microRNAs – kleine Moleküle ganz groß

microRNAs, regulators of complex phenotypes microRNAs (miRNAs) are small, non‐coding RNAs that regulate a number of biological processes, including development. Due to their mode of action some miRNAs are also causally involved in diseases like cancer. miRNAs bind base‐complementary mRNAs and lead to either degradation of the bound mRNAs or translational repression. Both result in decreased protein levels of the particular target. miRNA selectivity is governed via a very short, just six to seven nucleotides long, ”seed" sequence which is likely to exist in many mRNAs. miRNAs are, therefore, believed to target a larger number of mRNAs, each, leading to the concerted regulation of functionally connected proteins and to thus substantially contribute to phenotypes. We have performed several screenings which suggest that, indeed, many miRNAs regulate a larger number of proteins. However, we also showed that the proteins we tested were regulated by a larger number of miRNAs each. The complexity of miRNA regulation opens new avenues towards reaching a molecular understanding of disease phenotypes via the integrated consideration of coordinated regulations.     

Particle and crystal symmetry of erysimum latent virus

Erysimum latent virus, a tymovirus, contains 180 protein subunits arranged in a T = 3 icosahedral surface lattice. A cubic crystal form (with space group P2₁3 and $\text{a = 414}\text{A}\text{?}$) and a monoclinic form (space group B2, $\text{a = 442}\text{A}\text{?}\text{, b = 422}\text{A}\text{?}\text{, c = 387}\text{A}\text{?}\text{, γ = 95 °}$) have been observed. The asymmetric units of the two crystal forms contain one-third and one whole virus particle, respectively. Two possible packing arrangements of the virus particles in the monoclinic unit cell have been deduced from the low-angle diffraction patterns. X-ray diffraction data from the monoclinic crystals extend to at least 3·7 Å resolution.     

Excess folate during adolescence suppresses thyroid function with permanent deficits in motivation and spatial memory

Cognitive and memory deficits can be caused or exacerbated by dietary folate deficiency, which has been combatted by the addition of folate to grains and dietary supplements. The recommended dose of the B9 vitamin folate is 400 μg/day for adolescents and non-pregnant adults, and consumption above the recommended daily allowance is not considered to be detrimental. However, the effects of excess folate have not been tested in adolescence when neuro and endocrine development suggest possible vulnerability to long-term cognitive effects. We administered folate-supplemented (8.0 mg folic acid/kg diet) or control lab chow (2.7 mg folic acid/kg diet) to rats ad libitum from 30 to 60 days of age, and subsequently tested their motivation and learning and memory in the Morris water maze. We found that folate-supplemented animals had deficits in motivation and spatial memory, but they showed no changes of the learning- and memory-related molecules growth-associated protein-43 or Gs-α subunit protein in the hippocampus. They had decreased levels of thyroxine (T4) and triiodothyronine (T3) in the periphery and decreased protein levels of thyroid receptor-α1 and -α2 (TRα1 and TRα2) in the hippocampus. The latter may have been due to an observed increase of cytosine-phosphate-guanosine island methylation within the putative thyroid hormone receptor-α promoter, which we have mapped for the first time in the rat. Overall, folate supplementation in adolescence led to motivational and spatial memory deficits that may have been mediated by suppressed thyroid hormone function in the periphery and hippocampus.     

Inter-simple-sequence repeat (ISSR)-PCR for the identification of saprophytic strains of Leptospira

The inter-simple-sequence repeat (ISSR) primers that anneal to a simple repeat of various length and at non-repetitive motifs at 3 and 5 end were attempted for PCR amplification of Leptospira genome. Of the six ISSR primers tested, namely, (AG)₈T, (AG)₈C, (AG)₈G, (CA)₈A, (TG)₈C and (TG)₈G, only primer (AG)₈T produced amplification of 1000 bp in the two non-pathogenic Leptospira species tested, viz; Leptospira biflexa serovar patoc and L. meyeri serovar ranarum, with no amplification in any of the 16 standard pathogenic serovars tested. The remaining five ISSR primers did not exhibit any amplification of the Leptospira genome in either pathogenic or non-pathogenic species. From among 35 Leptospira isolates recovered from hospitalized patients with pyrexia of unknown origin and/or febrile jaundice (12 in number) and from different environmental water sources (23 in number), (AG)₈T ISSR-PCR correctly identified all the 22 isolates from water sources that were confirmed to be non-pathogenic by conventional tests. The results therefore, confirmed the ability of a primer, based on simple-sequence repeat motif, to produce a fragment that is useful as a group genetic marker in Leptospira species. The single nucleotide anchor, T, at the 3 end of the primer appeared to play an important role in differentiation of pathogenic and non-pathogenic species of Leptospira. Multiplex PCR, using ISSR primer, (AG)₈T and the reported 16S rRNA gene primers, specific for pathogenic Leptospira species, or the 23S rRNA Leptospira genus specific primers, provided clear identification of serovars and isolates into pathogenic or non-pathogenic groups.     

