Conjugation with polyamines enhances the antibacterial and anticancer activity of chloramphenicol

Chloramphenicol (CAM) is a broad-spectrum antibiotic, limited to occasional only use in developed countries because of its potential toxicity. To explore the influence of polyamines on the uptake and activity of CAM into cells, a series of polyamine–CAM conjugates were synthesized. Both polyamine architecture and the position of CAM-scaffold substitution were crucial in augmenting the antibacterial and anticancer potency of the synthesized conjugates. Compounds 4 and 5, prepared by replacement of dichloro-acetyl group of CAM with succinic acid attached to N4 and N1 positions of N⁸,N⁸-dibenzylspermidine, respectively, exhibited higher activity than CAM in inhibiting the puromycin reaction in a bacterial cell-free system. Kinetic and footprinting analysis revealed that whereas the CAM-scaffold preserved its role in competing with the binding of aminoacyl-tRNA 3′-terminus to ribosomal A-site, the polyamine-tail could interfere with the rotatory motion of aminoacyl-tRNA 3′-terminus toward the P-site. Compared to CAM, compounds 4 and 5 exhibited comparable or improved antibacterial activity, particularly against CAM-resistant strains. Compound 4 also possessed enhanced toxicity against human cancer cells, and lower toxicity against healthy human cells. Thus, the designed conjugates proved to be suitable tools in investigating the ribosomal catalytic center plasticity and some of them exhibited greater efficacy than CAM itself.     

Reticulon 4 Is Necessary for Endoplasmic Reticulum Tubulation,STIM1-Orai1 Coupling,and Store-operated Calcium Entry

Despite recent advances in understanding store-operated calcium entry (SOCE) regulation, the fundamental question of how ER morphology affects this process remains unanswered. Here we show that the loss of RTN4, is sufficient to alter ER morphology and severely compromise SOCE. Mechanistically, we show this to be the result of defective STIM1-Orai1 coupling because of loss of ER tubulation and redistribution of STIM1 to ER sheets. As a functional consequence, RTN4-depleted cells fail to sustain elevated cytoplasmic Ca²⁺ levels via SOCE and therefor are less susceptible to Ca²⁺ overload induced apoptosis. Thus, for the first time, our results show a direct correlation between ER morphology and SOCE and highlight the importance of RTN4 in cellular Ca²⁺ homeostasis.     

Investigation of rifampicin resistance mechanisms in Brucella abortus using MS-driven comparative proteomics

Mutations in the rpoB gene have already been shown to contribute to rifampicin resistance in many bacterial strains including Brucella species. Resistance against this antibiotic easily occurs and resistant strains have already been detected in human samples. We here present the first research project that combines proteomic, genomic, and microbiological analysis to investigate rifampicin resistance in an in vitro developed rifampicin resistant strain of Brucella abortus 2308. In silico analysis of the rpoB gene was performed and several antibiotics used in the therapy of Brucellosis were used for cross resistance testing. The proteomic profiles were examined and compared using MS-driven comparative proteomics. The resistant strain contained an already described mutation in the rpoB gene, V154F. A correlation between rifampicin resistance and reduced susceptibility on trimethoprim/sulfamethoxazole was detected by E-test and supported by the proteomics results. Using 12?836 MS/MS spectra we identified 6753 peptides corresponding to 456 proteins. The resistant strain presented 39 differentially regulated proteins most of which are involved in various metabolic pathways. Results from our research suggest that rifampicin resistance in Brucella mostly involves mutations in the rpoB gene, excitation of several metabolic processes, and perhaps the use of the already existing secretion mechanisms at a more efficient level.     

Escaping the mouse trap: the selection of new Evo-Devo model species

Among the many, sometimes contradictory, criteria that have been used for promoting model species, the most prominent has probably been their relevance for understanding human biology. Recently however, the debate has partly shifted from the search for evolutionary conservation (medicine-driven models) to a better understanding of the generative mechanisms underlying biological diversity (Evo-Devo-driven models). Integration of multiple disciplines, beyond developmental genetics and evolutionary molecular genetics, as well as of innovative technologies will help biologists to open the massive realm of living species to genome manipulation and phenotypic investigation. However, a consensual list of model species must still be reached for optimizing the interplay between in silico analyses and in vivo experiments, and we claim that the Evo-Devo community should play a more energetic role in this endeavor. We discuss here a few criteria and limitations of major relevance to the choice of model species for Evo-Devo studies, and promote the use of a pragmatic approach. Finally, given the difficulties related to manipulating and breeding model species, we suggest the development of Evo-Devo virtual zoos maintaining breeding colonies of a selected set of species and from which eggs or staged embryos are available on order.     

The Control Region of Maternally and Paternally Inherited Mitochondrial Genomes of Three Species of the Sea Mussel Genus Mytilus

Species of the mussel genus Mytilus possess maternally and paternally transmitted mitochondrial genomes. In the interbreeding taxa Mytilus edulis and M. galloprovincialis, several genomes of both types have been fully sequenced. The genome consists of the coding part (which, in addition to protein and RNA genes, contains several small noncoding sequences) and the main control region (CR), which in turn consists of three distinct parts: the first variable (VD1), the conserved (CD), and the second variable (VD2) domain. The maternal and paternal genomes are very similar in gene content and organization, even though they differ by >20% in primary sequence. They differ even more at VD1 and VD2, yet they are remarkably similar at CD. The complete sequence of a genome from the closely related species M. trossulus was previously reported and found to consist of a maternal-like coding part and a paternal-like and a maternal-like CR. From this and from the fact that it was extracted from a male individual, it was inferred that this is a genome that switched from maternal to paternal transmission. Here we provide clear evidence that this genome is the maternal genome of M. trossulus. We have found that in this genome the tRNA^Gln in the coding region is apparently defective and that an intact copy of this tRNA occurs in the CR, that one of the two conserved domains is missing essential motifs, and that one of the two first variable domains has a high rate of divergence. These features may explain the large size and mosaic structure of the CR of the maternal genome of M. trossulus. We have also obtained CR sequences of the maternal and paternal genomes of M. californianus, a more distantly related species. We compare the control regions from all three species, focusing on the divergence among genomes of different species origin and among genomes of different transmission routes.     

