Functional analysis of the invariant residue G791 of Escherichia coli 16S rRNA

The nucleotide at position 791(G791) of E. coli 16S rRNA was previously identified as an invariant residue for ribosomal function. In order to characterize the functional role of G791, base substitutions were introduced at this position, and mutant ribosomes were analyzed with regard to their protein synthesis ability, via the use of a specialized ribosome system. These ribosomal RNA mutations attenuated the ability of ribosomes to conduct protein synthesis by more than 65%. A transition mutation (G to A) exerted a moderate effect on ribosomal function, whereas a transversion mutation (G to C or U) resulted in a loss of protein synthesis ability of more than 90%. The sucrose gradient profiles of ribosomes and primer extension analysis showed that the loss of protein-synthesis ability of mutant ribosomes harboring a base substitution from G to U at position 791 stems partially from its inability to form 70S ribosomes. These findings show the involvement of the nucleotide at position 791 in the association of ribosomal subunits and protein synthesis steps after 70S formation, as well as the possibility of using 16S rRNA mutated at position 791 for the selection of second-site revertants in order to identify ligands that interact with G791 in protein synthesis.     

Mithramycin inhibits etoposide resistance in glucose-deprived HT-29 human colon carcinoma cells

Physiological cell conditions such as glucose deprivation and hypoxia play roles in the development of drug resistance in solid tumors. These tumor-specific conditions cause decreased expression of DNA topoisomerase IIalpha, rendering cells resistant to topo II target drugs such as etoposide. Thus, targeting tumor-specific conditions such as a low glucose environment may be a novel strategy in the development of anticancer drugs. On this basis, we established a novel screening program for anticancer agents with preferential cytotoxic activity in cancer cells under glucose-deprived conditions. We recently isolated an active compound, AA-98, from Streptomyces sp. AA030098 that can prevent stress-induced etoposide resistance in vitro. Furthermore, LC-MS and various NMR spectroscopic methods identified AA-98 as mithramycin, which belongs to the aureolic acid group of antitumor compounds. We found that mithramycin prevents the etoposide resistance that is induced by glucose deprivation. The etoposide-chemosensitive action of mithramycin was just dependent on strict low glucose conditions, and resulted in the selective cell death of etoposide-resistant HT-29 human colon cancer cells.     

Synthesis, in vitro assay, and molecular modeling of new piperidine derivatives having dual inhibitory potency against acetylcholinesterase and Abeta1-42 aggregation for Alzheimer's disease therapeutics

With the goal of developing Alzheimer's disease therapeutics, we have designed and synthesized new piperidine derivatives having dual action of acetylcholinesterase (AChE) and beta-amyloid peptide (Abeta) aggregation inhibition. For binding with the catalytic site of AChE, an ester with aromatic group was designed, and for the peripheral site, another aromatic group was considered. And for intercalating amyloid-beta oligomerization, long and linear conformation with a lipophilic group was considered. The synthetic methods employed for the structure with dual action depended on alcohols with an aromatic ring and the substituted benzoic acids, which are esterificated in the last step of the synthetic pathway. We screened these new derivatives through inhibition tests of acetylcholinesterase, butyrylcholinesterase (BChE), and Abeta(1-42) peptide aggregation, AChE-induced Abeta(1-42) aggregation. Our results displayed that compound 12 showed the best inhibitory potency and selectivity of AChE, and 29 showed the highest selectivity of BChE inhibition. Compounds 15 and 12 had inhibitory activities against Abeta(1-42) aggregation and AChE-induced Abeta aggregation. In the docking model, we confirmed that 4-chlorobenzene of 12 plays the parallel pi-pi stacking against the indole ring of Trp84 in the bottom gorge of AChE. Because the benzyhydryl moiety of 12 covered the peripheral site of AChE in a funnel-like shape, 12 showed good inhibitory potency against AChE and could inhibit AChE-induced Abeta(1-42) peptide aggregation.     

Selective estrogen receptor degraders with novel structural motifs induce regression in a tamoxifen-resistant breast cancer xenograft

Potent estrogen receptor ligands typically contain a phenolic hydrogen-bond donor. The indazole of the selective estrogen receptor degrader (SERD) ARN-810 is believed to mimic this. Disclosed herein is the discovery of ARN-810 analogs which lack this hydrogen-bond donor. These SERDs induced tumor regression in a tamoxifen-resistant breast cancer xenograft, demonstrating that the indazole NH is not necessary for robust ER-modulation and anti-tumor activity.     