ε-PolyLysine production from sugar cane molasses by a new isolates of Bacillus sp. and optimization of the fermentation condition

An attempt was made to use cane molasses as a culture medium for ε-PolyLysine (ε-PL) production by a natural bacterial isolate. The bacterium was identified as Bacillus sp., as confirmed by 16S rDNA sequence analysis. A BLAST result of the sequence indicated that the closest relative of this Bacillus BHU strain was B. thuringiensis, with 97 % homology. The molasses was found to be a better culture medium compared to commonly used culture media comprised of either glucose or glycerol as a carbon source. The various physicochemical parameters were studied for culture growth and polymer production, and were further optimized using response surface methodology (RSM). The correlation coefficient of the resulting model was found to be R ²?=?0.9828. The RSM predicted optimum conditions for ε-PL production (2.46 g/l) by the Bacillus strain was achieved by using molasses, 59.7 g/l; yeast extract, 15.2 mg/l; pH, 6.8 and fermentation time, 42 h at 30 °C. This study represents the first report on the potential application of cane molasses (a byproduct of sugarcane industries) as a culture medium for ε-PL production by Bacillus species. The specific Bacillus strain used in the present study can be exploited for developing a novel technology using inexpensive renewable resources for ε-PL production, a polymer of commercial interest.     

Induction of heme oxygenase-1 by chamomile protects murine macrophages against oxidative stress

AimsProtection of cells from oxidative insult may be possible through direct scavenging of reactive oxygen species, or through stimulation of intracellular antioxidant defense mechanisms by induction of antioxidant gene expression. In this study we investigated the cytoprotective effect of chamomile and elucidated the underlying mechanisms.Main methodsThe cytoprotective effect of chamomile was examined on H₂O₂-induced cellular stress in RAW 264.7 murine macrophages.Key findingsRAW 264.7 murine macrophages treated with chamomile were protected from cell death caused by H₂O₂. Treatment with 50 μM H₂O₂ for 6 h caused significant increase in cellular stress accompanied by cell death in RAW 264.7 macrophages. Pretreatment with chamomile at 10–20 μg/mL for 16 h followed by H₂O₂ treatment protected the macrophages against cell death. Chamomile exposure significantly increased the expression of antioxidant enzymes viz. heme oxygenase-1 (HO-1), peroxiredoxin-1 (Prx-1), and thioredoxin-1 (Trx-1) in a dose-dependent manner, compared with their respective controls. Chamomile increased nuclear translocation of Nrf2 with increased phosphorylated Nrf2 levels, and binding to the antioxidant response element in the nucleus.SignificanceThese molecular findings for the first time provide insights into the mechanisms underlying the induction of phase 2 enzymes through the Keap1-Nrf2 signaling pathway by chamomile, and provide evidence that chamomile possesses antioxidant and cytoprotective properties.     

Disulfiram is a potent modulator of multidrug transporter Cdr1p of Candida albicans

To find novel drugs for effective antifungal therapy in candidiasis, we examined disulfiram, a drug used for the treatment of alcoholism, for its role as a potential modulator of Candida multidrug transporter Cdr1p. We show that disulfiram inhibits the oligomycin-sensitive ATPase activity of Cdr1p and 2.5mM dithiothreitol reverses this inhibition. Disulfiram inhibited the binding of photoaffinity analogs of both ATP ([alpha-(32)P]8-azidoATP; IC(50)=0.76 microM) and drug-substrates ([(3)H]azidopine and [(125)I]iodoarylazidoprazosin; IC(50) approximately 12 microM) to Cdr1p in a concentration-dependent manner, suggesting that it can interact with both ATP and substrate-binding site(s) of Cdr1p. Furthermore, a non-toxic concentration of disulfiram (1 microM) increased the sensitivity of Cdr1p expressing Saccharomyces cerevisiae cells to antifungal agents (fluconazole, miconazole, nystatin, and cycloheximide). Collectively these results demonstrate that disulfiram reverses Cdr1p-mediated drug resistance by interaction with both ATP and substrate-binding sites of the transporter and may be useful for antifungal therapy.     

Search for new pharmacophores for antimalarial activity. Part I: Synthesis and antimalarial activity of new 2-methyl-6-ureido-4-quinolinamides

A total of 80 new 2-methyl-6-ureido-4-quinolinamides were synthesized and evaluated for their antimalarial activity. Several analogs elicited the antimalarial effect at MIC of 0.25 mg/mL against the chlooquine-sensitive P. falciparum strain. The IC₅₀ values of the active compounds were observed to be in ng/mL range and two of the analogs have better IC₅₀ value than the standard chloroquine. In the in vivo assay against mdr CQ resistant P. yoelii N67/P. yoelii nigeriensis, however, none of the compound showed complete suppression of parasitemia on day 7. One of the compounds displayed significant antibacterial effect against several strains of bacteria and was many-fold better than the standard drug gentamicin.     

Improved bioethanol production through simultaneous saccharification and fermentation of lignocellulosic agricultural wastes by <Emphasis Type="Italic">Kluyveromyces marxianus</Emphasis> 6556

The combined effect of simultaneous saccharification and fermentation and separate hydrolysis and fermentation (SHF) for ethanol production by Kluyveromyces marxianus 6556 was studied using two lignocellulosic feedstocks viz., corncob and soybean cake. The ethanologenic efficiency of K. marxianus 6556 was observed as 28% (theoretical yield) in a fermentation medium containing glucose, but, there was no ethanol production by cells grown on xylose. A maximum sugar release of 888 mg/g corncob and 552 mg/g soybean cake was achieved through acid hydrolysis pretreatment. Furthermore, corncob and soybean cake treated with commercial cellulase (100 IU for 48 h) from Trichoderma reesei yielded reducing sugars of 205 and 100 mg/g, respectively. Simultaneous saccharification and fermentation resulted in highest ethanol production of 5.68 g/l on corncob and 2.14 g/l on soybean cake after 48 h of incubation. On the contrary, the presence of inhibitors decreased the overall ethanol yield in the hydrolysates obtained through SHF of corncob and soybean cake.