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At least 397 records · Page 22Linked to original sources
PubMed1991

Organisation and expression of small subunit ribosomal RNA genes encoded by a 35-kilobase circular DNA in Plasmodium falciparum.

A restriction map of the 35-kb circular DNA molecule of Plasmodium falciparum showed that a region of about 6 kb, encoding both a large and a small subunit ribosomal RNA gene, has been duplicated in inverted orientation. The complete sequence of one small subunit rRNA gene is presented as well as an analysis of transcripts from erythrocytic stage parasites. Comparative sequence analysis of the rRNA gene and the proposed secondary structure of the rRNA suggest that it is of organellar origin. Intriguingly, while some characteristics of the small subunit rRNA gene are similar to mitochondrial sequences, others are more like those of plastids. The origin of the circular DNA molecule and evolutionary implications of its genetic content are discussed.

Animals↗
PubMed1978

Electron microscopic evidence for splicing of Moloney murine leukemia virus RNAs.

Poly (A) containing RNA extracted from Moloney murine leukemia virus infected mouse cells was hybridized with long single-stranded complementary DNA, prepared in detergent disrupted virions. Visualization of the hybrids in the electron microscope revealed among the structures, circles and circles with tails. Measurements performed on the circular molecules revealed two major species with circumferences corresponding to 3 and 8.2 kilobases. The latter structures had identical size to circles obtained after annealing of cDNA with the viral genome, 35S RNA. Circularization of a small viral RNA (3 kb) from infected cells in the RNA-cDNA hybrids is a direct evidence that like the 35S RNA it shares similar nucleotide sequences at both the 5' and 3' ends. The presence of 5' end sequences common to the two RNA species indicates the existence of a spliced viral RNA. Furthermore, based on the circularization of viral RNA in the hybrids, we suggest a new way to quantitate and determine the lengths of spliced RNA in retrovirus infected cells.

Base Sequence↗
PubMed2003

Perturbed folding kinetics of circularly permuted RNAs with altered topology.

The folding pathway of the Tetrahymena ribozyme correlates inversely with the sequence distance between native interactions, or contact order. The rapidly folding P4-P6 domain has a low contact order, while the slowly folding P3-P7 region has a high contact order. To examine the role of topology and contact order in RNA folding, we screened for circular permutants of the ribozyme that retain catalytic activity. Permutants beginning in the P4-P6 domain fold 5 to 20 times more slowly than the wild-type ribozyme. By contrast, 50% of a permuted RNA that disjoins a non-native interaction in P3 folds tenfold faster than the wild-type ribozyme. Hence, the probability of rapidly folding to the native state depends on the topology of tertiary domains.

Animals↗
PubMed1977

A heteroduplex study of the sequence relationships between the RNAs of M-MSV and M-MLV.

The regions of sequence homology between the RNA genomes of a murine sarcoma virus (clone 124 Moloney-MSV) and its parental helper virus (clone 3 Moloney-murine leukemia virus (M-MLV)) have been mapped. Long complementary DNA transcripts of the M-MLV RNA were hybridized to M-MSV RNA, and the structures of the hybrids were observed in the electron microscope. Beginning at the 5' end, the two RNAs are homologous for a region of length 2.25 kb (kilobases). In the next region, of length approximately 4.2 kb on the MLV genome, there are several homology segments between MLV and MSV, but there are also several short sequences present on MLV and deleted in MSV. There is then a major substitution loop; with a sequence (beta L) of length 2.9 kb present on MLV and missing on MSV, and a sequence (beta S) of length 1.5 kb present on MSV and missing on MLV. At the 3' end, there is a homology sequence of length 0.8 kb. On the basis of these results, other data on genes expressed in M-MSV-transformed cells, and by analogy with the avian gene map, we suggest that the gag genes (internal structural proteins) lie in the 2.25 kb region of homology near the 5' ends of M-MSV and M-MLV RNAs, and that the beta S segment contains a sarcoma (src) gene. Some of the heteroduplexes and some of the MLV cDNA/MLV RNA homoduplexes are circular, thus showing that cDNA transcription is initiated at an internal position in the RNA, proceeds to the 5' end, and them "jumps" to the 3' end.