Genomic analysis of facultatively oligotrophic haloarchaea of the genera Halarchaeum,Halorubrum, and Halolamina,isolated from solar salt

Archives of Microbiology - Extremely halophilic archaea (haloarchaea) belonging to the phylum Euryarchaeota have been found in high-salinity environments. In this study, Halarchaeum sp. CBA1220,...     

Neurogenesis of bone marrow-derived mesenchymal stem cells onto β-mercaptoethanol-loaded PLGA film

Bone marrow-derived mesenchymal stem cells (BMSCs) are of particular interest in the field of tissue engineering because of their potential to differentiate into osteoblasts, chondrocytes, and neuronal cells. In order to promote the differentiation of BMSCs into specific cell types, appropriate scaffold biomaterials and bioactive molecules that can support the differentiation of BMSCs into specific cell types are needed. We hypothesized that β-mercaptoethanol (BME), which has been reported to induce the differentiation of BMSCs into neural-like cells, promotes BMSCs to differentiate into neural-like cells when BME is added to polymeric scaffolds containing the BMSCs. We fabricated biocompatible film shaped scaffolds composed of poly(lacti-co-glycolic) acid (PLGA) and various concentrations of BME to confirm that BME-promoted differentiation of BMSCs is concentration-dependent. Cell proliferation increased as the BME concentration in the films increased at the early stage, and the proliferation rate remained similar on the PLGA films for 3 weeks following the BMSC seeding. The expression of neuronal markers in differentiated BMSCs was assessed by RT-PCR. At 2- and 3-week time-points, mRNA expression of neurofilament and neuron specific enolase was significantly increased in PLGA/BME films containing 400 μM BME compared to PLGA films. Thus, we have identified BMSC-seeded PLGA/BME films with 200 μM and 400 μM BME as potentially useful candidates for neural tissue engineering applications by promoting BMSC proliferation and differentiation towards neural-like cells.     

Prioritization of SNPs for genome-wide association studies using an interaction model of genetic variation,gene expression,and trait variation

The identification of true causal loci to unravel the statistical evidence of genotype-phenotype correlations and the biological relevance of selected single-nucleotide polymorphisms (SNPs) is a challenging issue in genome-wide association studies (GWAS). Here, we introduced a novel method for the prioritization of SNPs based on p-values from GWAS. The method uses functional evidence from populations, including phenotype-associated gene expressions. Based on the concept of genetic interactions, such as perturbation of gene expression by genetic variation, phenotype and gene expression related SNPs were prioritized by adjusting the p-values of SNPs. We applied our method to GWAS data related to drug-induced cytotoxicity. Then, we prioritized loci that potentially play a role in druginduced cytotoxicity. By generating an interaction model, our approach allowed us not only to identify causal loci, but also to find intermediate nodes that regulate the flow of information among causal loci, perturbed gene expression, and resulting phenotypic variation.     

HIV-1 clinical isolates resistant to CCR5 antagonists exhibit delayed entry kinetics that are corrected in the presence of drug

HIV CCR5 antagonists select for env gene mutations that enable virus entry via drug-bound coreceptor. To investigate the mechanisms responsible for viral adaptation to drug-bound coreceptor-mediated entry, we studied viral isolates from three participants who developed CCR5 antagonist resistance during treatment with vicriviroc (VCV), an investigational small-molecule CCR5 antagonist. VCV-sensitive and -resistant viruses were isolated from one HIV subtype C- and two subtype B-infected participants; VCV-resistant isolates had mutations in the V3 loop of gp120 and were cross-resistant to TAK-779, an investigational antagonist, and maraviroc (MVC). All three resistant isolates contained a 306P mutation but had variable mutations elsewhere in the V3 stem. We used a virus-cell β-lactamase (BlaM) fusion assay to determine the entry kinetics of recombinant viruses that incorporated full-length VCV-sensitive and -resistant envelopes. VCV-resistant isolates exhibited delayed entry rates in the absence of drug, relative to pretherapy VCV-sensitive isolates. The addition of drug corrected these delays. These findings were generalizable across target cell types with a range of CD4 and CCR5 surface densities and were observed when either population-derived or clonal envelopes were used to construct recombinant viruses. V3 loop mutations alone were sufficient to restore virus entry in the presence of drug, and the accumulation of V3 mutations during VCV therapy led to progressively higher rates of viral entry. We propose that the restoration of pre-CCR5 antagonist therapy HIV entry kinetics drives the selection of V3 loop mutations and may represent a common mechanism that underlies the emergence of CCR5 antagonist resistance.