Role of a conserved pseudouridine in U2 snRNA on the structural and electrostatic features of the spliceosomal pre-mRNA branch site

2005 ◽  
pp. 205-221
Author(s):  
Nancy L. Greenbaum
Keyword(s):  
U2 Snrna ◽  
Branch Site

Role of small nuclear RNAs in eukaryotic gene expression

2013 ◽  
Vol 54 ◽  
pp. 79-90 ◽  
Author(s):  
Saba Valadkhan ◽  
Lalith S. Gunawardane
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Stability ◽  
Splice Sites ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Eukaryotic Gene Expression ◽  
Eukaryotic Gene ◽  
Rna Biogenesis

Eukaryotic cells contain small, highly abundant, nuclear-localized non-coding RNAs [snRNAs (small nuclear RNAs)] which play important roles in splicing of introns from primary genomic transcripts. Through a combination of RNA–RNA and RNA–protein interactions, two of the snRNPs, U1 and U2, recognize the splice sites and the branch site of introns. A complex remodelling of RNA–RNA and protein-based interactions follows, resulting in the assembly of catalytically competent spliceosomes, in which the snRNAs and their bound proteins play central roles. This process involves formation of extensive base-pairing interactions between U2 and U6, U6 and the 5′ splice site, and U5 and the exonic sequences immediately adjacent to the 5′ and 3′ splice sites. Thus RNA–RNA interactions involving U2, U5 and U6 help position the reacting groups of the first and second steps of splicing. In addition, U6 is also thought to participate in formation of the spliceosomal active site. Furthermore, emerging evidence suggests additional roles for snRNAs in regulation of various aspects of RNA biogenesis, from transcription to polyadenylation and RNA stability. These snRNP-mediated regulatory roles probably serve to ensure the co-ordination of the different processes involved in biogenesis of RNAs and point to the central importance of snRNAs in eukaryotic gene expression.

An mRNA-type intron is present in the Rhodotorula hasegawae U2 small nuclear RNA gene

1993 ◽  
Vol 13 (9) ◽  
pp. 5613-5619
Author(s):  
Y Takahashi ◽  
S Urushiyama ◽  
T Tani ◽  
Y Ohshima
Keyword(s):  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
U2 Snrna ◽  
U6 Snrna ◽  
Branch Site ◽  
Snrna Gene ◽  
Nuclear Rna ◽  
Catalytic Role ◽  
Mrna Precursor ◽  
Site Recognition

Splicing an mRNA precursor requires multiple factors involving five small nuclear RNA (snRNA) species called U1, U2, U4, U5, and U6. The presence of mRNA-type introns in the U6 snRNA genes of some yeasts led to the hypothesis that U6 snRNA may play a catalytic role in pre-mRNA splicing and that the U6 introns occurred through reverse splicing of an intron from an mRNA precursor into a catalytic site of U6 snRNA. We characterized the U2 snRNA gene of the yeast Rhodotorula hasegawae, which has four mRNA-type introns in the U6 snRNA gene, and found an mRNA-type intron of 60 bp. The intron of the U2 snRNA gene is present in the highly conserved region immediately downstream of the branch site recognition domain. Interestingly, we found that this region can form a novel base pairing with U6 snRNA. We discuss the possible implications of these findings for the mechanisms of intron acquisition and for the role of U2 snRNA in pre-mRNA splicing.

scholarly journals Interactions of U2 Gene Loci and Their Nuclear Transcripts with Cajal (Coiled) Bodies: Evidence for PreU2 within Cajal Bodies

2000 ◽  
Vol 11 (9) ◽  
pp. 2987-2998 ◽  
Author(s):  
Kelly P. Smith ◽  
Jeanne Bentley Lawrence
Keyword(s):  
Potential Role ◽  
Coiled Body ◽  
Oligonucleotide Probes ◽  
Primary Transcript ◽  
U2 Snrna ◽  
Rna Foci ◽  
Coiled Bodies ◽  
Snrna Gene

