on the one hand, broad environmental significance, and, on the other hand, virulence, are highlighted in the context of E. coli survival in its niches. A focus is further placed on experimental studies on the survival of the different types of E. coli in soil, manure and water. Overall, the data suggest that E. coli can persist, for varying periods of time, in such terrestrial and aquatic habitats. In particular, the considerable persistence of the pathogenic E. coli O157:H7 is of importance, as its acid tolerance may be expected to confer a fitness asset in the more acidic environments. In this context, the extent to which E. coli interacts with its human/animal host and the organism''s survivability in natural environments are compared. In addition, the effect of the diversity and community structure of the indigenous microbiota on the fate of invading E. coli populations in the open environment is discussed. Such a relationship is of importance to our knowledge of both public and environmental health.