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At least 289 records · Page 16Linked to original sources
PubMed1988

Initiation of rrn transcription in chloroplasts of Euglena gracilis bacillaris.

The site of initiation of chloroplast rRNA synthesis was determined by S1-mapping and by sequencing primary rRNA transcripts specifically labeled at their 5'-end. Transcription initiates at a single site 53 nucleotides upstream of the 5'-end of the mature 16S rRNA under all growth conditions examined. The initiation site is within a DNA sequence that is highly homologous to and probably derived from a tRNA gene-region located elsewhere in the chloroplast genome. A nearly identical sequence (102 of 103 nucleotides) is present near the replication origin. The near identity of the two sequences suggests a common mode for control of transcription of the rRNA genes and initiation of chloroplast DNA replication. The related sequence in the tRNA gene-region does not appear to serve as a transcript initiation site.

Animals↗
PubMed1983

Use of adenosine 5'-0-(2-thiotriphosphate) for revealing of newly initiated transcripts in isolated rat liver nuclei.

Initiation of DNA-dependent RNA synthesis in isolated rat liver nuclei was studied with adenosine 5'-0-(2-thiotriphosphate), (beta-S-ATP), as a precursor. The newly made RNA labelled with sulfur at 5'-triphosphate termini (thio-RNA) was isolated by affinity chromatography on a mercury-agarose column. Sulfur label can be removed from thio-RNA by digestion with phosphodiesterase I and nucleotide pyrophosphatase. Gel electrophoresis revealed that thio-RNA synthesized during 30 min was composed of 4S-35S molecules with three prevailing classes grouped around 4S-5S, 16S and approximately 35S. Differential sensitivity of the thio-RNA classes to low (1 microgram/ml) and high (200 micrograms/ml) concentrations of alpha-amanitin disclosed that beta-S-ATP was used for initiation of transcription by all three classes of RNA polymerases, and that thio-RNA included molecules as large as 18S initiated by RNA polymerase II. Thio-RNA resistant even to high doses of alpha-amanitin represents probably a product of RNA polymerase I which was initiated and elongated up to 35S.

Adenosine Triphosphate↗
PubMed1997

Assignment of the human gene encoding eukaryotic initiation factor 4E (EIF4E) to the region q21-25 on chromosome 4.

We recently cloned genomic sequences containing the promoter region for the messenger RNA cap binding protein (eIF4E). As the rate-limiting step in translation, eukaryotic initiation factor 4E is important in cellular growth control. Using oligonucleotide primers specific for the promoter region in polymerase chain reactions (PCR), we amplified the human gene in a chromosome 4-specific human/rodent somatic cell panel. This panel mapped single copy genomic sequences for eIF4E in the region 4q21 to 4q25.

Animals↗
PubMed2006

The Nudix hydrolase superfamily.

Nudix hydrolases are found in all classes of organism and hydrolyse a wide range of organic pyrophosphates, including nucleoside di- and triphosphates, dinucleoside and diphosphoinositol polyphosphates, nucleotide sugars and RNA caps, with varying degrees of substrate specificity. Some superfamily members, such as Escherichia coli MicrotT, have the ability to degrade potentially mutagenic, oxidised nucleotides while others control the levels of metabolic intermediates and signalling compounds. In prokaryotes and simple eukaryo tes, the number of Nudix genes varies from 0 to over 30, reflecting the metabolic complexity and adaptability of the organism. Mammals have around 24 Nudix genes, several of which encode more than one variant. This review integrates the sizeable recent literature on these proteins with information from global functional genomic studies to provide some insights into the possible roles of different superfamily members in cellular metabolism and homeostasis and to stimulate discussion and further research into this ubiquitous protein family.

Animals↗
PubMed2004

Characterisation of a beta-tubulin gene from the monogenean parasite, Gyrodactylus salaris Malmberg, 1957.

The cDNA and two partial genomic sequences of beta-tubulin genes have been isolated from the monogenean parasite Gyrodactylus salaris. The cDNA sequence is not represented by either of the genomic sequences, implying that at least three isotypes of the gene exist in G. salaris. The sequences show regions of high homology with other helminth beta-tubulin genes. This represents the first isolation of a beta-tubulin gene from a monogenean and contributes to the overall characterisation of these genes within the helminths. This is an important area, as anthelmintic resistance is increasing against benzimidazole drugs that target the beta-tubulin gene. Benzimidazole drugs have been tested successfully against Gyrodactylus parasites, but their use is not widespread. Should it increase, analysis of the beta-tubulin gene may provide a tool for monitoring resistance development and improving management practises. Use of the beta-tubulin gene in the identification of Gyrodactylus species may prove complex due to the presence of different isotypes.

Amino Acid Sequence↗
PubMed2006

The intraleader AUG nucleotide sequence context is important for equine arteritis virus replication.

