A computational scan for U12-dependent introns in the human genome sequence

Levine, Aaron; Durbin, Richard

doi:10.1093/nar/29.19.4006

Cited by 121 publications

(177 citation statements)

References 28 publications

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“…Thus, it is likely that some of the dendritically spliced sequences represent splicing errors rather than noncanonical or cryptic splice sites. Although there are little data on the major spliceosome error rate, the minor spliceosome error rate is estimated to be as high as 1 in 280 splice events (60). Data in Fig.…”

Section: Discussionmentioning

confidence: 98%

RNA splicing capability of live neuronal dendrites

Glanzer

Miyashiro

Sul

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

101

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Dendrites are specialized extensions of the neuronal soma that contain components of the cellular machinery involved in RNA and protein metabolism. Several dendritically localized proteins are associated with the precursor-mRNA (pre-mRNA) splicing complex, or spliceosome. Although some spliceosome-related, RNA-binding proteins are known to subserve separate cytoplasmic functions when moving between the nucleus and cytoplasm, little is known about the pre-mRNA splicing capacity of intact dendrites. Here, we demonstrate the presence and functionality of pre-mRNA-splicing components in dendrites. When isolated dendrites are transfected with a chicken ␦-crystallin pre-mRNA or luciferase reporter premRNA, splicing junctions clustered at or near expected splice sites are observed. Additionally, in vitro synaptoneurosome experiments show that this subcellular fraction contains a similar complement of splicing factors that is capable of splicing chicken ␦-crystallin pre-mRNA. These observations suggest that pre-mRNAsplicing factors found in the dendroplasm retain the potential to promote pre-mRNA splicing.T he molecular properties of dendrites have been extensively examined since the discovery of components of the protein synthetic machinery in the dendroplasm, including ribosomes and membranous constituents of the endoplasmic reticulum and Golgi apparatus (1, 2). A more detailed analysis of ribosomal particles suggests that some serine-arginine (SR) proteins associate with ribosomes in the cytoplasm and may be involved in the translational regulation of associated mRNAs (3). These SR proteins, as well as select heterogeneous nuclear ribonucleoproteins (hnRNPs) (4), are known to constitutively move between the nucleus and cytoplasm in nonneuronal cells where they have roles in posttranscriptional gene expression separate from their characteristic activities in nuclear precursor mRNA (pre-mRNA) splicing as core parts of the spliceosome. The spliceosome is a multimegadalton complex of ribonucleoproteins and small nuclear RNAs (snRNAs) that catalyzes the ATP-dependent removal of introns and ligation of exons in nuclear pre-mRNA (5, 6). Some auxiliary constituents of the spliceosome [e.g., SMN (7) and SAM68 (8)] are found throughout the neuronal cytoplasm, and their localization extends into the dendrites. Although it is clear that these cytoplasmic RNA-binding proteins (RBPs) have critical roles in the postsplicing regulation of the intracellular transport, stability, nonsense-mediated decay (NMD), and translation of cellular mRNAs (9, 10), it is not known whether they retain the ability to assemble pre-mRNA splicingcompetent complexes in the dendritic milieu. Unspliced or incompletely spliced pre-mRNAs are often sequestered in the nucleus, yet removal of introns by splicing, in many cases, is not essential for mRNA export from the nucleus (11-16). It is believed that some viral mRNAs and alternatively spliced mRNAs are likely exported to the cytosol as intron-retaining transcripts (4).To determine the functional capacity ...

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Section: Discussionmentioning

confidence: 98%

RNA splicing capability of live neuronal dendrites

Glanzer

Miyashiro

Sul

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

101

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“…A further significant feature of U12 splicing is the short distance between the branchpoint and 39 splice site and the lack of a polypyrimidine tract (Dietrich et al, 2001a;Levine and Durbin, 2001). In the LD U12 intron, the authentic 39 splice site lies 11 nucleotides downstream of the branchpoint.…”

Section: Plant U12 Splice Site Signalsmentioning

confidence: 99%

“…The most abundant class consists of U2-dependent introns (U2 introns), whereas the second rarer class (<0.4% of introns) consists of U12-dependent introns (U12 introns). U12 introns have been found in the nuclear genomes of vertebrates, plants, and insects (Hall and Padgett, 1994;Sharp and Burge, 1997;Tarn and Steitz, 1997;Krainer, 1998, 1999;Burge et al, 1998;Levine and Durbin, 2001;Patel and Steitz, 2003). Introns belonging to these two distinct classes are spliced by two different spliceosomes: the major U2-type spliceosome and the less abundant U12-type spliceosome (Hall and Padgett, 1996;Tarn and Steitz 1996a.…”