Base Sequence↗
PubMed2002

In vivo RNA-directed transcription, with template switching, by a mammalian RNA polymerase.

These studies support the interpretation that a host polymerase, most likely RNA polymerase II, can not only carry out transcription that is RNA directed, but also achieve template switching on a discontinuous RNA template, and even perform non-templated nucleotide incorporation. As part of an in vivo analysis of the initiation of replication of the RNA genome of human hepatitis delta virus (HDV), a series of linear RNAs containing HDV sequences was tested in order to explain the ability of this host polymerase to initiate RNA-directed RNA synthesis in vivo and produce replicating circular HDV species. The data support the hypothesis that the input linear template RNAs were not converted to circles before transcription but rather that in the process of transcription, the polymerase was able to make an intra-molecular template switch. Furthermore, in certain cases this switch produced small deletions of template sequences, and in some cases even insertion of non-templated sequences. Thus, in an in vivo situation, polymerase II has several important capabilities in addition to what is considered typical DNA-directed transcription.

Base Sequence↗
PubMedDate not supplied

Circular DNA of Entamoeba histolytica encodes ribosomal RNA.

The presence of repeated DNA sequences encoding RNA in Entamoeba histolytica has been reported. In the present study we demonstrate by agarose gel electrophoresis. DNase digestion and electron microscopic analysis that these genes are located on extrachromosomal circular DNA molecules with an approximate size of 26 kb. Detection of replication intermediates suggests the episomal nature of these molecules. Amplified, extrachromosomal rRNA genes appear to be a common feature among the lower eukaryotes, occurring more commonly as linear molecules and less commonly as circles. Entamoeba histolytica is 1 of the few organisms studied in which rRNA genes are located predominantly on extrachromosomal circles.

Animals↗
PubMed1997

Probing the mechanisms of T7 RNA polymerase transcription initiation using photochemical conjugation of psoralen to a promoter.

We have dissected the steps in T7 RNA polymerase transcription initiation using psoralen cross-linking. DNA templates containing cross-links at either -14/-13, -2/-1, or -4/-3 were constructed. These cross-links are within the DNA-contacting region in the initiation complex. A cross-link at -2/-1 did not affect T7 RNA polymerase binding affinity, whereas a cross-link at -14/-13 reduced binding affinity by less than 2-fold. Transcription initiation was completely blocked by cross-links at -14/-13 or at -2/-1. A cross-link at -4/-3 inhibited neither binding nor the first RNA phosphodiester bond but greatly inhibited further RNA chain extension. Circular dichroism spectroscopy revealed that DNA melting in the -4/-3 cross-link was greatly inhibited, indicating that inhibition of RNA chain extension was a melting defect. Transcription shutoff on the -14/-13 cross-link may be due to inhibition of conformational changes in the polymerase-DNA complex. Because the -2/-1 cross-link is immediately upstream of the start site (+1), open complex formation may have been completely inhibited by this cross-link, accounting for the shutoff of transcription. Thus, depending on their location, psoralen cross-links affected different steps in the initiation process. We propose that promoter melting is progressive and that melting of one or two bp upstream of the +1 site is sufficient for formation of the first phosphodiester bond while further RNA chain extension within the promoter depends on greater upstream melting of the promoter, which may be required for stabilization of the initiation complex.

Bacteriophage T7↗
PubMed1980

Secondary structures of influenza and Sendai Virus RNAs.

The secondary structures of influenza and Sendai virus RNAs were investigated by thermal denaturation, circular dichroism and proflavine binding methods. In 0.1 M NaCl about 60% of the bases of both RNAs were involved in secondary structure. The melting temperatures (Tm) of both viral RNAs were linear functions of the logarithm of the sodium ion concentration in solution, but under all ionic conditions the melting temperatures of Sendai virus RNA were higher than those of influenza virus RNA. At all ionic strengths the melting range of Sendai virus RNA was less than influenza virus RNA, indicating that the helical regions in Sendai virus RNA were longer than those in influenza virus RNA. Although Sendai virus RNA had a higher thermal stability than influenza virus RNA, hyperchromicity and circular dichroism data showed that Sendai virus RNA had less G+C content (34%) within the double stranded regions than influenza virus RNA (48%). The binding isotherms of Sendai and influenza virus RNA-proflavine complexes were studied at different ionic strengths. The number of binding sites of proflavine with influenza virus RNA were significantly lower than those with Sendai virus RNA. These results demonstrate the essential difference between the secondary and tertiary structures of the RNAs under study.

