The intramolecular stem-loop structure of U6 snRNA can functionally replace the U6atac snRNA stem-loop

Shukla, Girish C.; Padgett, Richard A.

doi:10.1017/s1355838201000218

Cited by 29 publications

(20 citation statements)

References 29 publications

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“…The lack of stable U6-AGC base pairing to the U6-ISL in T. vaginalis might indicate folding instability of this structure. Nevertheless, our model supports the importance of the upper portion of this stem (McPheeters 1996;Shukla and Padgett 2001) as well as the formation of the U6 ISL pentaloop GNR(N)A, similar to that proposed in other organisms (Huppler et al 2002). Structure-function analyses of the T. vaginalis spliceosomal proteins and snRNAs will be necessary to test whether these structural differences impact splicing mechanisms in this divergent eukaryote.…”

Section: Discussionsupporting

confidence: 84%

Spliceosomal snRNAs in the unicellular eukaryote Trichomonas vaginalis are structurally conserved but lack a 5′-cap structure

Simões-Barbosa

Meloni²,

Wohlschlegel³

et al. 2008

RNA

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Few genes in the divergent eukaryote Trichomonas vaginalis have introns, despite the unusually large gene repertoire of this human-infective parasite. These introns are characterized by extended conserved regulatory motifs at the 59 and 39 boundaries, a feature shared with another divergent eukaryote, Giardia lamblia, but not with metazoan introns. This unusual characteristic of T. vaginalis introns led us to examine spliceosomal small nuclear RNAs (snRNAs) predicted to mediate splicing reactions via interaction with intron motifs. Here we identify T. vaginalis U1, U2, U4, U5, and U6 snRNAs, present predictions of their secondary structures, and provide evidence for interaction between the U2/U6 snRNA complex and a T. vaginalis intron. Structural models predict that T. vaginalis snRNAs contain conserved sequences and motifs similar to those found in other examined eukaryotes. These data indicate that mechanisms of intron recognition as well as coordination of the two catalytic steps of splicing have been conserved throughout eukaryotic evolution. Unexpectedly, we found that T. vaginalis spliceosomal snRNAs lack the 59 trimethylguanosine cap typical of snRNAs and appear to possess unmodified 59 ends. Despite the lack of a cap structure, U1, U2, U4, and U5 genes are transcribed by RNA polymerase II, whereas the U6 gene is transcribed by RNA polymerase III.

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Section: Discussionsupporting

confidence: 84%

Spliceosomal snRNAs in the unicellular eukaryote Trichomonas vaginalis are structurally conserved but lack a 5′-cap structure

Simões-Barbosa

Meloni²,

Wohlschlegel³

et al. 2008

RNA

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“…Some of this overlap is likely to be due to the presence of the U5 snRNP in both tri-snRNP complexes. In addition to the protein similarities, we have shown that an important RNA element of U6 snRNA can functionally replace the analogous element of U6atac snRNA (Shukla and Padgett 2001). More recently, we have also shown that U4atac snRNA can be functionally replaced by U4 snRNA providing that it can base pair with U6atac snRNA (Shukla and Padgett 2004).…”

Section: Introductionmentioning

confidence: 82%

“…Comparison of U6 and U6atac snRNAs showed that the central regions were either highly conserved or interchangeable based on our previous studies (Shukla and Padgett 2001). We therefore focused on the 59 and 39 ends of the RNAs, which are quite different (see Fig.…”

Section: Discussionmentioning

confidence: 99%

The conserved 3′ end domain of U6atac snRNA can direct U6 snRNA to the minor spliceosome

Dietrich¹,

Padgett²,

Shukla³

2009

RNA

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U6 and U6atac snRNAs play analogous critical roles in the major U2-dependent and minor U12-dependent spliceosomes, respectively. Previous results have shown that most of the functional cores of these two snRNAs are either highly similar in sequence or functionally interchangeable. Thus, a mechanism must exist to restrict each snRNA to its own spliceosome. Here we show that a chimeric U6 snRNA containing the unique and highly conserved 39 end domain of U6atac snRNA is able to function in vivo in U12-dependent spliceosomal splicing. Function of this chimera required the coexpression of a modified U4atac snRNA; U4 snRNA could not substitute. Partial deletions of this element in vivo, as well as in vitro antisense experiments, showed that the 39 end domain of U6atac snRNA is necessary to direct the U4atac/U6atac.U5 tri-snRNP to the forming U12-dependent spliceosome. In vitro experiments also uncovered a role for U4atac snRNA in this targeting.

