5(6)-carboxyfluorescein revisited: new protecting group, separation of isomers, and their spectral properties on oligonucleotides

Pentafluorophenyl esters of 5- and 6-carboxyfluorescein-3',6'-O-dipivalate can be easily separated in multigram quantities by column chromatography. The individual isomers were converted into stable phosphoramidites suitable for oligonucleotide synthesis. The use of the cyclohexylcarbonyl (Chc) protecting group instead of pivaloyl (Piv) facilitates the separation of isomers. The fluorescence spectra of 5- and 6-carboxyfluoresceins on oligonucleotides were compared.     

Cajal bodies and interchromatin granule clusters in cricket oocytes: composition, dynamics and interactions

The organization and molecular composition of complicated Cajal bodies (CBs) and interchromatin granule clusters (IGCs) in oocytes of the house cricket, Acheta domesticus, were studied using immunofluorescent/confocal and Immunogold labeling/electron microscopy. In A. domesticus oocytes, the CB consists of the fibrillar matrix and a central cavity containing a predominantly granular body with insertions of tightly packed fibrillar material. The latter structure was identified as an "internal" IGC, since it is enriched with the SC35 protein, a marker for IGCs. The IGCs located outside the CB were also identified. Microinjections of the fluorescein-tagged U7 snRNA into the ooplasm showed the targeting of the U7 to the matrix of the CB. Some other essential CB components (coilin, snRNPs, fibrillarin) were found to be colocalized in the matrix of the CB. Neither confocal nor Immunogold microscopy revealed significant amounts of RNA polymerase II (pol II) in the CB of A. domesticus oocytes. The splicing factor SC35 was detected in the matrix of the CB. In oocytes treated with DRB, the amount of IGCs in the nucleoplasm increased significantly, granular and fibrillar components of IGCs were segregated, and the fibrillar areas accumulated pol II. Additionally, IG-like granules were shown to display on the surface of the CB probably due to a shifting from the internal IGC. We believe that in A. domesticus oocytes, CBs are involved in nuclear distribution of splicing factors, but their role in pol II transport is less significant. We also suggest that the formation of complicated CBs is a result of interconnection between two different nuclear domains, CBs and IGCs.     

Interaction between kringle and growth-factor-like domains in the urokinase molecule: Possible role in stimulation of chemotaxis

The results presented in this paper suggest the presence of an interaction between the kringle- and the growth-factor-like urokinase domains. This interaction regulates chemotactic properties of urokinase. We also show that interaction of urokinase with its "classical" receptor (uPAR) has a "permissive" effect on the interactions between the kringle domain and other targets on the cell surface. On the basis of our data we can suggest that uPAR serves as an "adaptor" for urokinase, and the binding of urokinase kringle domain to its receptor causes immediate activation of intracellular signaling and induction of cell migration.     

Conformational analysis of the carboxy-terminal tails of human beta-tubulin isotypes

Several isotypes of the structural protein tubulin have been characterized. Their expression offers a plausible explanation for differences regarding microtubule function. Although sequence variation between tubulin isotypes occurs throughout the entire protein, it is the extreme carboxy-terminal tails (CTTs) that exhibit the greatest concentration of differences. In humans, the CTTs range in length from 9 to 25 residues and because of a considerable number of glutamic acid residues, contain over 1/3 of tubulin's total electrostatic charge. The CTTs are believed to be highly disordered and their precise function has yet to be determined. However, their absence has been shown to result in altered microtubule stability and a reduction in the interaction with several microtubule-associated proteins (MAPs). To characterize the role that CTTs play in microtubule function, we examined the global conformational differences within a set of nine human β-tubulin isotypes using replica exchange molecular dynamics simulations. Through the analysis of the resulting configuration ensembles, we quantified differences such as the CTTs sequence influence on overall flexibility and average secondary structure. Although only minor variations between each CTT were observed, we suggest that these differences may be significant enough to affect interactions with MAPs, thereby influencing important properties such as microtubule assembly and stability.     

Application of organic acids for plant protection against phytopathogens

The basic tendency in the field of plant protection concerns with reducing the use of pesticides and their replacement by environmentally acceptable biological preparations. The most promising approach to plant protection is application of microbial metabolites. In the last years, bactericidal, fungicidal, and nematodocidal activities were revealed for citric, succinic, α-ketoglutaric, palmitoleic, and other organic acids. It was shown that application of carboxylic acids resulted in acceleration of plant development and the yield increase. Of special interest is the use of arachidonic acid in very low concentrations as an inductor (elicitor) of protective functions in plants. The bottleneck in practical applications of these simple, nontoxic, and moderately priced preparations is the absence of industrial production of the mentioned organic acids of required quality since even small contaminations of synthetic preparations decrease their quality and make them dangerous for ecology and toxic for plants, animals, and human. This review gives a general conception on the use of organic acids for plant protection against the most dangerous pathogens and pests, as well as focuses on microbiological processes for production of these microbial metabolites of high quality from available, inexpensive, and renewable substrates.

     

Prognostic Values of Some Cardiovascular Risk Factors in Cosmonauts

A ten-year follow-up study was focused on comparison of risk factors of atherosclerosis and damage to target organs in a group of 40- to 65-year-old cosmonauts and volunteers in the same age range. It was shown that the incidence of supraventricular extrasystoles in the groups increased with aging by 85 and 80%, respectively, inferring the risk of supraventricular tachyarrhythmias. The primary predictors of atherosclerosis and coronary artery disease (CAD) in cosmonauts are decreased endothelium-dependent vasodilation in the right brachial artery, C-reactive protein (hsCRP) level > 2 mg/L, and high atherogenic index. These CAD predictors should be of particular concern during medical screening of candidates for cosmonauts.     

Localization of the chromatin-remodeling protein ATRX in the oocyte nucleus of some insects

A comparative study of nuclear distribution of the chromatin-remodeling protein ATRX in the oocytes of three species of insects in which the oocyte nucleus at the diplotene stage differs in structure, has been carried out using fluorescent and immunoelectron microscopy. In tóhe oocyte nucleus of the tenebrionid beetles, Tribolium castaneum and Tenebrio molitor, ATRX preferably associates with the karyosphere (karyosome) that represents a result of concentration of the condensed chromosomes in a limited volume of the nucleus. In the oocyte nucleus of the house cricket, Acheta domesticus, in which a karyosphere does not form, the protein ATRX is distributed in the entire nuclear volume in association with the chromatin. The fact of ATRX presence in the extrachromosomal structures of the insect oocyte nucleus, the karyosphere capsule and specific nuclear bodies, is documented for the first time.     

Genotyping of patients with phenylketonuria from different regions of Russia for determining BH4 responsiveness

To date, the efficacy of the phenylalanine hydroxylase (PAH) cofactor is proved for the treatment of both BH4-dependent hyperphenylalaninemia and phenylketonuria patients with mutations in the PAH gene. Since the patient’s response depends on the presence of residual PAH enzyme activity, it is advisable to search for mutations in the PAH gene to identify the potential responders and nonresponders to therapy. Four hundred thirty-five phenylketonuria patients from 13 regions of the Russian Federation were genotyped in order to identify responders and nonresponders to tetrahydrobiopterin (BH4) therapy. According to the results of this study, the number of probable nonresponders to the BH4 treatment exceeds 50% owing to a higher overall allelic frequency of “severe” PAH gene mutations. Responder patients with two “mild” mutations in the PAH gene were identified (1.6%).