Base Composition↗
PubMedDate not supplied

Some properties of the viroid inducing peach latent mosaic disease.

Analysis by polyacrylamide gel electrophoresis of nucleic acid extracts from different peach samples, healthy or infected with the peach latent mosaic (PLM) disease, demonstrated the association of this disease with an RNA exhibiting the electrophoretic properties typical of circular viroid molecules. This RNA was called peach latent mosaic viroid (PLMV), since a purified preparation of it, when inoculated into GF 305 peach seedlings induced characteristic symptoms of PLM disease. PLMV was estimated to have a molecular size in the range of 330-340 bases, by comparison of its electrophoretic mobility under denaturing conditions with those of several viroid RNA. Dot-blot analysis showed that PLMV had a sequence clearly different from other viroids, including citrus exocortis viroid, apple scar skin viroid (ASSV), hop stunt viroid (HSV) and avocado sunblotch viroid. The possible significance of the limited sequence homology shared by PLMV with HSV, and especially with ASSV, is discussed.

Fruit↗
PubMed1989

Circular RNAs: relics of precellular evolution?

The demonstration of enzymatic capabilities of certain RNAs, in addition to their well-known template properties, has led to the recognition that RNAs are the only biological macromolecules that can function both as genotype and phenotype, hence raising the possibility of Darwinian selection and precellular evolution at the RNA level in the absence of DNA or protein. Recent models of such precellular RNA systems are patterned after the properties of intron-derived ribozymes. On the basis of a phylogenetic analysis and known properties of certain small plant pathogenic RNAs (viroids and viroid-like satellite RNAs), I suggest that these plant RNAs are more plausible candidates than introns as "living fossils" of a precellular RNA world. Their small size and circularity would have enhanced probability of their survival in error-prone, primitive self-replicating RNA systems and assured complete replication without the need for initiation or termination signals. All of these RNAs possess efficient mechanisms for the precise cleavage of monomers from oligomeric replication intermediates. Some (most viroids) require a host factor, but others (viroid-like satellite RNAs and one viroid) function as self-cleaving RNA enzymes far smaller and simpler than those derived from introns. The question is raised whether introns could have evolved from viroids or viroid-like satellite RNAs rather than vice versa, as has been widely speculated.

Base Sequence↗
PubMed1997

RNA-binding properties of an unusual ribosomal protein TL5 from Thermus thermophilus.

The gene encoding the 5S rRNA-binding ribosomal protein TL5 from Thermus thermophilus, an extremely thermophilic species, was expressed in E. coli. A method for isolation of TL5 from the overproducing strain was developed. Samples of TL5 protein isolated from ribosomes and the overproducing strain displayed identical RNA-binding properties. Circular dichroic spectroscopy was used to calculate the secondary structure of the protein. TL5 was shown to form a stable complex with the 3'-terminal fragment of 5S rRNA, which is similar to the fragment of E. coli RNA that binds to L25 protein. The data suggest that TL5 from T. thermophilus and L25 from E. coli bind to similar sites on the 5S rRNA molecule.

Bacterial Proteins↗
PubMed1975

Physical properties of moloney murine leukemia virus high-molecular-weight RNA: a two subunit structure.