The Cajal (coiled) body (CB) is a structure enriched in proteins involved in mRNA, rRNA, and snRNA metabolism. CBs have been shown to interact with specific histone and snRNA gene loci. To examine the potential role of CBs in U2 snRNA metabolism, we used a variety of genomic and oligonucleotide probes to visualize in situ newly synthesized U2 snRNA relative to U2 loci and CBs. Results demonstrate that long spacer sequences between U2 coding repeats are transcribed, supporting other recent evidence that U2 transcription proceeds past the 3′ box. The presence of bright foci of this U2 locus RNA differed between alleles within the same nucleus; however, this did not correlate with the loci's association with a CB. Experiments with specific oligonucleotide probes revealed signal for preU2 RNA within CBs. PreU2 was also detected in the locus-associated RNA foci, whereas sequences 3′ of preU2 were found only in these foci, not in CBs. This suggests that a longer primary transcript is processed before entry into CBs. Although this work shows that direct contact of a U2 locus with a CB is not simply correlated with RNA at that locus, it provides the first evidence of new preU2 transcripts within CBs. We also show that, in contrast to CBs, SMN gems do not associate with U2 gene loci and do not contain preU2. Because other evidence indicates that preU2 is processed in the cytoplasm before assembly into snRNPs, results point to an involvement of CBs in modification or transport of preU2 RNA.

scholarly journals Genomic divergence footprints in the bracovirus of Cotesia sesamiae identified by targeted re-sequencing approach

2017 ◽  
Author(s):  
Jérémy Gauthier ◽  
Philippe Gayral ◽  
Bruno Pierre Le Ru ◽  
Séverine Jancek ◽  
Stéphane Dupas ◽  
...  
Keyword(s):  
Host Range ◽  
Parasitoid Wasp ◽  
Gene Families ◽  
Genetic Variations ◽  
Genomic Region ◽  
Loss Of Function ◽  
Cotesia Sesamiae ◽  
Large Deletions ◽  
Branch Site

AbstractThe African parasitoid wasp Cotesia sesamiae is structured in contrasted populations showing differences in host range and the recent discovery of a specialist related species, C. typhae, provide a good framework to study the mechanisms that link the parasitoid and their host range. To investigate the genomic bases of divergence between these populations, we used a targeted sequencing approach on 24 samples. We targeted a specific genomic region encoding the bracovirus, which is deeply involved in the interaction with the host. High sequencing coverage was obtained for all samples allowing the study of genetic variations between wasp populations and species. Combining population genetic estimations, the diversity (π), the relative differentiation (FST) and the absolute differentiation (dxy), and branch-site dN/dS measures, we identified six divergent genes impacted by positive selection belonging to different gene families. These genes are potentially involved in host adaptation and in the specialization process. Fine scale analyses of the genetic variations also revealed deleterious mutations and large deletions on certain genes inducing pseudogenization and loss of function. These results highlight the crucial role of the bracovirus in the molecular interactions between the wasp and its hosts and in the evolutionary processes of specialization.

scholarly journals An mRNA-type intron is present in the Rhodotorula hasegawae U2 small nuclear RNA gene.

1993 ◽  
Vol 13 (9) ◽  
pp. 5613-5619 ◽  
Author(s):  
Y Takahashi ◽  
S Urushiyama ◽  
T Tani ◽  
Y Ohshima
Keyword(s):  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
U2 Snrna ◽  
U6 Snrna ◽  
Branch Site ◽  
Snrna Gene ◽  
Nuclear Rna ◽  
Catalytic Role ◽  
Mrna Precursor ◽  
Site Recognition

Splicing an mRNA precursor requires multiple factors involving five small nuclear RNA (snRNA) species called U1, U2, U4, U5, and U6. The presence of mRNA-type introns in the U6 snRNA genes of some yeasts led to the hypothesis that U6 snRNA may play a catalytic role in pre-mRNA splicing and that the U6 introns occurred through reverse splicing of an intron from an mRNA precursor into a catalytic site of U6 snRNA. We characterized the U2 snRNA gene of the yeast Rhodotorula hasegawae, which has four mRNA-type introns in the U6 snRNA gene, and found an mRNA-type intron of 60 bp. The intron of the U2 snRNA gene is present in the highly conserved region immediately downstream of the branch site recognition domain. Interestingly, we found that this region can form a novel base pairing with U6 snRNA. We discuss the possible implications of these findings for the mechanisms of intron acquisition and for the role of U2 snRNA in pre-mRNA splicing.

scholarly journals Transcription on lampbrush chromosome loops in the absence of U2 snRNA.

