NMR Structure of the Amino-Terminal Domain of the Lambda Integrase Protein in Complex with DNA: Immobilization of a Flexible Tail Facilitates Beta-Sheet Recognition of the Major Groove

The integrase protein (Int) from bacteriophage lambda is the archetypal member of the tyrosine recombinase family, a large group of enzymes that rearrange DNA in all domains of life. Int catalyzes the insertion and excision of the viral genome into and out of the Escherichia coli chromosome. Recombination transpires within higher-order nucleoprotein complexes that form when its amino-terminal domain binds to arm-type DNA sequences that are located distal to the site of strand exchange. Arm-site binding by Int is essential for catalysis, as it promotes Int-mediated bridge structures that stabilize the recombination machinery. We have elucidated how Int is able to sequence specifically recognize the arm-type site sequence by determining the solution structure of its amino-terminal domain (Int^N, residues Met1 to Leu64) in complex with its P′2 DNA binding site. Previous studies have shown that Int^N adopts a rare monomeric DNA binding fold that consists of a three-stranded antiparallel beta-sheet that is packed against a carboxy-terminal alpha helix. A low-resolution crystal structure of the full-length protein also revealed that the sheet is inserted into the major groove of the arm-type site. The solution structure presented here reveals how Int^N specifically recognizes the arm-type site sequence. A novel feature of the new solution structure is the use of an 11-residue tail that is located at the amino terminus. DNA binding induces the folding of a 3₁₀ helix in the tail that projects the amino terminus of the protein deep into the minor groove for stabilizing DNA contacts. This finding reveals the structural basis for the observation that the “unstructured” amino terminus is required for recombination.     

Multikinase inhibitor Sorafenib and HDAC inhibitor suberoylanilide hydroxamic acid suppress confluent resistance of cancer cells to recombinant protein izTRAIL

Suppression of human tumor cell resistance to TRAIL-induced apoptosis in confluent cultures, using molecular target drugs (sorafenib and SAHA) at non-toxic concentrations was studied. Sorafenib, a multikinase inhibitor, and SAHA, an inhibitor of histone deacetylase, effectively suppressed resistance of confluent human cells derived from the skin carcinoma (A431 cell line) and fibrosarcoma (HT-1080 cell line). The effectiveness of suppression of confluent resistance with these inhibitors for human carcinoma A431 cells was significantly higher than that for the human ovarian carcinoma OVCAR-3 cells. For all cell lines studied, suppression of confluent resistance with SAHA was more effective than when sorafenib was used. The possible reason for increasing tumor cell resistance in confluent cultures and the importance of this phenomenon for understanding drug resistance of cells in the tumor tissue are discussed.     

A systems approach to morphogenesis in Arabidopsis thaliana: II. Modeling the regulation of shoot apical meristem structure

Two models of the mechanism maintaining a zonal structure in the shoot apical meristem (SAM) were built based on the analysis of experimental data on the interactions between CLV1, CLV2, CLV3, and WUS genes and the concepts of their role in this mechanism. The first model, a simple one-dimensional model with two morphogens, which is a variant of Wolpert’s French flag model [1], describes the regulation of zone distribution along the SAM vertical axis. Despite a number of simplifications, this model has stationary solutions with biologically meaningful interpretation. The simplifying assumptions were successively abandoned in constructing a two-dimensional model of the mechanism underlying the regulation of SAM structure. This model provides a better understanding of the distributed system that regulates the SAM structure, and allows more detailed formalization of the modern concepts and experimental data concerning this mechanism.

     

Regulatory Contexts in the 5'-Region of mRNA from Arabidopsis thaliana Plants and Their Role in Translation Efficiency

Russian Journal of Plant Physiology - In this study, the polysome profiling method was used for the separation of mRNA depending on their loading by ribosomes into polysomal and monosomal...     

