Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Introns”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,495 records · Page 83Linked to original sources
PubMed1984

Antibodies against synthetic oligopeptides allow identification of the mRNA-maturase encoded by the second intron of the yeast cob-box gene.

Genetic and biochemical evidence has strongly suggested that several introns located in yeast mitochondrial genes specifying apocytochrome b or cytochrome oxidase encode trans-acting proteins (termed mRNA-maturases) responsible for splicing the cognate intron and maturation of the mRNA. We have chemically synthesized three oligopeptides, predicted from the DNA sequence of the open reading frame (ORF) present in the second intron of the cob-box gene, and raised antibodies against them. These antibodies have allowed us to identify a protein of 42 kd as the product translated from the ORF of the wild-type intron. In two splicing-deficient mutants this protein is replaced by shorter polypeptides whose lengths and antigenic properties are in full agreement with the positions of TAA codons established by the DNA sequence of the intron's ORF.

Antibodies, Fungal↗
PubMed1985

Role of the 5' hairpin structure in the splicing accuracy of the fourth intron of the yeast cob-box gene.

The splicing mechanism of the maturase-coding introns is poorly understood. We have systematically examined the phenotypes of a large number of revertants from the mitochondrial mutation G2457. This mutation results from a single base change near the 5' splicing site. We show here that this base change does not completely block the splicing of the intron but rather affects the specificity of the splicing process. We examine four classes of revertants which allow us to characterize the crucial role of a stem and loop structure in the accuracy of the intron excision process. An unexpected class of revertant suggests that other elements are involved in this mechanism. Reversion of G2457 can also occur via the excision in the mitochondrial genome of the intron coding sequence. These results are discussed in relation to the possible role fulfilled by the maturase in the control of intron splicing.

Animals↗
PubMed1985

Nucleotide sequence of the L1 ribosomal protein gene of Xenopus laevis: remarkable sequence homology among introns.

Ribosomal protein L1 is encoded by two genes in Xenopus laevis. The comparison of two cDNA sequences shows that the two L1 gene copies (L1a and L1b) have diverged in many silent sites and very few substitution sites; moreover a small duplication occurred at the very end of the coding region of the L1b gene which thus codes for a product five amino acids longer than that coded by L1a. Quantitatively the divergence between the two L1 genes confirms that a whole genome duplication took place in Xenopus laevis approximately 30 million years ago. A genomic fragment containing one of the two L1 gene copies (L1a), with its nine introns and flanking regions, has been completely sequenced. The 5' end of this gene has been mapped within a 20-pyridimine stretch as already found for other vertebrate ribosomal protein genes. Four of the nine introns have a 60-nucleotide sequence with 80% homology; within this region some boxes, one of which is 16 nucleotides long, are 100% homologous among the four introns. This feature of L1a gene introns is interesting since we have previously shown that the activity of this gene is regulated at a post-transcriptional level and it involves the block of the normal splicing of some intron sequences.

Animals↗
PubMed1986

Intron mutations that affect the splicing efficiency of the CYH2 gene of Saccharomyces cerevisiae.

To define the extent of intervening sequences required for efficient splicing of the CYH2 gene in Saccharomyces cerevisiae, we have constructed a series of intron mutations. Artificial intron extensions of more than 300 bp of the natural intron lead to an inhibition of splicing whereas intron deletions lead to a drastic improvement of the splicing efficiency. It is shown that deletion of a 32 bp sequence element within the intron is responsible for this drastic improvement.

Base Sequence↗
PubMed1981

Long range control circuits within mitochondria and between nucleus and mitochondria. II. Genetic and biochemical analyses of suppressors which selectively alleviate the mitochondrial intron mutations.

In the preceding paper of this series (Dujardin et al. 1980 a) we described general methods of selecting and genetically characterizing suppressor mutations that restore the respiratory capacity of mit- mitochondrial mutations. Two dominant nuclear (NAM1-1 and NAM2-1) and one mitochondrial (mim2-1) suppressors are more extensively studied in this paper. We have analysed the action spectrum of these suppressors on 433 mit- mutations located in various mitochondrial genes and found that they preferentially alleviate the effects of mutations located within intron open reading frames of the cob-box gene. We conclude that these suppressors permit the maturation of cytochrome b mRNA by restoring the synthesis of intron encoded protein(s) catalytically involved in splicing i.e. mRNA-maturase(s) (cf. Lazowska et al. 1980). NAM1-1 is allele specific and gene non-specific; it suppresses mutations located within different introns. NAM2-1 and mim2-1 are intron-specific: they suppress mutations all located in the same (box7) intron of the cob-box gene. Analyses of cytochrome absorption spectra and mitochondrial translation products of cells in which the suppressors are associated with various other mit- mutations show that the suppressors restore cytochrome b and/or cytochrome oxidase (cox I) synthesis, as expected from their growth phenotype. This suppression is, however, only partial: some new polypeptides characteristic of the mit- mutations can be still detected in the presence of suppressor. Interestingly enough when box7 specific suppressors NAM2-1 and mim2-1 are associated with a complete cob-box deletion (leading to a total deficiency of cytochrome b and oxidase) partial restoration of cox I synthesis is observed while cytochrome b is still totally absent. These results show that in strains carrying NAM2-1 or mim2-1 the presence of cytochrome b gene is no longer required for the expression of the oxi3 gene pointing out to the possibility of a mutational switch-on of silent genes, whether mitochondrial, mim2-1, or nuclear, NAM2-1. This switch-on would permit the synthesis of an active maturase acting as a substitute for the box7 maturase in order to splice the cytochrome b and oxidase mRNAs.