The 5(-terminal leader sequence of the equine arteritis virus (EAV) genome contains an open reading frame (ORF) with an AUG codon in a suboptimal context for initiation of protein synthesis. To investigate the significance of this intraleader ORF (ILO), an expression plasmid was generated carrying a DNA copy of the subgenomic mRNA7 behind a T7 promoter. Capped RNA transcribed from this construct was shown to direct, in an in vitro translation system, the synthesis of leader peptide as well as N protein. Site-directed mutations aimed to either optimize or weaken the sequence context of the ILO start codon affected leader peptide synthesis as predicted; no peptide was detected when the initiation codon was incapacitated. Translation of the downstream N gene was inversely affected by leader peptide production, consistent with a ribosomal leaky scanning mechanism. To investigate the role of the leader peptide in the EAV replication life cycle we generated, using an infectious EAV cDNA clone, two mutant viruses in one of which the ILO start codon was in an optimal Kozak context for translation initiation while in the other the codon was again incapacitated. Surprisingly, both mutant viruses were equally viable and exhibited similar phenotypes in BHK-21 cells. However, their replication kinetics and viral yields were reduced relative to that of the wild-type parental virus, as were their plaque sizes. Importantly, the mutations introduced into the viruses appeared to be rapidly and precisely repaired upon passaging. Already after one viral passage a significant fraction of the viruses had regained the wild-type sequence as well as its phenotype. The results demonstrate that EAV replication is not dependent on the synthesis of the intraleader peptide. Rather, the leader peptide does not seem to have any function in the EAV life cycle. As we discuss, the available data indicate that the ILO 5( nucleotide sequence per se, not its functioning in translation initiation, is of critical importance for EAV replication.

Animals↗
PubMed2006

Production of a full-length infectious GFP-tagged cDNA clone of Beet mild yellowing virus for the study of plant-polerovirus interactions.

The full-length cDNA of Beet mild yellowing virus (Broom's Barn isolate) was sequenced and cloned into the vector pLitmus 29 (pBMYV-BBfl). The sequence of BMYV-BBfl (5721 bases) shared 96% and 98% nucleotide identity with the other complete sequences of BMYV (BMYV-2ITB, France and BMYV-IPP, Germany respectively). Full-length capped RNA transcripts of pBMYV-BBfl were synthesised and found to be biologically active in Arabidopsis thaliana protoplasts following electroporation or PEG inoculation when the protoplasts were subsequently analysed using serological and molecular methods. The BMYV sequence was modified by inserting DNA that encoded the jellyfish green fluorescent protein (GFP) into the P5 gene close to its 3' end. A. thaliana protoplasts electroporated with these RNA transcripts were biologically active and up to 2% of transfected protoplasts showed GFP-specific fluorescence. The exploitation of these cDNA clones for the study of the biology of beet poleroviruses is discussed.

Beta vulgaris↗
PubMed1981

The region of transcriptional initiation in Lytechinus variegatus rRNA genes.

A sea urchin ribosomal DNA 1.9 kilobase BamHI fragment adjacent to the 5' end of the 18 S gene has been mapped with the restriction enzymes, XhoI, EcoRi, SmaI and HinfI. A 270 basepair fragment which most likely contains the 5' end of the presumed primary transcript of rRNA was identified by hybridization of [32P]DNA fragments to total nuclear RNA separated on methylmercury hydroxide gels and bound to diazobenzyloxymethyl paper. Under these denaturing conditions the size of Lytechinus variegatus precursor rRNA was determined to be 7.2 kilobases (33 S).

Animals↗
PubMed1987

Reversion of 7-methylguanosine 5'-phosphate inhibition of mRNA translation by polysomal and soluble factors isolated from Saccharomyces cerevisiae.

Protein fractions that overcome m7GMP inhibition of mRNA translation have been purified from the yeast S. cerevisiae. An active fraction isolated from polysomes contains two polypeptides of 220- and 190-kDa. The active fraction isolated from postribosomal supernatant contains a major polypeptide of 28-kDa and other species of 32-, 24-, 22- and 21-kDa, and sediments in sucrose gradients as a high molecular weight complex of about 200,000. This fraction restored yeast mRNA translation in reticulocyte lysates under conditions of yeast and globin mRNA competition; however, this effect was not observed with the 220- and 190-kDa polypeptides from polysomes. Nevertheless, translation of yeast mRNA was stimulated by a partially purified fraction containing a 28-kDa polypeptide from polysomes.

Fungal Proteins↗
PubMed1989

Use of antisense RNA to help identify a genomic clone for the 5' region of mouse beta-glucuronidase.

It was possible to gauge the inhibition of mouse beta-glucuronidase expression by injecting RNA, made from both strands of subclones of a cosmid containing the complete gene, into mouse blastomeres at the four-cell stage. Although our initial screen did not identify the 5' region, we were able to isolate a subclone containing homology to 20 bp coding for N-terminal amino acids of rat and human beta-glucuronidase structural genes. Antisense RNA prepared from one strand of the 350 bp Pst I subclone inhibited beta-glucuronidase expression by 89% while RNA prepared from the other strand had little effect. The subclone appears to correspond to the 350 bp fragment identified by others as one including the ATG start site of mouse beta-glucuronidase.

Animals↗
PubMed1992

Duck hepatitis B virus polymerase produced by in vitro transcription and translation possesses DNA polymerase and reverse transcriptase activities.

Activities of the hepadnavirus polymerases are known to include those of DNA polymerase, reverse transcriptase and RNase H. To date, it has been difficult or impossible to clone and express the product as an active enzyme. In this study, full length capped RNA encoding Duck Hepatitis B Virus (DHBV) polymerase was produced by in vitro transcription from a T7 promoter. The RNA was translated in a rabbit reticulocyte lysate system and produced an 35S-Methionine labelled 79 Kd band on SDS-polyacrylamide gel electrophoresis. The translation product showed DNA polymerase and reverse transcriptase activities on exogenous templates (respectively) of DNA or RNA with random DNA hexamer primers. The same RNA transcripts were also microinjected into Xenopus oocytes, but appeared to be toxic and gave no detectable translation product. Production of hepadnavirus polymerase by in vitro transcription/translation may provide a useful tool for structure/function and pharmacological studies on this important group of polymerases.

Animals↗