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“…Interestingly, it could be shown that the intramolecular stem-loop structure of U6 snRNA can indeed functionally replace the U6atac snRNA stem-loop (32). Despite the fact that the U4/U6 and U4atac/U6atac snRNA duplexes exhibit analogous secondary structures, the overall sequence similarity between the corresponding snRNAs is not very high (approximately 40% between both U6 and U6atac and U4 and U4atac snRNAs) (35).…”

Section: Discussionmentioning

confidence: 99%

Human U4/U6.U5 and U4atac/U6atac.U5 Tri-snRNPs Exhibit Similar Protein Compositions

Schneider

Will

Makarova

et al. 2002

Molecular and Cellular Biology

111

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In the U12-dependent spliceosome, the U4atac/U6atac snRNP represents the functional analogue of the major U4/U6 snRNP. Little information is available presently regarding the protein composition of the former snRNP and its association with other snRNPs. In this report we show that human U4atac/U6atac di-snRNPs associate with U5 snRNPs to form a 25S U4atac/U6atac.U5 trimeric particle. Comparative analysis of minor and major tri-snRNPs by using immunoprecipitation experiments revealed that their protein compositions are very similar, if not identical. Not only U5-specific proteins but, surprisingly, all tested U4/U6-and major tri-snRNPspecific proteins were detected in the minor tri-snRNP complex. Significantly, the major tri-snRNP-specific proteins 65K and 110K, which are required for integration of the major tri-snRNP into the U2-dependent spliceosome, were among those proteins detected in the minor tri-snRNP, raising an interesting question as to how the specificity of addition of tri-snRNP to the corresponding spliceosome is maintained. Moreover, immunodepletion studies demonstrated that the U4/U6-specific 61K protein, which is involved in the formation of major tri-snRNPs, is essential for the association of the U4atac/U6atac di-snRNP with U5 to form the U4atac/U6atac.U5 tri-snRNP. Subsequent immunoprecipitation studies demonstrated that those proteins detected in the minor tri-snRNP complex are also incorporated into U12-dependent spliceosomes. This remarkable conservation of polypeptides between minor and major spliceosomes, coupled with the absence of significant sequence similarity between the functionally analogous snRNAs, supports an evolutionary model in which most major and minor spliceosomal proteins, but not snRNAs, are derived from a common ancestor.The major or U2-dependent spliceosome, which catalyzes the removal of the vast majority of introns from nuclear premRNA molecules, is a large 50S-60S ribonucleoprotein complex composed of the so-called major snRNPs U1, U2, U4/U6, and U5 and numerous non-snRNP-associated proteins (reviewed in references 4 and 27). It assembles on the pre-mRNA via an ordered pathway that involves dynamic interactions between the different snRNPs and also between snRNPs and the pre-mRNA (27,28,33). First, the U1 and the U2 snRNPs recognize the 5Ј splice site and the branch site, respectively, forming the prespliceosome. After the integration of the U4/U6 and U5 snRNPs as a preformed 25S U4/U6.U5 trisnRNP complex and subsequent rearrangements, a mature spliceosome is formed.More recently, a second spliceosome has been identified. This minor or U12-dependent spliceosome recognizes a distinct set of very rare nuclear pre-mRNA introns (so-called U12-type introns) that contain unique 5Ј splice site, branch site, and 3Ј splice site sequences (5, 11). The minor spliceosome contains a different set of snRNPs, namely, U11, U12, and U4atac/U6atac, but shares the U5 snRNP with the major spliceosome (12, 34-36). As shown both in vitro and in vivo, the U11, U12, and U4atac/U6atac snRNP...

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The intramolecular stem-loop structure of U6 snRNA can functionally replace the U6atac snRNA stem-loop

Cited by 29 publications

References 29 publications

Spliceosomal snRNAs in the unicellular eukaryote Trichomonas vaginalis are structurally conserved but lack a 5′-cap structure

Spliceosomal snRNAs in the unicellular eukaryote Trichomonas vaginalis are structurally conserved but lack a 5′-cap structure

The conserved 3′ end domain of U6atac snRNA can direct U6 snRNA to the minor spliceosome

Human U4/U6.U5 and U4atac/U6atac.U5 Tri-snRNPs Exhibit Similar Protein Compositions

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