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At least 325 records · Page 18Linked to original sources
PubMed1999

Phylogenetic position of the kinetoplastids, Cryptobia bullocki, Cryptobia catostomi, and Cryptobia salmositica and monophyly of the genus Trypanosoma inferred from small subunit ribosomal RNA sequences.

Phylogenetic relationships within the kinetoplastid flagellates were inferred from comparisons of small-subunit ribosomal RNA gene sequences. These included three new gene sequences from Cryptobia bullocki, (2091 bp), Cryptobia catostomi (2090 bp), and Cryptobia salmositica (2091 bp). Trees produced using maximum parsimony and distance-matrix methods (least squares and neighbor-joining) demonstrated with strong bootstrap support, that the kinetoplastids are a monophyletic group divided into two major lineages consistent with the two suborders, Trypanosomatina and Bodonina. Within the trypanosomatid clade, the genus Trypanosoma is a monophyletic group that divides into two groups, the salivarian trypanosomes and the stercorarian trypanosomes. Dimastigella and Rhynchobodo, currently classified in the Bodonina, are basal to the trypanosomatid-bodonid clade, suggesting that the suborder Bodonina is paraphyletic. Further, Trypanoplasma borreli grouped within the Cryptobia clade, and was more closely related to C. salmositica than to either C. bullocki or C. catostomi. This new molecular evidence, coupled with morphological similarities of the two genera, again calls into question the validity of the genus Trypanoplasma.

Animals↗
PubMed2005

Effective inhibition of human cytomegalovirus gene expression and growth by intracellular expression of external guide sequence RNA.

RNase P complexed with external guide sequence (EGS) represents a novel nucleic-acid-based gene interference approach to modulate gene expression. In this study, a functional EGS RNA was constructed to target the overlapping mRNA region of two human cytomegalovirus (HCMV) capsid proteins, the capsid scaffolding protein (CSP) and assemblin. The EGS RNA was shown to be able to direct human RNase P to cleave the target mRNA sequence efficiently in vitro. A reduction of approximately 75%-80% in the mRNA and protein expression levels of both CSP and assemblin and a reduction of 800-fold in viral growth were observed in human cells that expressed the functional EGS, but not in cells that either did not express the EGS or produced a "disabled" EGS that carried nucleotide mutations that precluded RNase P recognition. The action of the EGS is specific as the RNase P-mediated cleavage only reduces the expression of the CSP and assemblin but not other viral genes examined. Further studies of the antiviral effects of the EGS indicate that the expression of the functional EGS has no effect on HCMV genome replication but blocks viral capsid maturation, consistent with the notion that CSP and assemblin play essential roles in HCMV capsid formation. Our study provides the first direct evidence that EGS RNAs effectively inhibit HCMV gene expression and growth. Moreover, these results demonstrate the utility of EGS RNAs in gene therapy applications, including the treatment of HCMV infection by inhibiting the expression of virus-encoded essential proteins.

Animals↗
PubMed1992

Distinct RNA sequences in the gag region of human immunodeficiency virus type 1 decrease RNA stability and inhibit expression in the absence of Rev protein.

The expression of Gag, Pol, Vif, Vpr, Vpu, and Env proteins from unspliced and partially spliced human immunodeficiency virus type 1 (HIV-1) mRNAs depends on the viral protein Rev, while the production of Tat, Rev, and Nef from multiply spliced mRNAs does not require Rev. To investigate the difference between gag and tat mRNAs, we generated plasmids expressing tat-gag hybrid mRNAs. Insertion of the gag gene downstream of the tat open reading frame in the tat cDNA resulted in the inhibition of Tat production. This inhibition was caused, at least in part, by a decrease in the stability of the produced mRNA. Deletions in gag defined a 218-nucleotide inhibitory sequence named INS-1 and located at the 5' end of the gag gene. Further experiments indicated the presence of more than one inhibitory sequence in the gag-protease gene region of the viral genome. The inhibitory effect of INS-1 was counteracted by the positive effect mediated by the Rev-Rev-responsive element interaction, indicating that this sequence is important for Rev-regulated gag expression. The INS-1 sequence did not contain any known HIV-1 splice sites and acted independently of splicing. It was found to have an unusually high AU content (61.5% AU), a common feature among cellular mRNAs with short half-lives. These results suggest that HIV-1 and possibly other lentiviruses have evolved to express unstable mRNAs which require additional regulatory factors for their expression. This strategy may offer the virus several advantages, including the ability to enter a state of low or latent expression in the host.

Blotting, Northern↗
PubMed1984

Highly conserved 5S ribosomal RNA sequences in four rust fungi and atypical 5S rRNA secondary structure in Microstroma juglandis.

