Synthesis and biological activity of oxo-7H-benzo[e]perimidine-4-carboxylic acid derivatives as potent, nonpeptide corticotropin releasing factor (CRF) receptor antagonists

A novel series of derivatives of oxo-7H-benzo[e]perimidine-4-carboxylic acid (I) potently displaced radioligand binding of 125I-CRF to both CRF1 and CRF2 receptors. The members of this series antagonized CRF-stimulated cAMP formation and CRF-stimulated corticotropin release from rat pituitary in vivo. These are the first nonpeptide antagonists to show activity at both CRF1 and CRF2 receptors.     

NKG2D splice variants: a reexamination of adaptor molecule associations

NKG2D is a homodimeric C-type lectin-related receptor expressed on natural killer (NK) cells and T cells. In mice, alternative deoxyribonucleic acid (DNA) splicing generates two isoforms of NKG2D that differ in the length of their cytoplasmic domains. Their ability to induce cellular activation is mediated via association with two membrane-bound, signaling adaptor molecules, DAP10 and DAP12. It has been reported that the long form of NKG2D associates exclusively with DAP10, whereas the short variant can interact with either adaptor. The short isoform was reported to be almost undetectable in naïve NK cells. Using two distinct cell types, we demonstrate that like the short isoform, the long variant of NKG2D also associates not only with DAP10 but also with DAP12. Using reporter cells (70Z/3), we demonstrate that DAP12 can compete equally with DAP10 for association with both variants of NKG2D when DAP10 and DAP12 are coexpressed. Cross-linking either isoform of NKG2D induces a calcium flux when associated exclusively with DAP10 or DAP12. Moreover, using quantitative polymerase chain reaction (PCR), we also show that the short isoform of NKG2D is expressed in naïve NK cells. Our data suggest that signaling via mouse NKG2D isoforms is more complex than originally presented.     

Properties of neutral cellobiose dehydrogenase from the ascomycete Chaetomium sp. INBI 2-26(-) and comparison with basidiomycetous cellobiose dehydrogenases

The extracellular cellobiose dehydrogenase (CDH) obtained from Chaetomium sp. INBI 2-26(-) has a molecular mass of 95 kDa and an isoelectric point of 5. This novel CDH is highly specific for the oxidation of cellobiose (K(m,app) 4.5 microM) and lactose (K(m,app) 56 microM). With 2,6-dichloroindophenol (DCIP) and cytochrome c(3+) (cyt c(3+)) as electron acceptors, CDH was most active at pH 6. The turnover number of the enzyme for cellobiose, lactose, DCIP and cyt c(3+) was in the range of 9-14s(-1) at 20 degrees C and pH 6. The UV-visible spectrum revealed the flavohemoprotein nature of the enzyme. The cytochrome b domain of the enzyme was reduced by ascorbate, dithionite, as well as specifically by cellobiose in a wide range of pH. The apparent first order rate constants of the spontaneous re-oxidation of the reduced heme domain were estimated as 0.01 and 0.00039 s(-1) at pH 4.5 and 6.5, respectively. The half-inactivation time of CDH at pH 6 and 55 degrees C was ca. 100 min; the stability at pH 8 and, particularly, pH 4 was remarkably lower. Cellobiose stabilized the enzyme against thermal inactivation, whereas DCIP in turn sensitized the enzyme. The new enzyme revealed low affinity for crystalline cellulose, but was capable of binding onto H(3)PO(4)-swollen filter paper. The results show significant differences to already known CDHs and perspectives for several biotechnological applications, where CDH with maximal activity at neutral pH and high affinity for cellobiose and lactose night have some advantages.     

Nucleotide Sequences of Plasmodium vivaxA-Type rDNA ITS1, 5.8S,and ITS2. Elaboration of Retrospective PCR Diagnostics of Malaria Using Stained Thick Blood Smears