Hydrolysis of 2-arachidonoylglycerol in Tetrahymena thermophila. Identification and partial characterization of a Monoacylglycerol Lipase-like enzyme

Tetrahymena thermophila is a model organism for molecular and cellular biology. Previous studies from our group showed that Tetrahymena contains major components of the endocannabinoid system, such as various endocannabinoids and FAAH. In mammalian cells the endocannabinoid 2-arachidonoylglycerol is inactivated mainly by MAGL. In this study we showed that 2-arachidonoylglycerol and 2-oleoylglycerol are hydrolyzed by the combined actions of MAGL and FAAH. MAGL-like activity was examined in the presence of FAAH specific inhibitors, URB597 or AM374 and showed optimum pH of 8-9, apparent K(M) of 14.1μM and V(max) of 5.8nmol/min×mg. The enzyme was present in membrane bound and cytosolic isoforms; molecular mass was determined at ～45 and ～40kDa. MAGL and FAAH could also inactivate endogenous signaling lipids, which might play an important role in Tetrahymena as suggested in mammals. Tetrahymena could be used as a model system for testing drugs targeting enzymes of the endocannabinoid system.     

The effects of a twenty-four-week aquatic training program on muscular strength performance in healthy elderly women

The purpose of the present study was to determine the effectiveness of a 24-week aquatic training (AT) program, which included both aerobic and resistance components, on muscle strength (isometric and dynamic), flexibility, and functional mobility in healthy women over 60 years of age. Twenty-two subjects were assigned randomly to either an AT (n = 12) or a control (C, n = 10) group. Volunteers participated in a supervised shallow-water exercise program for 60 minutes a day, 3 days a week; the exercise program consisted of a 10-minute warm-up and stretching, 25 minutes of endurance-type exercise (dancing) at 80% of heart rate (HR)(max), 20 minutes of upper- and lower-body resistance exercises with specialized water-resistance equipment, and a 5-minute cool down. Maximal isometric torque of knee extensors (KEXT) and knee flexors (KFLEX) were evaluated by a Cybex Norm dynamometer, grip strength (HGR) was evaluated using a Jamar hydraulic dynamometer, and dynamic strength was evaluated via the 3 repetition maximum (3RM) test for chest press, knee extension, lat pull down, and leg press. Jumping performance was evaluated using the squat jump (SJ), functional mobility with the timed up-and-go (TUG) test, and trunk flexion with the sit-and-reach test. Body composition was measured using the bioelectrical impedance method. The AT induced significant improvements in KEXT (10.5%) and KFLEX (13.4%) peak torque, HGR strength (13%), 3RM (25.7-29.4%), SJ (24.6%), sit-and-reach (11.6%), and TUG (19.8%) performance. The AT group demonstrated a significant increase in lean body mass (3.4%). No significant changes in these variables were observed in the C group. The results indicate that AT, with both aerobic and resistance components, is an alternative training method for improving neuromuscular and functional fitness performance in healthy elderly women.     

Synthesis of pyrimidinopyridine-triazene conjugates targeted to abl tyrosine kinase

The synthesis and abl tyrosine kinase inhibitory activities of alkyltriazenes conjugated to phenylaminopyrimidines are described. Significant abl inhibitory activities were observed only when a benzamido spacer was inserted between the 1,2,3-triazene chain and the 2-phenyaminopyridopyrimidine moiety.     

Endogenous Interleukin 6 Conveys Resistance to cis-Diamminedichloroplatinum-Mediated Apoptosis of the K562 Human Leukemic Cell Line

Cisplatin is an effective chemotherapeutic agent that elicits its antineoplastic activity by binding to DNA and disrupting template functions. IL-6 is a cytokine which has been shown to play a central role in host immunological defense mechanisms. Although K562 leukemic cells have been shown to secrete IL-6, little is known of whether there exists a correlation between the expression of IL-6 and the resistance of these cells to anticancer chemotherapeutic agents. To determine the contribution of IL-6 to the regulation of cisplatin-induced apoptosis in K562 cells, we examined whether treatment of K562 cells and cisplatin-resistant K562 subclones with anti-IL-6 mAb enhances their sensitivity to cisplatin. The results show that cis-diamminedichloroplatinum (CDDP) resistance was overcome by treatment with nontoxic doses of CDDP in combination with anti-IL-6 mAb. When we tested if the synergistic effect of anti-IL-6 and cisplatin could restore the ability of K562 mutant cells to undergo apoptosis, we found the typical DNA laddering in these cells, even in the presence of a nontoxic dose of the drug. Treatment of cells with anti-IL-6 reduced the levels of glutathione. The current studies show that anti-IL-6 mAb sensitized CDDP-resistant K562 cells to CDDP by induction of apoptotic death and the reduction of glutathione levels might be implicated in the enhanced cytotoxicity observed.