The 5S ribosomal RNA nucleotide sequences of five basidiomycetous fungi, Coleosporium tussilaginis , Gymnosporangium clavariaeforme , Puccinia poarum , Endophyllum sempervivi and Microstroma juglandis were determined. Despite high differentiation in their host spectra the four rust species are highly conserved with respect to their 5S rRna sequences, which fit with the basidiomycete cluster 5 described by Walker and Doolittle (1). The sequences obtained from the first three rust fungi were proven to be identical while the sequence from Endophyllum sempervivi showed two base substitutions compared with the other rust fungi. The Microstroma juglandis 5S rRNA sequence differs from all other basidiomycete 5S rRNA sequences published so far in respect to its secondary structure which shows an atypical 'CCA' loop in helix D, but it reveals typical basidiomycetous signature nucleotides. Therefore Microstroma juglandis represents a cluster of its own within the Basidiomycetes. A dendrogram was constructed based on Kimura's "Neutral Theory of Molecular Evolution".

Base Sequence↗
PubMed1993

Translocation of repetitive RNA sequences with the germ plasm in Xenopus oocytes.

Xlsirts are a family of interspersed repeat RNAs from Xenopus laevis that contain from 3 to 13 repeat units (each 79 to 81 nucleotides long) flanked by unique sequences. They are homologous to the mammalian Xist gene that is involved in X chromosome inactivation. Xlsirt RNA appears first in the mitochondrial cloud (Balbiani body) in stage 2 oocytes and is then translocated as island-like structures to the vegetal cortex at early stage 3 coincident with the localization of the germ plasm. Exogenous Xlsirt RNA injected into oocytes translocates to the location of the endogenous RNA at that particular stage. The Xlsirt RNA repeat sequences are required for translocation and can cause the translocation of heterologous unique RNAs to the vegetal cortex.

Animals↗
PubMed2000

The small subunit ribosomal RNA sequence of Strongyloides stercoralis.

The published small subunit rRNA (ssrRNA) gene sequences for Strongyloides ratti and Strongyloides stercoralis are remarkably divergent, particularly in the 5' 400 bases of the approximately 1700 base pair (bp) sequences. This level of divergence between species nominally in the same genus was unprecedented. We have redetermined the ssrRNA sequence of S. stercoralis and find that the published sequence is a chimaera of parasite and fungal segments. The true sequence for S. stercoralis ssrRNA is very similar to that of S. ratti.

Animals↗
PubMed1982

The nucleotide sequence of tRNA4Val of Drosophila melanogaster. Chloroacetaldehyde modification as an aid to RNA sequencing.

The nucleotide sequence of tRNA4Val from Drosophila melanogaster was determined to be pGUUUmCCGUm1GGUG psi AGCGGDU(acp3U)AUCACA psi CUGCCmUIACAm5CGCAGAAGm7GCCCCCGGT psi CGm1AUCCCGGGCGGAAACACCA. It is probable that residue C 49 is modified to m5C. The use of tRNA modified with chloroacetaldehyde to overcome secondary structure problems in sequencing is described.

Acetaldehyde↗
PubMed1989

Specific oligodeoxynucleotide probes obtained through RNA sequencing.

By combining several established techniques we developed a method to test the specificity of mixed oligodeoxynucleotide hybridization probes and to provide the information for the design of long nondegenerate, and therefore more specific probes. Mixed oligodeoxynucleotide probes derived from known peptide sequences are first used to initiate primer extension reactions with poly(A)+RNA as template in the presence of three dNTPs and one ddNTP to generate cDNA transcripts of defined lengths. Comparing the lengths of the cDNA transcripts with the possible nucleic acid sequence coding for the known oligopeptide indicates whether the oligodeoxynucleotide mix hybridizes predominantly to the RNA of interest. In a second step, the oligodeoxynucleotide mix with the highest specificity is used for indirect RNA sequence analysis. This confirms the specificity of the probe and provides information to design a long, highly specific oligodeoxynucleotide probe for the gene of interest. This simple two-step-procedure helps to circumvent the time-consuming procedures of subcloning and sequencing of cross-hybridizing fragments.

Base Sequence↗
PubMed1980

Recent developments in methods for RNA sequencing using in vitro 32P-labeling.

A variety of approaches that utilize in vitro 32P-labeling of RNA and of oligonucleotides in the sequence analysis of RNAs are described. These include 1) methods for 5'- and 3'- end labeling of RNAs; 2) end labeling and sequencing of oligonucleotides present in complete T1 RNase or pancreatic RNase digests of RNA; 3) use of random endonucleases, such as nuclease P1, for terminal sequence analysis of end labeled RNAs; and 4) use of base specific enzymes or chemical reagents in the sequence analysis of end-labeled RNAs. Also described is an approach to RNA sequencing, applied so far to tRNAs, which is based on partial and random alkaline cleavage of an RNA to generate a series of overlapping oligonucleotide fragments, all containing the original 3'-end of the RNA. Analysis of the 5'- end group of each of these oligonucleotides (following 5'-end labeling with 32P) provides the sequence of most of the tRNA. The above methods have been used to derive the sequences of several tRNAs, the ribosomal 5S and 5 x 8S RNAs, a viroid RNA, and large segments of both prokaryotic and eukaryotic ribosomal and messenger RNAs.

Autoradiography↗
PubMed1990

Protection against tobacco mosaic virus infection in transgenic plants requires accumulation of coat protein rather than coat protein RNA sequences.

