An intact histone 3'-processing site is required for transcription termination in a mouse histone H2a gene

1991 ◽  
Vol 11 (1) ◽  
pp. 497-509
Author(s):  
N Chodchoy ◽  
N B Pandey ◽  
W F Marzluff
Keyword(s):  
Transcription Termination ◽  
Histone H2a ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Histone 3 ◽  
Transcription Termination Site ◽  
Processing Signal ◽  
Processing Site ◽  
Polyadenylation Sites ◽  
Nuclear Ribonucleoprotein

A transcription termination site has been characterized between the mouse histone H2a-614 and H3-614 genes. There is a poly(A)- RNA present in small amounts in the nucleus which ends 600 nucleotides 3' to the H2a-614 gene. Nuclear transcription studies demonstrate that transcription extends at least 600 nucleotides 3' to the gene but is greatly reduced 700 nucleotides 3' to the gene. If all or part of the normal 3'-processing signal, consisting of the stem-loop and the U7 small nuclear ribonucleoprotein binding site, is deleted, transcription then continues past the putative termination site and RNAs which end at the 3' end of the downstream H3-614 gene accumulate. Insertion of a 150-nucleotide fragment containing the termination site between the histone 3' end and downstream polyadenylation sites reduces usage of polyadenylation sites 85 to 90%. Taken together these results suggest there is a transcription termination site which requires an intact histone 3'-processing signal to function.

scholarly journals An intact histone 3'-processing site is required for transcription termination in a mouse histone H2a gene.

1991 ◽  
Vol 11 (1) ◽  
pp. 497-509 ◽  
Author(s):  
N Chodchoy ◽  
N B Pandey ◽  
W F Marzluff
Keyword(s):  
Transcription Termination ◽  
Histone H2a ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Histone 3 ◽  
Transcription Termination Site ◽  
Processing Signal ◽  
Processing Site ◽  
Polyadenylation Sites ◽  
Nuclear Ribonucleoprotein

A transcription termination site has been characterized between the mouse histone H2a-614 and H3-614 genes. There is a poly(A)- RNA present in small amounts in the nucleus which ends 600 nucleotides 3' to the H2a-614 gene. Nuclear transcription studies demonstrate that transcription extends at least 600 nucleotides 3' to the gene but is greatly reduced 700 nucleotides 3' to the gene. If all or part of the normal 3'-processing signal, consisting of the stem-loop and the U7 small nuclear ribonucleoprotein binding site, is deleted, transcription then continues past the putative termination site and RNAs which end at the 3' end of the downstream H3-614 gene accumulate. Insertion of a 150-nucleotide fragment containing the termination site between the histone 3' end and downstream polyadenylation sites reduces usage of polyadenylation sites 85 to 90%. Taken together these results suggest there is a transcription termination site which requires an intact histone 3'-processing signal to function.

scholarly journals Interactions between highly conserved U2 small nuclear RNA structures and Prp5p, Prp9p, Prp11p, and Prp21p proteins are required to ensure integrity of the U2 small nuclear ribonucleoprotein in Saccharomyces cerevisiae.

1994 ◽  
Vol 14 (9) ◽  
pp. 6337-6349 ◽  
Author(s):  
S E Wells ◽  
M Ares
Keyword(s):  
Synthetic Lethality ◽  
Small Nuclear Rna ◽  
Rna Structures ◽  
Stem Loop ◽  
U2 Snrna ◽  
Small Nuclear Ribonucleoprotein ◽  
U2 Snrnp ◽  
Nuclear Rna ◽  
Nuclear Ribonucleoprotein

Binding of U2 small nuclear ribonucleoprotein (snRNP) to the pre-mRNA is an early and important step in spliceosome assembly. We searched for evidence of cooperative function between yeast U2 small nuclear RNA (snRNA) and several genetically identified splicing (Prp) proteins required for the first chemical step of splicing, using the phenotype of synthetic lethality. We constructed yeast strains with pairwise combinations of 28 different U2 alleles with 10 prp mutations and found lethal double-mutant combinations with prp5, -9, -11, and -21 but not with prp3, -4, -8, or -19. Many U2 mutations in highly conserved or invariant RNA structures show no phenotype in a wild-type PRP background but render mutant prp strains inviable, suggesting that the conserved but dispensable U2 elements are essential for efficient cooperative function with specific Prp proteins. Mutant U2 snRNA fails to accumulate in synthetic lethal strains, demonstrating that interaction between U2 RNA and these four Prp proteins contributes to U2 snRNP assembly or stability. Three of the proteins (Prp9p, Prp11p, and Prp21p) are associated with each other and pre-mRNA in U2-dependent splicing complexes in vitro and bind specifically to synthetic U2 snRNA added to crude splicing extracts depleted of endogenous U2 snRNPs. Taken together, the results suggest that Prp9p, -11p, and -21p are U2 snRNP proteins that interact with a structured region including U2 stem loop IIa and mediate the association of the U2 snRNP with pre-mRNA.