Stages life cycle of the malaria parasite differ in the rate of replication and the structural properties of functionally active A-, S-, and O-type ribosomes. Regions of A-type rDNA including ITS1, 5.8S, and ITS2 from two strains of Plasmodium vivaxwith different incubation periods were amplified and sequenced. No substantial differences in the sequences of two strains were revealed. Phylogenetic analysis of the obtained and homologous sequences of ITS1 rDNA of A, S, and O types of P. vivax; A and S types of P. falciparum; and Cryptosporidium parvum, Eimeria maxima, Toxoplasma gondiias outgroup, by the maximum parsimony method using PAUP 4.0 revealed that divergence of ITS1 might have occurred after speciation and at different rates in individual lineages of the Plasmodiumgenus. Basing on the results of the analysis of orthologous sequences of P. vivaxand P. falciparum, we developed genus- and species-specific primers for PCR diagnostics of malaria, as well as a one-step effective method of DNA isolation from Giemsa–Romanovsky-stained thick blood smears. It was demonstrated that stained preparations could be a reliable source of plasmodial DNA, and the quality of preparations and storage time (10–20 years) did not interfere with the results of PCR analysis.     

Galectins: an evolutionarily conserved family of animal lectins with multifunctional properties; a trip from the gene to clinical therapy.

Galectins constitute a family of evolutionarily conserved animal lectins, which are defined by their affinity for poly-N-acetyllactosamine-enriched glycoconjugates and sequence similarities in the carbohydrate recognition domain. During the past decade, attempts to dissect the functional role for galectins in vivo have been unsuccessful in comparison to the overwhelming information reached at the biochemical and molecular levels. The present review deals with the latest advances in galectin research and is aimed at validating the functional significance of these carbohydrate-binding proteins. Novel implications of galectins in cell adhesion, cell growth regulation, immunomodulation, apoptosis, inflammation, embryogenesis, metastasis and pre-mRNA splicing will be particularly discussed in a trip from the gene to the clinical therapy. Elucidation of the molecular mechanisms involved in galectin functions will certainly open new avenues not only in biomedical research, but also at the level of disease diagnosis and clinical intervention, attempting to delineate new therapeutic strategies in autoimmune diseases, inflammatory processes, allergic reactions and tumor spreading.     

Immunoenzyme analysis of angiogenin in cow's milk]

Competitive enzyme immunoassay based on polyclonal antibodies can be used for determining the content of angiogenin in milk. These polyclonal antibodies had no cross-reactions with ribonuclease or other milk whey proteins. Milk angiogenin levels in samples taken fvom animals of a separate population varied from 2.09 to 4.85 mg/l. Unlike cow milk productivity, the number of calvings affects the milk angiogenin content.     

The role of auxin and sugar signaling in dominance inhibition of inflorescence growth by fruit load

Dominance inhibition of shoot growth by fruit load is a major factor that regulates shoot architecture and limits yield in agriculture and horticulture crops. In annual plants, the inhibition of inflorescence growth by fruit load occurs at a late stage of inflorescence development termed the end of flowering transition. Physiological studies show this transition is mediated by production and export of auxin from developing fruits in close proximity to the inflorescence apex. In the meristem, cessation of inflorescence growth is controlled in part by the age-dependent pathway, which regulates the timing of arrest. Here, we show the end of flowering transition is a two-step process in Arabidopsis (Arabidopsis thaliana). The first stage is characterized by a cessation of inflorescence growth, while immature fruit continues to develop. At this stage, dominance inhibition of inflorescence growth by fruit load is associated with a selective dampening of auxin transport in the apical region of the stem. Subsequently, an increase in auxin response in the vascular tissues of the apical stem where developing fruits are attached marks the second stage for the end of flowering transition. Similar to the vegetative and floral transition, the end of flowering transition is associated with a change in sugar signaling and metabolism in the inflorescence apex. Taken together, our results suggest that during the end of flowering transition, dominance inhibition of inflorescence shoot growth by fruit load is mediated by auxin and sugar signaling.

Dominance inhibition of inflorescence shoot growth by fruit load involves auxin and sugar signaling during the end of flowering transition.     

The potential distribution of German wasps (Vespula germanica) in Argentina

Abstract

In the early 1980s, the German wasp (Vespula germanica) arrived in Argentina and became established in northern Patagonia. We use an ecoclimatic computer‐based program (CLIMEX) to predict the potential spread of wasps to other areas of the country. The model suggests that much of Argentina does not appear to be very suitable for the species’ establishment. However, more suitable areas than those currently colonised by the wasp occur in the centre of the country. Based on CLIMEX results, we suggest that, if the wasps ever break through the barrier of unfavourable habitat that surrounds their current distribution, they will continue dispersing to those areas. Notwithstanding this, factors other than climate may explain why German wasps are firmly established in areas otherwise considered to be inhospitable for the species.