Loss of a conserved 7-methylguanosine modification in 16S rRNA confers low-level streptomycin resistance in bacteria

Streptomycin has been an important drug for the treatment of tuberculosis since its discovery in 1944. But numerous strains of Mycobacterium tuberculosis, the bacterial pathogen that causes tuberculosis, are now streptomycin resistant. Although such resistance is often mediated by mutations within rrs, a 16S rRNA gene or rpsL, which encodes the ribosomal protein S12, these mutations are found in a limited proportion of clinically isolated streptomycin-resistant M. tuberculosis strains. Here we have succeeded in identifying a mutation that confers low-level streptomycin resistance to bacteria, including M. tuberculosis. We found that mutations within the gene gidB confer low-level streptomycin resistance and are an important cause of resistance found in 33% of resistant M. tuberculosis isolates. We further clarified that the gidB gene encodes a conserved 7-methylguanosine (m(7)G) methyltransferase specific for the 16S rRNA, apparently at position G527 located in the so-called 530 loop. Thus, we have identified gidB as a new streptomycin-resistance locus and uncovered a resistance mechanism that is mediated by loss of a conserved m(7)G modification in 16S rRNA. The clinical significance of M. tuberculosis gidB mutation also is noteworthy, as gidB mutations emerge spontaneously at a high frequency of 10(-6) and, once emerged, result in vigorous emergence of high-level streptomycin-resistant mutants at a frequency more than 2000 times greater than that seen in wild-type strains. Further studies on the precise function of GidB may provide a basis for developing strategies to suppress pathogenic bacteria, including M. tuberculosis.     

Synthesis of phosphorothioate oligonucleotide-peptide conjugates by solid phase fragment condensation

Phosphorothioate oligonucleotide-peptide conjugates were synthesized by solid phase fragment condensation (SPFC). Arginine rich peptides could be successfully conjugated in 2.8-13.4% isolated yields. All the products were fully characterized by reversed phase HPLC and MALDI-TOF-MS to give satisfactory results.     

Aminocarbonylation route to tolvaptan

Pd-catalyzed aminocarbonylation between aryl bromide and NH-benzazepinone was effectively carried out to furnish the key intermediate for tolvaptan (up to 85%) in one step.     

Design and synthesis of piperidine farnesyltransferase inhibitors with reduced glucuronidation potential

The design and synthesis of a novel piperidine series of farnesyltransferase (FTase) inhibitors with reduced potential for metabolic glucuronidation are described. The various substitution and exchange of the phenyl group at the C-2 position of the previously described 2-(4-hydroxy)phenyl-3-nitropiperidine 1a (FTase IC(50)=5.4nM) resulted in metabolically stable compounds with potent FTase inhibition (14a IC(50)=4.3nM, 20a IC(50)=3.0nM, and 50a IC(50)=16nM). Molecular modeling studies of these compounds complexed with FTase and farnesyl pyrophosphate are also described.     

Protective effects of carotenoids from saffron on neuronal injury in vitro and in vivo

Crocus sativus L. (saffron) has been used as a spice for flavoring and coloring food preparations, and in Chinese traditional medicine as an anodyne or tranquilizer. Our previous study demonstrated that crocin, a carotenoid pigment of saffron, can suppress the serum deprivation-induced death of PC12 cells by increasing glutathione (GSH) synthesis and thus inhibiting neutral sphingomyelinase (nSMase) activity and ceramide formation. The carotenoid pigments of saffron consist of crocetin di-(beta-d-glucosyl)-ester [dicrocin], crocetin-(beta-d-gentiobiosyl)-(beta-d-glucosyl)-ester [tricrocin] and crocetin-di-(beta-d-gentiobiosyl)-ester [crocin]. Saffron also contains picrocrocin, the substance causing saffron's bitter taste. In this study, to confirm whether neuroprotective effects of saffron are caused solely by crocin, we examined the antioxidant and GSH-synthetic activities of these crocins in PC12 cells under serum-free and hypoxic conditions. Measurements of cell viability, peroxidized membrane lipids and caspase-3 activity showed that the rank order of the neuroprotective potency at a concentration of 10 muM was crocin>tricrocin>dicrocin and picrocrocin (the latter two crocins had a little or no potency). In addition, we show that among these saffron's constituents, crocin most effectively promotes mRNA expression of gamma-glutamylcysteinyl synthase (gamma-GCS), which contributes to GSH synthesis as the rate-limiting enzyme, and that the carotenoid can significantly reduce infarcted areas caused by occlusion of the middle cerebral artery (MCA) in mice.     

