The determinants for Sm protein binding to Xenopus U1 and U5 snRNAs are complex and non-identical.

Jarmołowski, Artur; Mattaj, Iain W.

doi:10.1002/j.1460-2075.1993.tb05648.x

Cited by 65 publications

(70 citation statements)

References 39 publications

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“…The secondary structure, but not the sequence, of this stem loop is conserved. Interestingly, deletion of the terminal stem of U1 produced a stable RNA in Xenopus oocytes that was not Sm-precipitable (Jarmolowski and Mattaj 1993), whereas deletion of the terminal stem loop in HeLa extracts altered the sedimentation behavior of the resulting U1 particles (Patton et al 1987). Here, we observed that several phosphates within the terminal stem loop of U1 significantly affect RNP stability when changed to phosphorothioates.…”

Section: A Stem-loop Clamp For the Sm Ringmentioning

confidence: 71%

Assembly of the U1 snRNP involves interactions with the backbone of the terminal stem of U1 snRNA

McConnell¹,

Lokken²,

Steitz³

2003

RNA

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Nucleotide analog interference mapping (NAIM) is a powerful method for identifying RNA functional groups involved in protein-RNA interactions. We examined particles assembled on modified U1 small nuclear RNAs (snRNAs) in vitro and detected two categories of interferences. The first class affects the stability of two higher-order complexes and comprises changes in two adenosines, A65 and A70, in the loop region previously identified as the binding site for the U1 small nuclear ribonucleoprotein (snRNP)-specific U1A protein. Addition of an exocyclic amine to position 2 of A65 interferes strongly with protein binding, whereas removal or modification of the exocyclic amine at position 6 makes little difference. Modifications of A70 exhibit the opposite effects: Additions at position 2 are permitted, but modification of the exocyclic amine at position 6 significantly inhibits protein binding. These interactions, critical for U1A-U1 snRNA recognition in the context of in vitro snRNP assembly, are consistent with previous structural studies of the isolated protein with the RNA hairpin containing the U1A binding site. The second category of interferences affects all partially assembled U1-protein complexes by decreasing the stability of Sm core protein associations. Interestingly, most strong interferences occur at phosphates in the terminal stem-loop region of U1, rather than in the Sm binding site. These data argue that interactions with the phosphate backbone of the terminal stem loop are essential for the stable association of Sm core proteins with the U1 snRNA. We suggest that the stem loop of all Sm snRNAs may act as a clamp to hold the ring of Sm proteins in place.

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Section: A Stem-loop Clamp For the Sm Ringmentioning

confidence: 71%

Assembly of the U1 snRNP involves interactions with the backbone of the terminal stem of U1 snRNA

McConnell¹,

Lokken²,

Steitz³

2003

RNA

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“…In agreement with this finding, a subset of the U5 nucleotides implicated in the 5' splice site-U5 interaction have been cross-linked to exonic and intronic RNA sequences near the 5' splice site, indicating a close, physical juxtaposition of U5 and 5' splice site nucleotides during splicing (41, 44). Taken together, these studies suggest that the U5 snRNA plays an active role in the function of this snRNP.Aside from the few nucleotides implicated in the exon-U5 interaction, only the conserved Sm core protein binding site of U5 has a defined function (13,14,21 functional significance. An important first step towards this goal is the determination of U5 secondary structure, which is most effectively accomplished through phylogenetic analysis; this approach delineates structures that are presumed, by their evolutionary conservation, to be functionally significant (27).…”

mentioning

confidence: 83%

“…Aside from the few nucleotides implicated in the exon-U5 interaction, only the conserved Sm core protein binding site of U5 has a defined function (13,14,21 functional significance. An important first step towards this goal is the determination of U5 secondary structure, which is most effectively accomplished through phylogenetic analysis; this approach delineates structures that are presumed, by their evolutionary conservation, to be functionally significant (27).…”

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confidence: 99%

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Architecture of the U5 small nuclear RNA.

Frank¹,

Roiha²,

Guthrie³

1994

Mol. Cell. Biol.

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We have used comparative sequence analysis and deletion analysis to examine the secondary structure of the U5 small nuclear RNA (snRNA), an essential component of the pre-mRNA splicing apparatus. The secondary structure of Saccharomyces cerevisiae U5 snRNA was studied in detail, while sequences from six other fungal species were included in the phylogenetic analysis. Our results indicate that fungal U5 snRNAs, like their counterparts from other taxa, can be folded into a secondary structure characterized by a highly conserved stem-loop (stem-loop 1) that is flanked by a moderately conserved internal loop (internal loop 1). In addition, several of the fungal U5 snRNAs include a novel stem-loop structure (ca. 30 nucleotides) that is adjacent to stem-loop 1. By deletion analysis of the S. cerevisiae snRNA, we have demonstrated that the minimal U5 snRNA that can complement the lethal phenotype of a U5 gene disruption consists of (i) stem-loop 1, (ii) internal loop 1, (iii) a stem-closing internal loop 1, and (iv) the conserved Sm protein binding site. Remarkably, all essential, U5-specific primary sequence elements are encoded by a 39-nucleotide domain consisting of stem-loop 1 and internal loop 1. This domain must, therefore, contain all U5-specific sequences that are essential for splicing activity, including binding sites for U5-specific proteins.Although the U5 small nuclear ribonucleoprotein (snRNP) is an essential cofactor in pre-mRNA splicing, its precise function is unknown. The U5 snRNP has long been hypothesized to play a role in the second catalytic step of splicing, perhaps as a factor required for the recognition of the 3' splice site (6,28,38). Indeed, we have recently identified several proteins that genetically interact with the U5 small nuclear RNA (snRNA) and are required for the second step of splicing (8). One of these proteins, the product of the SLU7 gene, functions in the selection of distant 3' splice sites (7). Whether any of these proteins is an integral U5 snRNP protein remains an open question; nevertheless, they provide strong evidence linking U5 function to the recognition and utilization of the 3' splice site. Moreover, several studies indicate that conserved sequences in the U5 snRNA, itself, can interact with exon sequences at the intron-exon junctions. Newman and Norman (25) have demonstrated that mutations in the Saccharomyces cerevisiae U5 snRNA can suppress mutations in both 5' and 3' splice sites, suggesting that the U5 snRNA can form hydrogen bonds with both 5' and 3' exon sequences. In agreement with this finding, a subset of the U5 nucleotides implicated in the 5' splice site-U5 interaction have been cross-linked to exonic and intronic RNA sequences near the 5' splice site, indicating a close, physical juxtaposition of U5 and 5' splice site nucleotides during splicing (41, 44). Taken together, these studies suggest that the U5 snRNA plays an active role in the function of this snRNP.Aside from the few nucleotides implicated in the exon-U5 interaction, only the conserved Sm ...