Pigment Pattern Formation in the Guppy,Poecilia reticulata,Involves the Kita and Csf1ra Receptor Tyrosine Kinases

Males of the guppy (Poecilia reticulata) vary tremendously in their ornamental patterns, which are thought to have evolved in response to a complex interplay between natural and sexual selection. Although the selection pressures acting on the color patterns of the guppy have been extensively studied, little is known about the genes that control their ontogeny. Over 50 years ago, two autosomal color loci, blue and golden, were described, both of which play a decisive role in the formation of the guppy color pattern. Orange pigmentation is absent in the skin of guppies with a lesion in blue, suggesting a defect in xanthophore development. In golden mutants, the development of the melanophore pattern during embryogenesis and after birth is affected. Here, we show that blue and golden correspond to guppy orthologs of colony-stimulating factor 1 receptor a (csf1ra; previously called fms) and kita. Most excitingly, we found that both genes are required for the development of the black ornaments of guppy males, which in the case of csf1ra might be mediated by xanthophore–melanophore interactions. Furthermore, we provide evidence that two temporally and genetically distinct melanophore populations contribute to the adult camouflage pattern expressed in both sexes: one early appearing and kita-dependent and the other late-developing and kita-independent. The identification of csf1ra and kita mutants provides the first molecular insights into pigment pattern formation in this important model species for ecological and evolutionary genetics.     

Mitochondrial phylogeny of Locustella and related genera

We used maximum likelihood analysis of complete mitochondrial ND2 sequences (1041 bp) to clarify the taxonomy and relationships of various species and genera of grass and bush warblers. The tree revealed two clades of grass and bush warblers. One clade was comprised of all four western Palearctic Locustella and two species of Asian Bradypterus . The other clade included five eastern Palearctic Locustella (including the distinctive Sakhalin warbler Locustella amnicola ) and the marsh grassbird Megalurus pryeri . African Bradypterus and Australian little grassbird Megalurus gramineus were distantly related to their Asian congeners. Therefore, current taxonomy of these genera does not reflect their evolutionary history and needs revision. It is proposed that a phylogenetic analysis of morphology and ecological preferences would show that the current taxonomy of grass and bush warblers reflects species' habitat preferences and morphology related to locomotion and foraging in their habitats, rather than their shared ancestry. Distinct clades were found in grasshopper warbler Locustella naevia and Pallas's grasshopper warbler L. certhiola . Detailed phylogeographic studies are needed to elucidate the species status of the clades within these two species.     

Investigation of OCP-triggered dissipation of excitation energy in PSI/PSII-less Synechocystis sp. PCC 6803 mutant using non-linear laser fluorimetry

In order to prevent photodestruction by high light, photosynthetic organisms have evolved a number of mechanisms, known as non-photochemical quenching (NPQ), that deactivate the excited states of light harvesting pigments. Here we investigate the NPQ mechanism in the cyanobacterium Synechocystis sp. PCC 6803 mutant deficient in both photosystems. Using non-linear laser fluorimetry, we have determined molecular photophysical characteristics of phycocyanin and spectrally distinct forms of allophycocyanin for the cells in non-quenched and quenched states. Our analysis of non-linear fluorescence characteristics revealed that NPQ activation leads to an ~ 2-fold decrease in the relaxation times of both allophycocyanin fluorescence components, F660 and F680, and a 5-fold decrease in the effective excitation cross-section of F680, suggesting an emergence of a pathway of energy dissipation for both types of allophycocyanin. In contrast, NPQ does not affect the rates of singlet–singlet exciton annihilation. This indicates that, upon NPQ activation, the excess excitation energy is transferred from allophycocyanins to quencher molecules (presumably 3′hydroxyechinenone in the orange carotenoid protein), rather than being dissipated due to conformational changes of chromophores within the phycobilisome core. Kinetic measurements of fluorescence quenching in the Synechocystis mutant revealed the presence of several stages in NPQ development, as previously observed in the wild type. However, the lack of photosystems in the mutant enhanced the magnitude of NPQ as compared to the wild type, and allowed us to better characterize this process. Our results suggest a more complex kinetics of the NPQ process, thus clarifying a multistep model for the formation of the quenching center.     