Base Sequence↗
PubMed2002

The sugar beet mitochondrial nad4 gene: an intron loss and its phylogenetic implication in the Caryophyllales.

The sugar beet mitochondrial gene for subunit IV of NADH dehydrogenase ( nad4) has been characterized. Unlike the corresponding genes in wheat and turnip, sugar beet nad4 lacks the second intron ( nad4-i2). Northern-blot analysis demonstrates transcription of the gene. A total of 19 RNA editing sites were identified in the sugar beet nad4 transcripts; interestingly, there is no editing in the region which flanks the lost intron. This observation is in favour of intron loss via homologous recombination of an edited RNA intermediate. We also found that the nad4-i2 intron is absent from the mitochondrial genomes of all examined members of the Caryophyllales, but present in the closely related orders, Polygonales and Plumbaginales, which suggests that the intron was lost in the common ancestor of the Caryophyllales.

Journal Article↗
PubMed1984

A model for the excision of introns 1 and 2 from adenoviral major late pre-messenger RNAs.

Total steady-state RNA was extracted from nuclei of HeLa cells late after infection with adenovirus serotype 2. Most of the nuclear RNA is transcribed from the major late transcription unit (16.2 to 100.0 map units). To study the cleavage reactions involved in the splicing of leaders 1 and 2, we have used the S1 nuclease mapping technique with restriction fragments located in the region of intron 1 as DNA probes. The S1 mapping data showed that in total nuclear RNA, RNA species accumulate from which the 5' part of intron 1 has been excised, but which still contain the 3' part of the intron. This indicates that intron 1 can be removed in a stepwise fashion following the 5' to 3' direction. We have compared the nucleotide sequences from the ends of the putative processing intermediates. The internal cleavage sites do not resemble the consensus 5' or 3' splice site sequences. However, they show considerable homology to the sequence 5' A-T-G-A-T-G-G-C-A-T 3', which may act as a signal for internal cleavage. The intermediates are present in both the poly(A)+ and poly(A)- RNA fractions, although with different relative intensities. Primer extension experiments have been performed in which a primer, located with its left end in leader 2, is extended into intron 1. The results show that there may be a cleavage site as short as 35 nucleotides before the 3' splice site. Cleavage at the 3' splice site seems to be rapidly followed by ligation of leader 1 to leader 2. A model for RNA splicing based on these findings and data from the literature is presented.

Adenoviruses, Human↗
PubMed1983

Evidence for the biochemical role of an internal sequence in yeast nuclear mRNA introns: implications for U1 RNA and metazoan mRNA splicing.

Sequence comparison of the introns of two yeast genes (rp51A and rp51B) coding for the same ribosomal protein shows homology only in the last 50 bases of the intron. This region of the intron contains an internal conserved sequence (ICS) present near the 3' end of all sequenced yeast nuclear mRNA introns. Removal of a 29 bp sequence containing the ICS prevents splicing of an intron-containing hybrid gene. In cells containing the wild-type gene, we have detected RNA molecules that we suggest are normal splicing intermediates, generated by an endonucleolytic cut in the primary transcript at the ICS. The homology of the ICS with a sequence near the 5' end of U1 snRNA suggests a model in which an interaction in cis between the ICS and the 5' splice junction in yeast is the counterpart of the interaction in trans between U1 and 5' splice junctions in higher eucaryotes.

Base Sequence↗
PubMed1983

Evidence for an intron-contained sequence required for the splicing of yeast RNA polymerase II transcripts.

We constructed numerous deletion mutants in the 3' region of the yeast actin gene intron. When these were introduced into yeast on autonomously replicating shuttle vectors, a sequence element between 35 and 70 nucleotides upstream from the 3' splice site was required for splicing the actin gene transcripts. In mutant genes where this intron region was maintained but new sequences introduced immediately downstream replaced the normal 3' acceptor site sequences, alternative splicing occurred. The alternative splicing signal was generally the first AG dinucleotide following the intron sequence required for splicing. The intron-contained sequence is sufficient to define new intron-exon boundaries. It contains the octanucleotide 5'-TACTAACA-3', which occurs 20 to 55 nucleotides upstream from the 3' splice site in all split protein-coding nuclear genes from S. cerevisiae sequenced to date, and we suggest that it is an essential element of the yeast splicing mechanism, involved in the selection of splicing sites.