1992 ◽  
Vol 3 (3) ◽  
pp. 249-261 ◽  
Author(s):  
A Tsvetkov ◽  
M Jantsch ◽  
Z Wu ◽  
C Murphy ◽  
J G Gall
Keyword(s):  
Physiological Saline ◽  
Lampbrush Chromosome ◽  
Antisense Oligodeoxynucleotides ◽  
Lampbrush Chromosomes ◽  
U2 Snrna ◽  
Complete Destruction ◽  
Small Nuclear Rnas ◽  
Cytological Changes

The five small nuclear RNAs (snRNAs) involved in splicing occur on the loops of amphibian lampbrush chromosomes and in hundreds to thousands of extrachromosomal granules called B snurposomes. To assess the role of these snRNAs during transcription and to explore possible relationships between the loops and B snurposomes, we injected single-stranded antisense oligodeoxynucleotides (oligos) against U1 and U2 snRNA into toad and newt oocytes. As shown before, antisense U1 and U2 oligos caused truncation of U1 and complete destruction of U2 snRNAs, respectively. However, injection of any oligo, regardless of sequence, brought on dramatic cytological changes, including shortening of the chromosomes and retraction of the lateral loops, with concomitant shutdown of polymerase II transcription, as well as disappearance of some or all of the B snurposomes. When injected oocytes were incubated for 12 h or longer in physiological saline, these changes were reversible; that is, the chromosomes lengthened, transcription (detected by 3H-UTP incorporation) resumed on newly extended lateral loops, and B snurposomes reappeared. In situ hybridization showed that loops and B snurposomes had negligible amounts of U2 snRNA after recovery from injection of the anti-U2 oligo, whereas these structures had normal levels of U2 snRNA after recovery from a control oligo. Thus, the morphological integrity of B snurposomes and lampbrush chromosome loops is not dependent on the presence of U2 snRNA. Because transcription occurs in the absence of U2 snRNA, we conclude that splicing is not required for transcription on lampbrush chromosome loops.

scholarly journals A conserved pseudouridine modification in eukaryotic U2 snRNA induces a change in branch-site architecture

RNA ◽  
2001 ◽  
Vol 7 (6) ◽  
pp. 833-845 ◽  
Author(s):  
MEREDITH I. NEWBY ◽  
NANCY L. GREENBAUM
Keyword(s):  
U2 Snrna ◽  
Branch Site

scholarly journals Role of the 3′ Splice Site in U12-Dependent Intron Splicing

2001 ◽  
Vol 21 (6) ◽  
pp. 1942-1952 ◽  
Author(s):  
Rosemary C. Dietrich ◽  
Marian J. Peris ◽  
Andrew S. Seyboldt ◽  
Richard A. Padgett
Keyword(s):  
Splice Site ◽  
Site Selection ◽  
Intron Splicing ◽  
Splice Sites ◽  
Splice Site Selection ◽  
Branch Site ◽  
Site Function

ABSTRACT U12-dependent introns containing alterations of the 3′ splice site AC dinucleotide or alterations in the spacing between the branch site and the 3′ splice site were examined for their effects on splice site selection in vivo and in vitro. Using an intron with a 5′ splice site AU dinucleotide, any nucleotide could serve as the 3′-terminal nucleotide, although a C residue was most active, while a U residue was least active. The penultimate A residue, by contrast, was essential for 3′ splice site function. A branch site-to-3′ splice site spacing of less than 10 or more than 20 nucleotides strongly activated alternative 3′ splice sites. A strong preference for a spacing of about 12 nucleotides was observed. The combined in vivo and in vitro results suggest that the branch site is recognized in the absence of an active 3′ splice site but that formation of the prespliceosomal complex A requires an active 3′ splice site. Furthermore, the U12-type spliceosome appears to be unable to scan for a distal 3′ splice site.