The high-molecular-weight RNA of Moloney murine leukemia virus (MuLV) was analyzed by sedimentation equilibrium ultracentrifugation. Molecular weights of 7.2 x 10(6) and 3.4 x 10(6) were found for the native and subunit forms, respectively, indicating that the native structure is a dimer. S20,w and frictional coefficients were determined for MuLV RNA by analytical velocity centrifugation as a function of ionic strength. The apparent S20,w of native MuLV RNA was 47.3, 57.4, and 66.5 in 0.01, 0.1, and 0.20 M Na+, respectively; the corresponding frictional coefficients were 5.44, 4.48, and 3.87. Native RNA was estimated by circular dichroism to be 85% helical, whereas denatured RNA was 54% helical. Thermal denaturation profiles were obtained from uv absorbance scans. Melting temperatures of 57 and 68 C were obtained for high-molecular-weight RNA in 0.01 M Na+ and 0.122 M Na+, 1mM Mg2+, respectively. van't Hoff plots of the thermal denaturation data gave enthalpies for the helix-coil transition of 21,600 cal (ca. 90,500 J) per mol of cooperatively melting unit in high salt and 19,600 cal (ca. 82,100 J) per mol in low salt, consistent with both base stacking and pairing. The melting of Mu LV RNA occurred over a broad temprange and van't Hoff plots were linear over most of the melting range, indicating a noncooperative process of helix stabilization.

Centrifugation, Density Gradient↗
PubMed1980

Small circular deoxyribonucleic acid of Drosophila melanogaster: homologous transcripts in the nucleus and cytoplasm.

We have recently characterized small circular DNA of Drosophila cultured cells in terms of its average size, sequence complexity, and homology to intermediate repetitive DNA. We show here that transcripts homologous to small circular DNA are present in various RNA fractions. Nuclear poly(A+), nuclear poly(A-), and polysomal poly(A+) RNA drive 10, 7, and 20%, respectively, of in vitro labeled small circular DNA tracer into hybrid. Sequences complementary to small circular DNA are at least 10-fold more concentrated in nuclear poly(A+) RNA than in nuclear poly(A-) or polysomal poly(A+) RNA. We do not detect significant homology between poly(A-) cytoplasmic RNA and small circular DNA. Assuming that only the least complex component of small circular DNA is driven into hybrid and that transcription is asymmetric, we use the results obtained here and previously published data to calculate the sequence complexity and relative concentration of nuclear poly(A+), nuclear poly(A-), and polysomal poly(A+) RNA homologous to small circular DNA.

Animals↗
PubMed2001

Transcription factor Rho does not require a free end to act as an RNA-DNA helicase on an RNA.

Escherichia coli Rho factor is a ring-shaped, homohexameric protein that terminates synthesis of RNA through interactions with the nascent RNA transcript. Because its mechanism of action may involve translocation of the RNA transcript through the hole in its ring structure, its action could depend on the availability of a free 5' terminus. To determine whether Rho's activity is 5'-end-dependent, its ability to bind to and function on a circular derivative of lambda cro mRNA was investigated. The circular derivative was made in vitro by action of RNA ligase on a derivative of lambda cro RNA containing an extra 10-nucleotide sequence near the 5'-end that was complementary to a sequence located near the 3'-end. Rho bound nearly as tightly to the circular derivative RNA as to the standard cro transcript. Rho was also able to readily dissociate a DNA oligonucleotide from its helical complex with the circular RNA in an ATP-dependent reaction. Thus, the action of Rho on a transcript does not depend on the availability of a free 5' terminus.

Adenosine Triphosphate↗
PubMed2006

Isolation and crystallization of a unique size category of recombinant Rabies virus Nucleoprotein-RNA rings.

In order to study the packaging of rabies virus RNA inside the viral nucleocapsid, rabies nucleoprotein was expressed in insect cells. In the cells, it binds to cellular RNA to form long, helical or short circular complexes, depending on the length of the bound RNA. The circular complexes contained from 9 up to 13 N-protomers per ring. Separation of the rings into defined size classes was impossible through regular column chromatographies or gradient centrifugation. The size classes could be separated by native polyacrylamide gel electrophoresis. A large-scale separation was achieved with a 4% native gel using a preparative electrophoresis apparatus. Crystallization trials were set up with N-RNA rings from three size classes and crystals were obtained in all cases. The best diffracting crystals, diffracting up to 6A, contained rings with 11 N-protomers plus an RNA molecule of 99 nucleotides. The diffraction limit was improved to 3.5A by air dehydration prior to flash freezing.

Animals↗