Base Sequence↗
PubMed1984

RNA splicing in neurospora mitochondria: self-splicing of a mitochondrial intron in vitro.

We have used Neurospora nuclear mutant cyt-18-1, which accumulates a number of unspliced mitochondrial precursor RNAs, to identify rapidly mitochondrial introns that are self-splicing in vitro. Incubation of deproteinized whole mitochondrial RNA from the mutant with 32P-GTP resulted in strong labeling of a 1.3 kb RNA, subsequently identified as cytochrome b (cob) intron 1, and weaker labeling of additional RNAs. Self-splicing of cob intron 1, including precise cleavage and ligation, was confirmed using an in vitro transcript synthesized from the SP6 promoter. The in vitro splicing reaction was shown to be analogous to that for the Tetrahymena nuclear rRNA intron. Since splicing of cob intron 1 is inhibited in a recessive nuclear mutant, we infer that this essentially RNA-catalyzed splicing reaction must be facilitated by a protein in vivo.

Base Sequence↗
PubMed1985

The Tetrahymena rRNA intron self-splices in E. coli: in vivo evidence for the importance of key base-paired regions of RNA for RNA enzyme function.

We have developed an in vivo RNA splicing assay for the self-splicing rRNA intron of Tetrahymena thermophila using E. coli as the host. A DNA fragment containing the intron sequence has been cloned into M13mp83 so that expression of the beta-galactosidase alpha-fragment is dependent upon intron excision from the mRNA precursor. Plaque phenotypes correlate well with levels of excised intron RNA. Point mutations were made by oligonucleotide-directed mutagenesis in conserved sequences P, Q, and S. All showed reduced splicing, agreeing with mitochondrial genetic data for S and providing the first direct evidence that P and Q are functionally important. The results support the hypothesis that base-pairing of R with S and P with Q is important for intron structure and function.

Base Composition↗
PubMed1986

Protein-dependent splicing of a group I intron in ribonucleoprotein particles and soluble fractions.

The group I intron in the Neurospora mitochondrial large rRNA gene is not self-splicing in vitro. Here, we show that this intron can be spliced from 35S pre-rRNA in RNPs or from deproteinized 35S pre-rRNA or in vitro transcripts by a soluble activity that is present in mitochondrial lysates and can be released from RNPs. Splicing occurs by the same guanosine-initiated transesterification mechanism characteristic of self-splicing group I introns, but is absolutely dependent upon proteins that are presumably required for correct folding of the pre-rRNA. The soluble splicing activity is not simply associated with large subunit ribosomal proteins. Nuclear mutant cyt18-1, which is defective in splicing a number of group I introns in vivo, is grossly deficient in the soluble splicing activity. Our results suggest that the cyt18 gene encodes or regulates a component of an activity that functions in splicing group I introns in Neurospora mitochondria.

Base Sequence↗
PubMed1986

Characterization of the intron in the phage T4 thymidylate synthase gene and evidence for its self-excision from the primary transcript.

The td gene contains a 735 bp open reading frame within its 1017 bp intron. A 12 nucleotide stretch may form a stable secondary structure with the putative Shine-Dalgarno sequence of the intron open reading frame and thus impair its translation. SP6 RNA polymerase transcripts of the td gene synthesized in vitro at 40 degrees C encompass a 2.7 kb primary transcript, a 1.7 kb mRNA, and a 1 kb intron RNA. The excised intron RNA consisted of linear and cyclized forms. RNAase H studies and resistance of the cyclized intron to linearization by HeLa cell debranching enzyme suggest it to be circular. Self-splicing of isolated td primary transcript occurred only marginally at 28 degrees C, but increased progressively to 50 degrees C, and required the presence of both Mg++ and a guanosine cofactor. An internal guide sequence is evident which may align the 5' splice site with the 3' end, presumably for precise exon ligation.

Base Sequence↗
PubMed1986

The triosephosphate isomerase gene from maize: introns antedate the plant-animal divergence.

We have cloned and characterized a cDNA and genomic DNA for the triosephosphate isomerase expressed in maize roots. The gene is interrupted by eight introns. If we compare this gene with that for the protein in chicken, which has six introns, we see that five of the introns are at identical places, one has shifted by three codons, and two are totally new. This great matching leads us to conclude that the introns were in place before the plant-animal divergence, and that the parental gene had at least eight introns, two of which were lost in the line that leads to animals.