Transgenic tobacco plants which express a chimeric gene encoding the tobacco mosaic virus (TMV) coat protein (CP) and the TMV 3' untranslated region are protected against infection by TMV. In this study chimeric genes that encode the sequences representing the TMV CP subgenomic RNA, but do not produce protein (because of removal of the initiation codon), and RNA that lacks the tRNA-like sequence of the TMV 3' end were expressed in transgenic plants. Only plants that accumulated CP, regardless of the presence of absence of the 3' end of TMV-RNA, were protected against infection by TMV. The results indicate that the CP per se, rather than TMV RNA, is responsible for the resistance to infection by TMV. Furthermore, the degree of protection is dependent upon the level of accumulated CP.

Base Sequence↗
PubMed1992

Phylogenetic relationships of Sarcocystis species from sheep, goats, cattle and mice based on ribosomal RNA sequences.

Partial sequences of the small subunit ribosomal RNA of four species of Sarcocystis were obtained by reverse transcription of total cellular RNA. The semi-conserved regions of these four species were aligned with homologous sequences of two other Sarcocystis species and a range of other eukaryotes including Toxoplasma, Eimeria and Cryptosporidium. Parsimony analysis of the aligned sequences showed that Sarcocystis and Toxoplasma had a more recent common ancestor with Eimeria than with Cryptosporidium. The six Sarcocystis spp. did not cluster together in this analysis; two monophyletic groups were observed, one formed by the two Sarcocystis spp. with felids as definitive hosts, and another by the four Sarcocystis spp. with canids as definitive hosts. These two clades were segregated by Toxoplasma. An analysis of nucleotide divergence suggests that the difference between the two groups of Sarcocystis spp. is similar to that between invertebrates and vertebrates. The results obtained here question the validity of a separation of the genus Sarcocystis from Toxoplasma and refute classifications that place these two genera into two different subfamilies of the Sarcocystidae.

Animals↗
PubMed2006

Dissection of Prp8 protein defines multiple interactions with crucial RNA sequences in the catalytic core of the spliceosome.

Current models of the core of the spliceosome include a network of RNA-RNA interactions involving the pre-mRNA and the U2, U5, and U6 snRNAs. The essential spliceosomal protein Prp8 interacts with U5 and U6 snRNAs and with specific pre-mRNA sequences that participate in catalysis. This close association with crucial RNA sequences, together with extensive genetic evidence, suggests that Prp8 could directly affect the function of the catalytic core, perhaps acting as a splicing cofactor. However, the sequence of Prp8 is almost entirely novel, and it offers few clues to the molecular basis of Prp8-RNA interactions. We have used an innovative transposon-based strategy to establish that catalytic core RNAs make multiple contacts in the central region of Prp8, underscoring the intimate relationship between this protein and the catalytic center of the spliceosome. Our analysis of RNA interactions identifies a discrete, highly conserved region of Prp8 as a prime candidate for the role of cofactor for the spliceosome's RNA core.

Base Sequence↗
PubMed1980

Differential distribution of poly(A)-containing RNA sequences between the nucleus and post-nuclear supernatant of the lactating guinea-pig mammary gland.

RNA complexity analysis of poly(A)-containing RNA isolated from the lactating guinea-pig mammary gland demontrates that the complexity within the nuclear population was three--four-fold greater than that in the equivalent post-nuclear polyribosomal population [see Craig et al., Biochem. J. (1979) 181, 737-756]. Up to 21 000 different sequences distributed in two abundance groups were detected in the nucleus, whereas 5000 sequences distributed in three abundance groups were present in the post-nuclear population. All poly(A)-containing RNA sequences present in the post-nuclear fraction could be detected in the nuclear poly(A)-containing population. Analysis of the relative distribution of the three post-nuclear abundance populations within the nuclear poly(A)-containing RNA population demonstrates that the abundant and moderately abundant post-nuclear sequences were present a similar concentrations within the nucleus, and comprised the abundant nuclear population. The abundant and moderately abundant post-nuclear sequences were present at 62200 and 785 copies of each sequence/cell in the post-nuclear fraction respectively, and 44 and 118 copies of each sequence/cell in the nuclear fraction respectively. The scarce post-nuclear sequences (18.5 copies of each sequence/cell) were also present at low levels in the nuclear fraction (0.1 copy of each sequence/cell). the results are discussed in terms of the post-transcriptional regulation of gene expression in the lactating guinea-pig mammary gland.

Animals↗
PubMed1989

The genome-linked protein and 5' end RNA sequence of plum pox potyvirus.

The infectivity of plum pox potyvirus (PPV) RNA was decreased by treatment with proteases. Ribonuclease digestion of iodinated PPV RNA yielded material which had an electrophoretic mobility corresponding to Mr 22,000. This protein presumably corresponds to the protease-sensitive structure needed for infectivity. A protein-linked RNase T1-resistant oligonucleotide, 38 nucleotides long, was sequenced and shown to correspond to the 5' terminus of the RNA by sequence comparison to the RNAs of two other potyviruses, tobacco etch virus and tobacco vein mottling virus. A 12 nucleotide block was found to be completely conserved in the RNAs of the three viruses.

Base Sequence↗