scholarly journals Splicing-independent recruitment of spliceosomal small nuclear RNPs to nascent RNA polymerase II transcripts

2007 ◽  
Vol 178 (6) ◽  
pp. 937-949 ◽  
Author(s):  
Snehal Bhikhu Patel ◽  
Natalya Novikova ◽  
Michel Bellini
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Messenger Rnas ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Small Nuclear Rnas ◽  
Transcriptional Units ◽  
Necessary And Sufficient ◽  
Nuclear Ribonucleoprotein ◽  
Nascent Transcripts

In amphibian oocytes, most lateral loops of the lampbrush chromosomes correspond to active transcriptional sites for RNA polymerase II. We show that newly assembled small nuclear ribonucleoprotein (RNP [snRNP]) particles, which are formed upon cytoplasmic injection of fluorescently labeled spliceosomal small nuclear RNAs (snRNAs), target the nascent transcripts of the chromosomal loops. With this new targeting assay, we demonstrate that nonfunctional forms of U1 and U2 snRNAs still associate with the active transcriptional units. In particular, we find that their association with nascent RNP fibrils is independent of their base pairing with pre–messenger RNAs. Additionally, stem loop I of the U1 snRNA is identified as a discrete domain that is both necessary and sufficient for association with nascent transcripts. Finally, in oocytes deficient in splicing, the recruitment of U1, U4, and U5 snRNPs to transcriptional units is not affected. Collectively, these data indicate that the recruitment of snRNPs to nascent transcripts and the assembly of the spliceosome are uncoupled events.

Model of the interaction between the UI A small nuclear ribonucleoprotein and UI small nuclear RNA stem/loop II

1993 ◽  
Vol 21 (3) ◽  
pp. 605-609 ◽  
Author(s):  
P. R. Evans ◽  
C. Oubridge ◽  
T.-H. Jessen ◽  
J. Li ◽  
C. H. Teo ◽  
...  
Keyword(s):  
Small Nuclear Rna ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Nuclear Rna ◽  
Nuclear Ribonucleoprotein

scholarly journals A specific 31-nucleotide domain of U1 RNA directly interacts with the 70K small nuclear ribonucleoprotein component.

1989 ◽  
Vol 9 (11) ◽  
pp. 4872-4881 ◽  
Author(s):  
C C Query ◽  
R C Bentley ◽  
J D Keene
Keyword(s):  
Nucleotide Sequence ◽  
Splice Site ◽  
Rna Binding ◽  
Binding Domain ◽  
Specific Domain ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Associated Proteins ◽  
Nuclear Ribonucleoprotein

We have defined the nucleotide sequence of a protein-binding domain within U1 RNA that specifically recognizes and binds both to a U1 small nuclear ribonucleoprotein component (the 70K protein) and to the previously defined RNA-binding domain of the 70K protein. We have investigated direct interactions between purified U1 RNA and 70K protein by reconstitution in vitro. Thirty-one nucleotides of U1 RNA, corresponding to stem-loop I, were required for this interaction. Nucleotides at the 5' end of U1 RNA that are involved in base pairing with the 5' splice site of pre-mRNA were not required for binding. In contrast to other reports, these findings demonstrate that a specific domain of U1 RNA can bind directly to the 70K protein independently of any other snRNP-associated proteins.

scholarly journals Mutations in an essential U2 small nuclear RNA structure cause cold-sensitive U2 small nuclear ribonucleoprotein function by favoring competing alternative U2 RNA structures.