Animals↗
PubMed2005

Structural analysis of the group II intron splicing factor CRS2 yields insights into its protein and RNA interaction surfaces.

Chloroplast RNA splicing 2 (CRS2) is a nuclear-encoded protein required for the splicing of nine group II introns in maize chloroplasts. CRS2 functions in the context of splicing complexes that include one of two CRS2-associated factors (CAF1 and CAF2). The CRS2-CAF1 and CRS2-CAF2 complexes are required for the splicing of different subsets of CRS2-dependent introns, and they bind tightly and specifically to their genetically defined intron targets in vivo. The CRS2 amino acid sequence is closely related to those of bacterial peptidyl-tRNA hydrolases (PTHs). To identify the structural differences between CRS2 and bacterial PTHs responsible for CRS2's gains of CAF binding and intron splicing functions, we determined the structure of CRS2 by X-ray crystallography. The fold of CRS2 is the same as that of Escherichia coli PTH, but CRS2 has two surfaces that differ from the corresponding surfaces in PTH. One of these is more hydrophobic in CRS2 than in PTH. Site-directed mutagenesis of this surface blocked CRS2-CAF complex formation, indicating that it is the CAF binding site. The CRS2 surface corresponding to the putative tRNA binding face of PTH is considerably more basic than in PTH, suggesting that CRS2 interacts with group II intron substrates via this surface. Both the sequence and the structural context of the amino acid residues essential for peptidyl-tRNA hydrolase activity are conserved in CRS2, yet expression of CRS2 is incapable of rescuing a pth(ts)E.coli strain.

Amino Acid Sequence↗
PubMed2002

Pseudouridylation of U(35) in the anticodon of Arabidopsis thaliana pre-tRNA(Tyr) depends on length rather than structure of an intron.

In order to establish the structure and sequence requirements for pseudouridine (Psi(35)) biosynthesis in Arabidopsis thaliana tRNA(Tyr) five mutants of nuclear pre-tRNA(Tyr) have been prepared and analyzed: DeltaI-tRNA(Tyr) transcript depleted of an intron, and 5UI, 7UI, 9UI and 12UI transcripts containing tracts of five, seven, nine and 12 U residues, respectively, instead of the wild type tRNA(Tyr) intron. The in vitro transcripts were incubated in a lupin seed extract containing Psi(35) synthase activity, and those containing an artificial intron composed of 12 or nine U residues turned out to be good substrates for Psi(35) synthase. The transcript with an intron composed of seven uridine residues was pseudouridylated up to 40%, whereas the remaining two were not pseudouridylated at all. The secondary structures of all transcripts were determined using enzymatic and chemical probes: S(1), V(1), T(1), A, P(1) and Pb(2+). All mutant pre-tRNAs show similar structural features: their anticodon arm contains a five base pair stem and a large loop which consists of five natural tRNA(Tyr) AC loop nucleotides to which five, seven, nine and 12 U residues are added. As the structure of the wild type pre-tRNA(Tyr) is different we propose that the role of its intron in the process of U(35) pseudouridylation is simply to expand the anticodon region to the required critical length.

Anticodon↗
PubMedDate not supplied

Enzymatic activity of the conserved core of a group I self-splicing intron.

Splicing of the Tetrahymena ribosomal intron was first studied by Cech et al., who subsequently demonstrated that the intron RNA catalyses its own excision from a primary transcript to yield mature ribosomal RNA. This intron shares several short conserved sequences and a common secondary structure with several other introns, some of which have also been shown to self-splice. Here I show that the conserved core of the Tetrahymena intron can act in trans to catalyse the sequence-specific cleavage and addition of guanosine to a separate RNA substrate.

Animals↗
PubMedDate not supplied

Mechanism of recognition of the 5' splice site in self-splicing group I introns.

Group I introns include many mitochondrial ribosomal RNA and messenger RNA introns and the nuclear rRNA introns of Tetrahymena and Physarum. The splicing of precursor RNAs containing these introns is a two-step reaction. Cleavage at the 5' splice site precedes cleavage at the 3' splice site, the latter cleavage being coupled with exon ligation. Following the first cleavage, the 5' exon must somehow be held in place for ligation. We have now tested the reactivity of two self-splicing group I RNAs, the Tetrahymena pre-rRNA and the intron 1 portion of the Neurospora mitochondrial cytochrome b (cob) pre-mRNA, in the intermolecular exon ligation reaction (splicing in trans) described by Inoue et al. The different sequence specificity of the reactions supports the idea that the nucleotides immediately upstream from the 5' splice site are base-paired to an internal, 5' exon-binding site, in agreement with RNA structure models proposed by Davies and co-workers and others. The internal binding site is proposed to be involved in the formation of a structure that specifies the 5' splice site and, following the first step of splicing, to hold the 5' exon in place for exon ligation.

Animals↗