Rice NON-YELLOW COLORING1 is involved in light-harvesting complex II and grana degradation during leaf senescence

Chlorophyll degradation is an aspect of leaf senescence, which is an active process to salvage nutrients from old tissues. non-yellow coloring1 (nyc1) is a rice (Oryza sativa) stay-green mutant in which chlorophyll degradation during senescence is impaired. Pigment analysis revealed that degradation of not only chlorophylls but also light-harvesting complex II (LHCII)-bound carotenoids was repressed in nyc1, in which most LHCII isoforms were selectively retained during senescence. Ultrastructural analysis of nyc1 chloroplasts revealed that large and thick grana were present even in the late stage of senescence, suggesting that degradation of LHCII is required for the proper degeneration of thylakoid membranes. Map-based cloning of NYC1 revealed that it encodes a chloroplast-localized short-chain dehydrogenase/reductase (SDR) with three transmembrane domains. The predicted structure of the NYC1 protein and the phenotype of the nyc1 mutant suggest the possibility that NYC1 is a chlorophyll b reductase. Although we were unable to detect the chlorophyll b reductase activity of NYC1, NOL (for NYC1-like), a protein closely related to NYC1 in rice, showed chlorophyll b reductase activity in vitro. We suggest that NYC1 and NOL encode chlorophyll b reductases with divergent functions. Our data collectively suggest that the identified SDR protein NYC1 plays essential roles in the regulation of LHCII and thylakoid membrane degradation during senescence.     

Myostatin and MyoD family expression in skeletal muscle of IGF-1 knockout mice

Insulin-like growth factor-1 (IGF-1) is a positive regulator in proliferation and differentiation of skeletal muscle cells, while myostatin (MSTN) is a member of transforming growth factor beta superfamily that acts as a negative regulator of skeletal muscle mass. The present study was performed to detail whether a correlation exists between MSTN and IGF-1 in skeletal muscle of IGF-1 knockout mice (IGF-1(-/-)) and their wild type (WT; i.e., IGF-1(+/+)) littermates. The body weight of IGF-1(-/-) animals was 32% that of WT littermates. The fiber cross-sectional areas (CSA) and number of fibers in M. rectus femoris of IGF-1(-/-) animals were 49 and 59% those of WT animals, respectively. Thus, muscle hypoplasia of IGF-1(-/-) undoubtedly was confirmed. Myostatin mRNA levels and protein levels were similar between M. gastrocnemius of IGF-1(-/-) and WT animals. Myostatin immunoreactivity was similarly localized in muscle fibers of both IGF-1(-/-) and WT M. rectus femoris. The mRNA levels of MyoD family (Myf5, MyoD, MRF4, myogenin) were differentially expressed in IGF-1(-/-)M. gastrocnemius, in which the mRNA expression of MRF4 and myogenin was significantly lower, whereas there were no changes in the mRNA expression of Myf5 and MyoD. These findings first describe that myostatin expression is not influenced by intrinsic failure of IGF-1, although MRF4 and myogenin are downregulated.     

E-cadherin gene promoter hypermethylation in H. pylori-induced enlarged fold gastritis

BACKGROUND: Promoter hypermethylation of E-cadherin plays an important role on gastric carcinogenesis. We have previously reported that the odds ratio for gastric carcinoma and the prevalence of diffuse-type early gastric carcinoma in Helicobacter pylori-induced enlarged fold gastritis increased with increasing fold width. Thus, we examined E-cadherin methylation in gastric mucosa from H. pylori-induced enlarged fold gastritis before and after H. pylori eradication. Moreover, we analyzed the mechanism of H. pylori infection-induced E-cadherin hypermethylation. MATERIALS AND METHODS: Twenty-three H. pylori-positive patients with enlarged folds, 18 H. pylori-positive and seven H. pylori-negative patients without enlarged folds, were involved in the study. E-cadherin promoter methylation was studied using quantitative methylation-specific polymerase chain reaction. We investigated methylation percentage and DNA methyltransferase activity in gastric cancer cell lines treated with EGF, TNFalpha, and MG132. RESULTS: E-cadherin methylation percentage of the gastric antral and body mucosa in H. pylori-positive patients with enlarged folds was much greater than that in both H. pylori-positive and -negative patients without enlarged folds. After H. pylori eradication, the methylation percentage in six patients with enlarged fold gastritis decreased significantly from 15.6 +/- 3.9 to 8.8 +/- 2.2 (p < .05). Moreover, the methylation was induced by TNFalpha, MG132, and EGF treatment, and DNA methyltransferase activity was induced by EGF treatment in MKN-1 cells. CONCLUSIONS: Our findings suggest that the hypermethylation of E-cadherin promoter might be involved in the process of gastric carcinoma through the specialized factors in H. pylori-induced enlarged fold gastritis.     

Implication of N-glycolylneuraminic acid in regulation of cell adhesiveness of C2C12 myoblast cells during differentiation into myotube cells

Glycoconjugate Journal - A transition of sialic acid (Sia) species on GM3 ganglioside from N-acetylneuraminic acid (Neu5Ac) to N-glycolylneuraminic acid (Neu5Gc) takes place in mouse C2C12 myoblast...     

Folate receptor-targeted novel boron compound for boron neutron capture therapy on F98 glioma-bearing rats

Radiation and Environmental Biophysics - Folic acid (FA) has high affinity for the folate receptor (FR), which is limited expressed in normal human tissues, but over-expressed in several tumor...