1994 ◽  
Vol 14 (3) ◽  
pp. 1689-1697 ◽  
Author(s):  
M I Zavanelli ◽  
J S Britton ◽  
A H Igel ◽  
M Ares
Keyword(s):  
Rna Structure ◽  
Cold Sensitivity ◽  
Small Nuclear Rna ◽  
Stem Loop ◽  
Alternative Structures ◽  
Small Nuclear Ribonucleoprotein ◽  
Alternative Conformation ◽  
Cold Sensitive ◽  
The Stability ◽  
Nuclear Ribonucleoprotein

Mutations in stem-loop IIa of yeast U2 RNA cause cold-sensitive growth and cold-sensitive U2 small nuclear ribonucleoprotein function in vitro. Cold-sensitive U2 small nuclear RNA adopts an alternative conformation that occludes the loop and disrupts the stem but does so at both restrictive and permissive temperatures. To determine whether alternative U2 RNA structure causes the defects, we tested second-site mutations in U2 predicted to disrupt the alternative conformation. We find that such mutations efficiently suppress the cold-sensitive phenotypes and partially restore correct U2 RNA folding. A genetic search for additional suppressors of cold sensitivity revealed two unexpected mutations in the base of an adjacent stem-loop. Direct probing of RNA structure in vivo indicates that the suppressors of cold sensitivity act to improve the stability of the essential stem relative to competing alternative structures by disrupting the alternative structures. We suggest that many of the numerous cold-sensitive mutations in a variety of RNAs and RNA-binding proteins could be a result of changes in the stability of a functional RNA conformation relative to a competing structure. The presence of an evolutionarily conserved U2 sequence positioned to form an alternative structure argues that this region of U2 is dynamic during the assembly or function of the U2 small nuclear ribonucleoprotein.

scholarly journals The U1 RNA-binding site of the U1 small nuclear ribonucleoprotein (snRNP)-associated A protein suggests a similarity with U2 snRNPs.

1989 ◽  
Vol 9 (7) ◽  
pp. 2975-2982 ◽  
Author(s):  
C Lutz-Freyermuth ◽  
J D Keene ◽  
C Lutz-Reyermuth
Keyword(s):  
Binding Site ◽  
Rna Binding ◽  
Sequence Similarity ◽  
Binding Assays ◽  
Stem Loop ◽  
Small Nuclear Ribonucleoprotein ◽  
Vitro Binding ◽  
Associated Proteins ◽  
Nuclear Ribonucleoprotein

The site of interaction between human U1 RNA and one of its uniquely associated proteins, A, was examined with in vitro binding assays. The A protein bound directly to stem-loop II of U1 RNA in a region which exhibits sequence similarity to U2 RNA. The similarity with U2 RNA was in a region that has been shown to interact with a U2 RNA-associated protein. The A protein-binding site on U1 RNA overlapped a previously described epitope for an RNA-specific human autoantibody (S. L. Deutscher and J. D. Keene, Proc. Natl. Acad. Sci. USA 85:3299-3303, 1988), supporting the hypothesis that the anti-RNA antibody originated as an anti-idiotypic response to A protein-specific autoantibodies.

scholarly journals Mutations in an essential U2 small nuclear RNA structure cause cold-sensitive U2 small nuclear ribonucleoprotein function by favoring competing alternative U2 RNA structures

1994 ◽  
Vol 14 (3) ◽  
pp. 1689-1697
Author(s):  
M I Zavanelli ◽  
J S Britton ◽  
A H Igel ◽  
M Ares
Keyword(s):  
Rna Structure ◽  
Cold Sensitivity ◽  
Small Nuclear Rna ◽  
Stem Loop ◽  
Alternative Structures ◽  
Small Nuclear Ribonucleoprotein ◽  
Alternative Conformation ◽  
Cold Sensitive ◽  
The Stability ◽  
Nuclear Ribonucleoprotein

Mutations in stem-loop IIa of yeast U2 RNA cause cold-sensitive growth and cold-sensitive U2 small nuclear ribonucleoprotein function in vitro. Cold-sensitive U2 small nuclear RNA adopts an alternative conformation that occludes the loop and disrupts the stem but does so at both restrictive and permissive temperatures. To determine whether alternative U2 RNA structure causes the defects, we tested second-site mutations in U2 predicted to disrupt the alternative conformation. We find that such mutations efficiently suppress the cold-sensitive phenotypes and partially restore correct U2 RNA folding. A genetic search for additional suppressors of cold sensitivity revealed two unexpected mutations in the base of an adjacent stem-loop. Direct probing of RNA structure in vivo indicates that the suppressors of cold sensitivity act to improve the stability of the essential stem relative to competing alternative structures by disrupting the alternative structures. We suggest that many of the numerous cold-sensitive mutations in a variety of RNAs and RNA-binding proteins could be a result of changes in the stability of a functional RNA conformation relative to a competing structure. The presence of an evolutionarily conserved U2 sequence positioned to form an alternative structure argues that this region of U2 is dynamic during the assembly or function of the U2 small nuclear ribonucleoprotein.

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