Section: Introductionmentioning

confidence: 99%

“…Introns belonging to these two distinct classes are spliced by two different spliceosomes: the major U2-type spliceosome and the less abundant U12-type spliceosome (Hall and Padgett, 1996;Tarn and Steitz 1996a. Although the first U12 introns to be described had AT-AC-terminal dinucleotides, the majority of U12-type introns contain GT-AG, and a small number contain other noncanonical terminal dinucleotides, such as AT-AA, AT-AG, AT-AT, GT-AT, or GT-GG (Jackson, 1991;Hall and Padgett, 1994;Dietrich et al, 1997Dietrich et al, , 2001aSharp and Burge, 1997;Burge et al, 1998;Wu and Krainer, 1999;Levine and Durbin, 2001;Zhu and Brendel, 2003). Moreover, functional analyses have shown that AT-AC-terminal dinucleotides are not a defining feature of U12 introns (Dietrich et al, 1997(Dietrich et al, , 2001a.…”

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confidence: 99%

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Determinants of Plant U12-Dependent Intron Splicing Efficiency

Lewandowska

Simpson²,

Clark³

et al. 2004

Plant Cell

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Factors affecting splicing of plant U12-dependent introns have been examined by extensive mutational analyses in an in vivo tobacco (Nicotiana tabacum) protoplast system using introns from three different Arabidopsis thaliana genes: CBP20, GSH2, and LD. The results provide evidence that splicing efficiency of plant U12 introns depends on a combination of factors, including UA content, exon bridging interactions between the U12 intron and flanking U2-dependent introns, and exon splicing enhancer sequences (ESEs). Unexpectedly, all three plant U12 introns required an adenosine at the upstream purine position in the branchpoint consensus UCCUURAUY. The exon upstream of the LD U12 intron is a major determinant of its higher level of splicing efficiency and potentially contains two ESE regions. These results suggest that in plants, U12 introns represent a level at which expression of their host genes can be regulated. INTRODUCTIONPre-mRNA splicing in eukaryotes is a fundamental step in gene expression and represents an important level at which the expression of protein-coding genes can be regulated (Reed, 2000;Smith and Valcá rcel, 2000;Graveley, 2001;Hastings and Krainer, 2001;Cartegni et al., 2002;Black, 2003). In higher eukaryotes, there are two classes of nuclear pre-mRNA introns. The most abundant class consists of U2-dependent introns (U2 introns), whereas the second rarer class (<0.4% of introns) consists of U12-dependent introns (U12 introns). U12 introns have been found in the nuclear genomes of vertebrates, plants, and insects (Hall and Padgett, 1994;Sharp and Burge, 1997;Tarn and Steitz, 1997; Krainer, 1998, 1999;Burge et al., 1998;Levine and Durbin, 2001;Patel and Steitz, 2003). Introns belonging to these two distinct classes are spliced by two different spliceosomes: the major U2-type spliceosome and the less abundant U12-type spliceosome (Hall and Padgett, 1996;Tarn and Steitz 1996a. Although the first U12 introns to be described had AT-AC-terminal dinucleotides, the majority of U12-type introns contain GT-AG, and a small number contain other noncanonical terminal dinucleotides, such as AT-AA, AT-AG, AT-AT, GT-AT, or GT-GG (Jackson, 1991;Hall and Padgett, 1994;Dietrich et al., 1997Dietrich et al., , 2001aSharp and Burge, 1997;Burge et al., 1998;Wu and Krainer, 1999;Levine and Durbin, 2001;Zhu and Brendel, 2003). Moreover, functional analyses have shown that AT-AC-terminal dinucleotides are not a defining feature of U12 introns (Dietrich et al., 1997(Dietrich et al., , 2001a. Instead, U12 introns contain highly conserved sequences in the 59 splice site (exon:G/ATATCCTY) and branchpoint region (TCCTTRAY) (Hall and Padgett, 1994;Sharp and Burge, 1997;Burge et al., 1998), which are both required for prespliceosome complex formation (Frilander and Steitz, 1999). The 39 splice site consensus sequence of U12 introns (YAC/G:exon) is less informative than their 59 splice site and branchpoint sequences. U12 introns also lack a polypyrimidine tract and have a short distance between the branchpoint and 39 splice si...