Mitochondrial DNA and plumage evolution in the white wagtail Motacilla alba

We analyzed sequences of two mitochondrial DNA (mtDNA) gene regions (control region and ND2) totaling 1477 base‐pairs from 232 specimens of the white wagtail Motacilla alba obtained from 27 localities throughout Eurasia. Although overall haplotype diversity was relatively low (0.79) and the most common haplotype was shared by 45% of individuals, belonging to six subspecies, a high level of population differentiation was detected. The mtDNA tree revealed three clades: (1) most individuals from Krasnodar (belonging to M. a. alba subspecies), (2) all individuals from Almaty and some from Primor'e (belonging to M. a. personata, M. a. lugens and M. a. leucopsis subspecies), and (3) the remaining individuals (representing all subspecies and all localities except Almaty). We suggest that these three clades represent historically isolated populations that relatively recently came into secondary contact in Krasnodar and Primor'e. None of the six subspecies were reciprocally monophyletic in the mtDNA tree. The Krasnodar population appeared to receive immigrants from other localities, but distinctive haplotypes from this locality did not appear elsewhere, suggesting asymmetric gene flow. Signatures of recent gene flow between northern populations were detected, and there was no evidence of isolation by distance within the northern group of populations. Mismatch distributions for most localities were consistent with population expansions. We also analyzed 12 male plumage characters from 93 study skins sampled from 24 populations. Phylogenetic trees resulting from separate genetic and morphological analyses were incongruent. Plumage evolution seems to be under strong sexual or natural selection, which favors particular phenotypes in various areas irrespective of the mitochondrial background. Dispersal events at different evolutionary times could have obscured the effects of earlier isolation events. The mtDNA data does not support species status for M. a. lugens and M. a. personata, which shared haplotypes with other subspecies of M. alba. We recommend that M. lugens and M. personata are placed as junior synonyms of M. alba.     

Circadian rhythms of antibacterial resistance, coagulase and antilysozyme activity of Staphylococcus aureus

AIM: To determine daily dynamics of antibacterial resistance as well as antilysozyme and coagulase activity of S. aureus strains. MATERIALS AND METHODS: On an example of clinical strains of S.aureus isolated from patients with surgical infections daily dynamics of biological characteristics of staphylococci was studied. After 12 hours of incubation strains were tested for coagulase activity by standard method (test tube method), antilysozyme activity by photometric method, and antibacterial resistance by method of serial dilutions in agar. Tests were repeated each 3-hours during a day. RESULTS: Variation of levels of studied biological characteristics of staphylococci during a day was revealed. Structures of coagulase and antilysozyme circadian rhythms had some differences in different S. aureus strains. Alongside with it, similarity in temporal expression of such biological characteristics of staphylococci as antibacterial resistance and antilysozyme activity was noted. CONCLUSION: Obtained data open prospect to use biorhythmological approach in study of biological characteristics of microorganisms during evaluation of their mechanisms of adaptation to changing environmental conditions. Chronobiological approach allows to reveal periods of maximal expression of S. aureus characteristics that could be used for increasing of effectiveness of antibacterial treatment by the choice of optimal time for administration of antibiotic.     

Lymphoprostatic scintigraphy in regional lymphatic mapping in patients with prostate cancer

The purpose of the investigation was to upgrade the diagnosis quality for metastatic lesion to the regional lymp nodes in prostate cancer. The results of the most widely used diagnostic techniques (computed tomography (CT), magnetic resonance Imaging, (MRI), radionuclide studies (RNS), and ultrasonography (USG)), which had been compared with those of histology of removed lymph nodes, were assessed. For this, 74 patients with prostate cancer were comprehensively examined and treated at the Central Research X-ray Radiolological Institute. The comparative assessment of radiodiagnostic techniques (USG, CT, MRI, and RNS) showed the high informative value of retroperitoneal lymph nodal MRI in the detection of structural-and-morphological and anatomic-and-topographic changes in the lymph nodes (87.5% sensitivity, 83.3% specificity, and 85.7% accuracy). The developed lymphoprostatic scintigraphy (LPSG) is an informative technique that allows the better diagnosis of prostate metastases to the regional lymph nodes (92.4% sensitivity, 91.7% specificity, and 92.1% accuracy; the prognostic value of a positive result is 90.7%). LPSG is superior in its informative value to small pelvic USG and CT and inferior to indirect lymphoscintigraphy and yields valuable additional information on the physiological function of the small pelvic lymphatic collector.