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The Journal of Biological Chemistry Apr 2004The African trypanosome Trypanosoma brucei transcribes the active variant surface glycoprotein (VSG) gene from one of about 20 VSG expression sites (ESs). In order to...
The African trypanosome Trypanosoma brucei transcribes the active variant surface glycoprotein (VSG) gene from one of about 20 VSG expression sites (ESs). In order to study ES control, we made reporter lines with a green fluorescent protein gene inserted behind the promoter of different ESs. We attempted to disrupt the silencing machinery, and we used fluorescence-activated cell sorter analysis for the rapid and sensitive detection of ES up-regulation. We find that a range of treatments that either block nuclear DNA synthesis, like aphidicolin, or modify DNA-like cisplatin and 1-methyl-3-nitro-1-nitrosoguanidine results in up-regulation of silent ESs. Aphidicolin treatment was the most effective, with almost 80% of the cells expressing green fluorescent protein from a silent ES. All of these treatments blocked the cells in S phase. In contrast, a range of toxic chemicals had little or no effect on expression. These included berenil and pentamidine, which selectively cleave the mitochondrial kinetoplast DNA, the metabolic inhibitors suramin and difluoromethylornithine, and the mitotic inhibitor rhizoxin. Up-regulation also affected other RNA polymerase I (pol I) transcription units, as procyclin genes were also up-regulated after cells were treated with either aphidicolin or DNA-modifying agents. Strikingly, this up-regulation of silent pol I transcription units was bloodstream form-specific and was not observed in insect form T. brucei. We postulate that the redistribution of a limiting bloodstream form-specific factor involved in both silencing and DNA repair results in the derepression of normally silenced pol I transcription units after DNA damage.
Topics: Animals; Antiprotozoal Agents; Aphidicolin; Blood; DNA Damage; Diminazene; Enzyme Inhibitors; Gene Silencing; Genes, Reporter; Green Fluorescent Proteins; Luminescent Proteins; Membrane Glycoproteins; Pentamidine; Protozoan Proteins; RNA Polymerase I; S Phase; Suramin; Transcription, Genetic; Trypanocidal Agents; Trypanosoma brucei brucei; Up-Regulation; Variant Surface Glycoproteins, Trypanosoma
PubMed: 14726511
DOI: 10.1074/jbc.M312307200 -
DNA Research : An International Journal... 1995Complete nucleotide sequences of three kinds of rice beta-tubulin cDNA clones (pTUB22, R1623 and R2242) were determined. Southern hybridization indicated that these...
Complete nucleotide sequences of three kinds of rice beta-tubulin cDNA clones (pTUB22, R1623 and R2242) were determined. Southern hybridization indicated that these beta-tubulins consist of one gene family. Using RFLP mapping, these three beta-tubulin cDNAs were mapped to different chromosomes indicating at least three loci for the beta-tubulin gene. The deduced amino acid sequences of these cDNAs showed a high similarity to other plant beta-tubulins. The asparagine residue located at the 100th amino acid from the N-terminus of plant beta-tubulins was also conserved with these three beta-tubulins. This asparagine is thought to be responsible for the sensitivity against rhizoxin, the toxin of the pathogen of rice seedling blight, Rhizopus sp. a soil-borne microorganism. Expression of the three beta-tubulin genes was analyzed by Northern blotting and all three clones were expressed in root, the possible target tissue of rhizoxin. These results suggest that these clones are candidates of beta-tubulins targeted by rhizoxin.
Topics: Amino Acid Sequence; Chromosome Mapping; Chromosomes, Fungal; Cloning, Molecular; DNA, Complementary; Molecular Sequence Data; Multigene Family; Oryza; Sequence Homology, Amino Acid; Structure-Activity Relationship; Tubulin
PubMed: 7788525
DOI: 10.1093/dnares/2.1.21 -
British Journal of Cancer Nov 1995Rhizoxin is a tubulin-binding anti-neoplastic agent which is active in a range of murine tumour models. The recommended schedule, of intravenous (i.v.) bolus... (Clinical Trial)
Clinical Trial
Rhizoxin is a tubulin-binding anti-neoplastic agent which is active in a range of murine tumour models. The recommended schedule, of intravenous (i.v.) bolus administration at a dose of 2 mg m-2 every 3 weeks, has been assessed in three phase II trials of ovarian, renal and colorectal cancer. In general terms the drug was fairly well tolerated, but the response rate was disappointing: 0/18, colorectal cancer; 0/18, renal cancer; 1 partial response (PR)/17, ovarian cancer.
Topics: Adult; Aged; Antineoplastic Agents, Phytogenic; Colorectal Neoplasms; Female; Humans; Kidney Neoplasms; Lactones; Macrolides; Middle Aged; Ovarian Neoplasms; Treatment Outcome
PubMed: 7577480
DOI: 10.1038/bjc.1995.498