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“…Chemical modification studies of nematode SL RNA indicate a functional role in trans-splicing for purines in the upstream but not the downstream stem-loop (Hannon et al 1992), however, the SL RNA of Schistosoma does not contain a similar upstream stem-loop (Rajkovic et al 1990). Stem-loops may quantitatively facilitate Sm protein binding to U snRNAs (Jarmalowski and Mattaj 1993;Hinz et al 1996) but they are not essential (Raker et al 1999). As Figure 6 shows, the 46-nucleotide Ciona SL RNA is more like the relatively simple SL RNA of Schistosoma than the more complex classical SL RNA secondary structure represented by Caenorhabditis SL1.…”

Section: Trans-splicing Of Tni/␤-gal Chimeric Transcriptsmentioning

confidence: 99%

mRNA 5′-leader trans-splicing in the chordates

Vandenberghe¹,

Meedel²,

Hastings³

2001

Genes Dev.

138

133

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We report the discovery of mRNA 5-leader trans-splicing (SL trans-splicing) in the chordates. In the ascidian protochordate Ciona intestinalis, the mRNAs of at least seven genes undergo trans-splicing of a 16-nucleotide 5-leader apparently derived from a 46-nucleotide RNA that shares features with previously characterized splice donor SL RNAs. SL trans-splicing was known previously to occur in several protist and metazoan phyla, however, this is the first report of SL trans-splicing within the deuterostome division of the metazoa. SL trans-splicing is not known to occur in the vertebrates. However, because ascidians are primitive chordates related to vertebrate ancestors, our findings raise the possibility of ancestral SL trans-splicing in the vertebrate lineage.[ mRNA 5Ј-leader trans-splicing is a mode of gene expression reported in several organisms in which the original 5Ј ends of pre-mRNAs are discarded and are replaced by the 5Ј segment of a spliced leader (SL) RNA (Bonen 1993;Blumenthal 1995;Davis 1996). The function of SL transsplicing is not clear in every case and may vary. Multiple roles have been proposed, including mediation of mRNA stability or translatability (Maroney et al. 1995), resolution of polycistronic pre-mRNAs (Agabian 1990;Blumenthal 1995), and production of functional mRNAs from RNA polymerase I transcripts (Lee and Van der Ploeg 1997). In some organisms, only a subset of mRNAs undergo SL trans-splicing but in others, most or all do (Agabian 1990;Bonen 1993;Davis 1996). SL trans-splicing occurs alongside of the conventional cis-splicing process that removes introns from pre-mRNAs (Bonen 1993;Blumenthal 1995;Mair et al. 2000). There are mechanistic parallels between SL trans-splicing and conventional cis-splicing, including the use of the same set of nucleotide sequence features to mark splice donor and acceptor sites, and a strong resemblance of SL RNAs to spliceosomal U snRNAs (Agabian 1990;Bonen 1993;Nilsen 1993). These similarities imply a close evolutionary relationship between cis-splicing and SL trans-splicing, but the nature of this relationship and the overall evolutionary history of SL trans-splicing are not clear, in part because the phylogenetic distribution of SL transsplicing has not been clearly delineated.The known phylogenetic distribution of SL transsplicing is uneven and includes several protist and metazoan groups (Bonen 1993;Blumenthal 1995;Davis 1996). It was first discovered in a protist group, the trypanosomes (Campbell et al. 1984;Kooter et al. 1984;Milhausen et al. 1984), then subsequently in two protosotome metazoan phyla, Nematoda (Krause and Hirsh 1987) and Platyhelminthes (flatworms) (Rajkovic et al. 1990) and in Euglena, a protist distantly related to trypanosomes (Tessier et al. 1991). SL trans-splicing has not been reported in advanced protostome phyla, that is, the arthropods, annelids, or molluscs, nor among the deuterostomes, the great division of the metazoa that includes chordates/vertebrates. However, because each discovery of SL trans-splicing was a...

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The determinants for Sm protein binding to Xenopus U1 and U5 snRNAs are complex and non-identical.

Cited by 65 publications

References 39 publications

Assembly of the U1 snRNP involves interactions with the backbone of the terminal stem of U1 snRNA

Assembly of the U1 snRNP involves interactions with the backbone of the terminal stem of U1 snRNA

Architecture of the U5 small nuclear RNA.

mRNA 5′-leader trans-splicing in the chordates

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