scholarly journals U2 snRNA-Protein Contacts in Purified Human 17S U2 snRNPs and in Spliceosomal A and B Complexes

2006 ◽  
Vol 26 (7) ◽  
pp. 2803-2816 ◽  
Author(s):  
Olexandr Dybkov ◽  
Cindy L. Will ◽  
Jochen Deckert ◽  
Nastaran Behzadnia ◽  
Klaus Hartmuth ◽  
...  
Keyword(s):  
Branch Point ◽  
Protein Interactions ◽  
Cross Linking ◽  
Molecular Architecture ◽  
Point Selection ◽  
Stem Loop ◽  
U2 Snrna ◽  
Branch Site ◽  
Branch Point Sequence ◽  
U2 Snrnp

ABSTRACT The 17S U2 snRNP plays an essential role in branch point selection and catalysis during pre-mRNA splicing. Much remains to be learned about the molecular architecture of the U2 snRNP, including which proteins contact the functionally important 5′ end of the U2 snRNA. Here, RNA-protein interactions within immunoaffinity-purified human 17S U2 snRNPs were analyzed by lead(II)-induced RNA cleavage and UV cross-linking. Contacts between the U2 snRNA and SF3a60, SF3b49, SF3b14a/p14 and SmG and SmB were detected. SF3b49 appears to make multiple contacts, interacting with the 5′ end of U2 and nucleotides in loops I and IIb. SF3a60 also contacted different regions of the U2 snRNA, including the base of stem-loop I and a bulge in stem-loop III. Consistent with it contacting the pre-mRNA branch point adenosine, SF3b14a/p14 interacted with the U2 snRNA near the region that base pairs with the branch point sequence. A comparison of U2 cross-linking patterns obtained with 17S U2 snRNP versus purified spliceosomal A and B complexes revealed that RNA-protein interactions with stem-loop I and the branch site-interacting region of U2 are dynamic. These studies provide important insights into the molecular architecture of 17S U2 snRNPs and reveal U2 snRNP remodeling events during spliceosome assembly.

scholarly journals Recognition of the 5' splice site by the spliceosome.

1998 ◽  
Vol 45 (4) ◽  
pp. 869-881 ◽  
Author(s):  
M M Konarska
Keyword(s):  
Splice Site ◽  
Uv Light ◽  
Direct Interaction ◽  
Catalytic Center ◽  
U6 Snrna ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Definition Of ◽  
Self Splicing

The splicing of nuclear pre-mRNAs is catalyzed by a large, multicomponent ribonucleoprotein complex termed the spliceosome. Elucidation of the molecular mechanism of splicing identified small nuclear RNAs (snRNAs) as important components of the spliceosome, which, by analogy to the self-splicing group II introns, are implicated in formation of the catalytic center. In particular, the 5' splice site (5'SS) and the branch site, which represent the two substrates for the first step of splicing, are first recognized by U1 and U2 snRNPs, respectively. This initial recognition of splice sites is responsible for the global definition of exons and introns, and represents the primary target for regulation of splicing. Subsequently, pairing interaction between the 5'SS and U1 snRNA is disrupted and replaced by a new interaction of the 5'SS with U6 snRNA. The 5'SS signal contains an invariant GU dinucleotide present at the 5' end of nearly all known introns, however, the mechanism by which the spliceosome recognizes this element is not known. We have identified and characterized a specific UV light-induced crosslink formed between the 5'SS RNA and hPrp8, a protein component of U5 snRNP in the spliceosome that is likely to reflect a specific recognition of the GU dinucleotide for splicing. Because recognition of the 5'SS must be linked to formation of the catalytic site, the identification of a specific and direct interaction between the 5'SS and Prp8 has significant implications for the role of this protein in the mechanism of mRNA splicing.

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