Human Splicing Factor ASF/SF2 Encodes for a Repressor Domain Required for Its Inhibitory Activity on Pre-mRNA Splicing

Dauksaite, Vita; Akusjärvi, Göran

doi:10.1074/jbc.m107867200

Cited by 38 publications

(46 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…can also suppress splicing in a heterologous ␤-globin intron (21). Thus, splicing enhancers can suppress splicing when located within the introns of different splicing substrates, and we have shown that this suppression can be observed both in vitro and in microinjected Xenopus oocytes.…”

Section: Discussionmentioning

confidence: 76%

“…In previous studies (20,21) and in the results reported here, the activities of ESEs were shown to be context-specific. ESEs promote early steps of spliceosome assembly when located upstream or downstream of 5Ј or 3Ј splice sites, respectively, but they suppress splicing when located within the intron (4).…”

Section: Discussionmentioning

confidence: 92%

“…At the same time, negative intronic regulatory elements with which SR proteins interact have also been found (20,29,33,34). For example, a splicing suppressor element that binds to SR proteins, 3RE, has been identified in the adenovirus major late IIIa pre-mRNA (20,21). 3RE is located immediately adjacent to the BPS.…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Serine/arginine-rich protein-dependent suppression of exon skipping by exonic splicing enhancers

Ibrahim

Schaal

Hertel

et al. 2005

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

112

View full text Add to dashboard Cite

The 5 and 3 splice sites within an intron can, in principle, be joined to those within any other intron during pre-mRNA splicing. However, exons are joined in a strict 5 to 3 linear order in constitutively spliced pre-mRNAs. Thus, specific mechanisms must exist to prevent the random joining of exons. Here we report that insertion of exon sequences into an intron can inhibit splicing to the downstream 3 splice site and that this inhibition is independent of intron size. The exon sequences required for splicing inhibition were found to be exonic enhancer elements, and their inhibitory activity requires the binding of serine͞arginine-rich splicing factors. We conclude that exonic enhancers can act as barriers to prevent exon skipping and thereby may play a key role in ensuring the correct 5 to 3 linear order of exons in spliced mRNA.intronic splicing silencers ͉ pre-mRNA splicing ͉ accurate splice-site recognition and pairing M ost genes in higher eukaryotes contain multiple introns and exons that are transcribed to generate pre-mRNA. The production of mature mRNA requires the precise removal of introns and joining of exons by splicing. In constitutively spliced pre-mRNAs all of the introns are excised, and the exons are joined in the correct 5Ј to 3Ј order. This is a consequence of the fact that 5Ј splice sites are ligated to a 3Ј splice site exclusively within the same intron, thus avoiding the exclusion of exons (''exon skipping''). Two key problems in understanding the mechanisms of accurate pre-mRNA splicing are how the splicing machinery recognizes exons and how exons are joined in the correct 5Ј to 3Ј order (i.e., how exon skipping is avoided).Significant advances have been made in understanding the mechanisms involved in accurate exon recognition (1-3). This process requires weakly conserved sequence elements located at the 5Ј and 3Ј splice sites and at the branchpoint sequence (BPS) within introns. In addition, exons contain sequence elements known as exonic splicing enhancers (ESEs), which function as binding sites for members of the serine͞arginine-rich (SR) family of splicing factors. The bound SR proteins promote the use of flanking 5Ј and 3Ј splice sites through both protein-protein and protein-RNA interactions (4-6). Although splicing enhancer͞SR protein complexes provide a mechanism for exon recognition, the mechanisms by which exon skipping is avoided are not understood.An important insight into this problem was provided by the observation that many human genetic diseases are the consequence of single base mutations in ESEs and that these mutations cause exon skipping (7). Further insights were provided by studies of regulated alternative splicing, where it was shown that the inclusion of an alternate exon requires an ESE and the presence of specific SR proteins (8). Thus, exonic enhancer͞SR protein complexes are required to avoid exon skipping in both constitutive and alternative splicing. Although these observations indicate that exon recognition is a necessary step in avoiding exon skipping, it ...

show abstract

Section: Discussionmentioning

confidence: 76%

Section: Discussionmentioning

confidence: 92%

See 1 more Smart Citation

Serine/arginine-rich protein-dependent suppression of exon skipping by exonic splicing enhancers

Ibrahim

Schaal

Hertel

et al. 2005

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

112

View full text Add to dashboard Cite

show abstract

“…We have evidence that SF2͞ASF ⌬RS is modified by phosphorylation of Ser 129 within RRM2 (data not shown). The conserved heptapeptide motif is unlikely to be directly involved in RNA binding, given its position within the predicted structure of RRM2, but could have a function in protein-protein interactions (29). Fig.…”

Section: Rrm2 Is Required For the Function Of Sf2͞asf In Translationmentioning

confidence: 99%

“…RRM2 is characterized by the presence of a phylogenetically conserved heptapeptide, SWQDLKD, which is located in the first ␣-helix of this domain (28). It has been shown that mutations in this heptapeptide motif affect SF2͞ASF functions in pre-mRNA splicing (29) and its recruitment to nuclear stress granules (30). We took advantage of an RRM2 mutant of SF2͞ASF in which amino acid substitutions were introduced in this conserved heptapeptide, replacing the WQD residues by three alanines (Fig.…”

