Antitumor reactivity of non-metallocene titanium complexes of oxygen-based ligands: is ligand lability essential?

In our attempt to define the parameters affecting anticancer activity of titanium complexes and to assess the role of hydrolytic stability, titanium compounds of oxygen-based ligands were studied. A homoleptic complex of hydroxyamino-1,3,5-triazine ligands was prepared and its hydrolysis was investigated by UV-vis spectroscopy at biologically relevant pH and temperature conditions based on its ligand to metal charge transfer absorption band. This complex exhibits very high hydrolytic stability under the conditions measured with negligible ligand dissociation. Its anticancer reactivity was investigated on ovarian OVCAR-1 and colon HT-29 cells, in comparison with the reference highly labile Ti(OiPr)(4) and TiCl(4)(THF)(2) (where THF is tetrahydrofuran), the inert thermodynamically stable TiO2, and the free aromatic hydroxyamino-1,3,5-triazine ligand. Whereas all reference titanium complexes were found to be completely unreactive against both tumor cell types, suggesting some moderate inertness is required, the homoleptic complex of the triazine ligands clearly possess some mild reactivity despite having no labile groups, and despite its incomplete solubility in the concentrations applied. As the free aromatic ligand is highly active under similar conditions, detailed time-dependence measurements were conducted and indicated that the cytotoxicity of the ligand is more affected by reducing incubation time, and that introducing the titanium complex to the medium prior to cell administration does not increase reactivity at a certain incubation time. These findings suggest that the reactivity of the complex does not result from that of the free ligand following dissociation, but rather involves the titanium center.     

Structural basis of diversity and homodimerization specificity of zinc-finger-associated domains in Drosophila

In arthropods, zinc finger-associated domains (ZADs) are found at the N-termini of many DNA-binding proteins with tandem arrays of Cys2-His2 zinc fingers (ZAD-C2H2 proteins). ZAD-C2H2 proteins undergo fast evolutionary lineage-specific expansion and functional diversification. Here, we show that all ZADs from Drosophila melanogaster form homodimers, but only certain ZADs with high homology can also heterodimerize. CG2712, for example, is unable to heterodimerize with its paralog, the previously characterized insulator protein Zw5, with which it shares 46% homology. We obtained a crystal structure of CG2712 protein''s ZAD domain that, in spite of a low sequence homology, has similar spatial organization with the only known ZAD structure (from Grauzone protein). Steric clashes prevented the formation of heterodimers between Grauzone and CG2712 ZADs. Using detailed structural analysis, site-directed mutagenesis, and molecular dynamics simulations, we demonstrated that rapid evolutionary acquisition of interaction specificity was mediated by the more energy-favorable formation of homodimers in comparison to heterodimers, and that this specificity was achieved by multiple amino acid substitutions resulting in the formation or breaking of stabilizing interactions. We speculate that specific homodimerization of ZAD-C2H2 proteins is important for their architectural role in genome organization.     

The Effect of TIGAR Knockdown on Apoptotic and Epithelial‐Mesenchymal Markers Expression in Doxorubicin‐Resistant Non‐Small Cell Lung Cancer A549 Cell Lines

Resistance to chemotherapeutic drugs is a critical problem in cancer therapy, but the underlying mechanism has not been fully elucidated. TP53‐induced glycolysis regulatory phosphatase (TIGAR), an important glycolysis and apoptosis regulator, plays a crucial role in cancer cell survival by protecting cells against oxidative stress‐induced apoptosis. In the present study, we investigated whether TIGAR is involved in epithelial‐mesenchymal transition (EMT) in doxorubicin (DOX)‐resistant human non‐small cell lung cancer (NSCLC), A549/DOX cells. We found that the expression of TIGAR was significantly higher in A549/DOX cells than in the parent A549 cell lines. siRNA‐mediated TIGAR knockdown reduced migration, viability and colony survival of doxorubicin‐resistant lung cancer cells. Also, TIGAR knockdown decreased pro‐survival protein Bcl‐2 and increased pro‐apoptotic Bax and cleaved poly (ADP‐ribose) polymerase (PARP). Moreover, TIGAR depletion significantly up‐regulated both caspase‐3 and caspase‐9 expression. Furthermore, TIGAR depletion up‐regulated the expression of E‐cadherin and down‐regulated the expression of vimentin. These results indicate that TIGAR knockdown may inhibit EMT in doxorubicin (DOX)‐resistant human NSCLC and may represent a therapeutic target for a non‐small lung cancer cells chemoresistance.     

Re-engineering an alphoidtetO-HAC-based vector to enable high-throughput analyses of gene function

Human artificial chromosome (HAC)-based vectors represent an alternative technology for gene delivery and expression with a potential to overcome the problems caused by the use of viral-based vectors. The recently developed alphoid^tetO-HAC has an advantage over other HAC vectors because it can be easily eliminated from cells by inactivation of the HAC kinetochore via binding of tTS chromatin modifiers to its centromeric tetO sequences. This provides unique control for phenotypes induced by genes loaded into the alphoid^tetO-HAC. However, inactivation of the HAC kinetochore requires transfection of cells by a retrovirus vector, a step that is potentially mutagenic. Here, we describe an approach to re-engineering the alphoid^tetO-HAC that allows verification of phenotypic changes attributed to expression of genes from the HAC without a transfection step. In the new HAC vector, a tTS-EYFP cassette is inserted into a gene-loading site along with a gene of interest. Expression of the tTS generates a self-regulating fluctuating heterochromatin on the alphoid^tetO-HAC that induces fast silencing of the genes on the HAC without significant effects on HAC segregation. This silencing of the HAC-encoded genes can be readily recovered by adding doxycycline. The newly modified alphoid^tetO-HAC-based system has multiple applications in gene function studies.     

Exceptional error minimization in putative primordial genetic codes

Background  

The standard genetic code is redundant and has a highly non-random structure. Codons for the same amino acids typically differ only by the nucleotide in the third position, whereas similar amino acids are encoded, mostly, by codon series that differ by a single base substitution in the third or the first position. As a result, the code is highly albeit not optimally robust to errors of translation, a property that has been interpreted either as a product of selection directed at the minimization of errors or as a non-adaptive by-product of evolution of the code driven by other forces.     

New Morphofunctional Criteria for Resistance Profile in Post-Traumatic Stress Disorder Models as Adrenal Dysfunction Trigger

Doklady Biological Sciences - For the first time in modeling posttraumatic stress disorder (PTSD), we have described the morphofunctional state of adrenal glands in Wistar rats resistant and...     

Structural changes of erythrocyte membrane revealed by 3D confocal optical profilometer

We examined hematological changes influenced by the experimental hypervitaminosis A. The 3D confocal optical profilometer was applied for assessment of the erythrocytes' membrane structural changes influenced by an overdose of vitamin A. The blood smears were evaluated in terms of alterations of geometrical and optical parameters of erythrocytes for two groups of animals: oil base and retinol palmitate (n = 9 animals for each group). The results demonstrate that an overdose of retinol palmitate causes changes in the torus curvature and pallor of discocytes, their surface area and volume. The observed structural malformations of the shape of red blood cells become visible at the earlier preclinical stage of changes in animal state and behavior. With this in mind, the results of the study open a new area of research in the certain dysfunction diagnosis of red blood cells and have a great potential in the further development of new curative protocols.