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“…The sequences upstream of the U12-type 5Јss in human, Arabidopsis, or Drosophila genes are not conserved (29,46,73), suggesting that cross-link formation between U12 and the P120 5Ј exon was facilitated in part by fortuitous sequence complementarity, which does not necessarily serve any essential role in intron recognition or splicing. This is supported by our in vitro splicing experiments in which splicing substrates containing mutations in the region binding to U12 snRNA did not show any significant differences in splicing activity as compared to control reactions (Frilander and Meng, unpublished; see Fig.…”

Section: Discussionmentioning

confidence: 99%

Proximity of the U12 snRNA with both the 5′ Splice Site and the Branch Point during Early Stages of Spliceosome Assembly

Frilander

Meng

2005

Molecular and Cellular Biology

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U12 snRNA is required for branch point recognition in the U12-dependent spliceosome. Using site-specific cross-linking, we have captured an unexpected interaction between the 5 end of the U12 snRNA and the ؊2 position upstream of the 5 splice site of P120 and SCN4a splicing substrates. The U12 snRNA nucleotides that contact the 5 exon are the same ones that form the catalytically important helix Ib with U6atac snRNA in the spliceosome catalytic core. However, the U12/5 exon interaction is transient, occurring prior to the entry of the U4atac/U6atac.U5 tri-snRNP to the spliceosome. This suggests that the helix Ib region of U12 snRNA is positioned near the 5 splice site early during spliceosome assembly and only later interacts with U6atac to form helix Ib. We also provide evidence that U12 snRNA can simultaneously interact with 5 exon sequences near 5 splice site and the branch point sequence, suggesting that the 5 splice site and branch point sequences are separated by <40 to 50 Å in the complex A of the U12-dependent spliceosome. Thus, no major rearrangements are subsequently needed to position these sites for the first step of catalysis.During pre-mRNA splicing, the 5Ј splice site (5Јss) and the branch point sequence (BPS), located at the opposite ends of each intron, must come into close proximity to facilitate the chemistry of the first catalytic step (4). In metazoan cells, two different types of introns, namely, the U2 type and U12 type, are recognized by their different 5Јss and BPS sequences. The U2-type introns comprise Ͼ99.5% of all spliceosomal introns and contain relatively poorly conserved 5Јss and BPS regions. In contrast, these sequences are highly conserved in the lowabundance U12-type introns (13, 47).Distinct U2-and U12-dependent spliceosomes facilitate the removal of the two different intron types. These two spliceosomes differ in their composition of small nuclear ribonucleoproteins (snRNPs), but their overall assembly pathways and catalytic mechanisms are similar. The U1, U2, U4, and U6 small nuclear RNAs (snRNAs) are specific to the U2-dependent spliceosome, whereas U11, U12, U4atac, and U6atac snRNAs are functionally analogous components in the U12-dependent spliceosome. Both spliceosomes utilize the U5 snRNA (42,56).A widely accepted paradigm of spliceosome assembly states that snRNPs and protein components associate with the premRNA in a stepwise manner (3,4,16). In the U2-dependent spliceosome, this process starts with intron recognition in complex E (the commitment complex in yeast), in which the 5Јss is recognized by the U1 snRNP, whereas the BPS, polypyrimidine tract (PPT), and 3Ј splice site (3Јss) are recognized by the protein factors SF1/BBP, U2AF65, and U2AF35, respectively (1,36,37,44,68,75). In the first ATP-dependent stage (complex A, or prespliceosome), SF1/BBP at the BPS is replaced by the U2 snRNP (43, 74). A spliceosome (complex B) is formed upon entry of the U4/U6.U5 tri-snRNP (20,24). Additional rearrangements of RNA-RNA interactions (39, 50), and changes in protein comp...

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A computational scan for U12-dependent introns in the human genome sequence

Cited by 121 publications

References 28 publications

RNA splicing capability of live neuronal dendrites

RNA splicing capability of live neuronal dendrites

Determinants of Plant U12-Dependent Intron Splicing Efficiency

Proximity of the U12 snRNA with both the 5′ Splice Site and the Branch Point during Early Stages of Spliceosome Assembly

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