Section: Rrm2 Is Required For the Function Of Sf2͞asf In Translationmentioning

confidence: 99%

Reversible phosphorylation differentially affects nuclear and cytoplasmic functions of splicing factor 2/alternative splicing factor

Sanford

Ellis²,

Cazalla³

et al. 2005

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

119

128

View full text Add to dashboard Cite

The Ser͞Arg-rich (SR) proteins constitute a family of highly conserved nuclear phosphoproteins that are involved in many steps of mRNA metabolism. Previously, we demonstrated that shuttling SR proteins can associate with translating ribosomes and enhance translation of reporter mRNAs both in vivo and in vitro. Here, we show that endogenous, cytoplasmic splicing factor 2͞alternative splicing factor (SF2͞ASF) associated with the translation machinery is hypophosphorylated, suggesting that the phosphorylation state of the Arg-Ser-rich (RS) domain may influence the role of SF2͞ASF in cytoplasmic RNA processing. In agreement, we show that mutations mimicking a hypophosphorylated RS domain strongly increased SF2͞ASF binding to cytoplasmic mRNA and its activity in translation. We also demonstrate that, whereas the RS domain is not required for the function of SF2͞ASF in mRNA translation in vivo or in vitro, its second RNA recognition motif (RRM)2 plays a critical role in this process. Taken together, these data suggest that RS-domain phosphorylation may influence the association of SF2͞ASF with mRNA, whereas RRM2 may play an important role in mediating protein-protein interactions during translation. These data are consistent with a model whereby reversible protein phosphorylation differentially regulates the subcellular localization and activity of shuttling SR proteins.splicing factors ͉ translation ͉ subcellular localization ͉ RNA binding T he Ser͞Arg-rich (SR) proteins are a family of phylogenetically conserved, structurally related, splicing factors that have dual roles in pre-mRNA splicing affecting both constitutive and alternative splicing (1). SR family proteins have a modular structure consisting of one or two copies of an N-terminal RNA-recognition motif (RRM) followed by a C-terminal domain rich in alternating Ser and Arg residues, known as the Arg-Ser-rich (RS) domain. The RRMs determine RNA binding specificity, whereas the RS domain functions as a proteinprotein interaction module by recruiting components of the core splicing apparatus to promote splice site pairing (reviewed in ref.2).SR proteins are primarily localized to the nuclear speckles (reviewed in ref.3), and a subset of SR proteins shuttle continuously between the nucleus and the cytoplasm (4). Within the cell, the RS domain acts as a nuclear localization signal by mediating the interaction with the SR protein nuclear import receptor transportin-SR (5-7) and also influences the nucleocytoplasmic shuttling of individual SR proteins (4). The shuttling ability of a subset of SR proteins suggested additional roles in mRNA transport, and͞or in cytoplasmic events, such as mRNA localization, stability, or regulation of translation. In agreement with this hypothesis, two shuttling SR proteins, SRp20 and 9G8, have been shown to promote mRNA export of intronless RNAs (8) and also act as adapter proteins for TAP-dependent mRNA export (9). SR proteins have also been implicated in RNA stability and quality control. For instance, splicing factor 2͞al-ternati...

show abstract

SF2/ASF regulates proteomic diversity by affecting the balance between translation initiation mechanisms

Blaustein

Quadrana

Risso

et al. 2009

J of Cellular Biochemistry

View full text Add to dashboard Cite

Post-splicing activities have been described for a subset of shuttling serine/arginine-rich splicing regulatory proteins, among them SF2/ASF. We showed that growth factors activate a Ras-PI 3-kinase-Akt/PKB signaling pathway that not only modifies alternative splicing of the fibronectin EDA exon, but also alters in vivo translation of reporter mRNAs containing the EDA binding motif for SF2/ASF, providing two co-regulated levels of isoform-specific amplification. Translation of most eukaryotic mRNAs is initiated via the scanning mechanism, which implicates recognition of the m7G cap at the mRNA 5'-terminus by the eIF4F protein complex. Several viral and cellular mRNAs are translated in a cap-independent manner by the action of cis-acting mRNA elements named internal ribosome entry sites that direct internal ribosome binding to the mRNA. Here we use bicistronic reporters that generate mRNAs carrying two open reading frames, one translated in a cap-dependent manner while the other by internal ribosome entry site-dependent initiation, to show that in vivo over-expression of SF2/ASF increases the ratio between cap-dependent and internal ribosome entry site-dependent translation. Consistently, knocking-down of SF2/ASF causes the opposite effect. Changes in expression levels of SF2/ASF also affect alternative translation of an endogenous mRNA, that one coding for fibroblast growth factor-2. These results strongly suggest a role for SF2/ASF as a regulator of alternative translation, meaning the generation of different proteins by the balance among these two translation initiation mechanisms, and expand the known potential of SF2/ASF to regulate proteomic diversity to the translation field.

show abstract

Human Splicing Factor ASF/SF2 Encodes for a Repressor Domain Required for Its Inhibitory Activity on Pre-mRNA Splicing

Cited by 38 publications

References 39 publications

Serine/arginine-rich protein-dependent suppression of exon skipping by exonic splicing enhancers

Serine/arginine-rich protein-dependent suppression of exon skipping by exonic splicing enhancers

Reversible phosphorylation differentially affects nuclear and cytoplasmic functions of splicing factor 2/alternative splicing factor

SF2/ASF regulates proteomic diversity by affecting the balance between translation initiation mechanisms

Contact Info

Product

Resources

About