Phosphorylation-Dependent Control of the Pre-mRNA Splicing Machinery

Soret, Johann; Tazi, Jamal

doi:10.1007/978-3-662-09728-1_4

Cited by 31 publications

(26 citation statements)

References 265 publications

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“…Because SF3a p120 is known to be a component of U2 small nuclear ribonucleoprotein particle, which forms the spliceosome along with the U1 complex (24, 25), we used coimmunoprecipitation to determine whether phosphorylated hER␣ associated with the spliceosome via SF3a p120. Coimmunoprecipitation experiments were performed with extracts from hER␣-expressing MCF7 cells, and results indicated that all U1͞U2 endogenous components tested (U2: SF3a p66, SF3a p60 small nuclear ribonucleoprotein particles; and U1: U1 70K small nuclear ribonucleoprotein particles) (27,28) were coimmunoprecipitated with endogenous hER␣, with these associations potentiated by EGF treatment (Fig. 2B, lanes 1-4).…”

Section: Phosphorylated Her␣ Associates With the Spliceosome Through mentioning

confidence: 94%

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Masuhiro

Mezaki²,

Sakari³

et al. 2005

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

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Mitogen-activated protein kinase-mediated growth factor signals are known to augment the ligand-induced transactivation function of nuclear estrogen receptor ␣ (ER␣) through phosphorylation of Ser-118 within the ER␣ N-terminal transactivation (activation function-1) domain. We identified the spliceosome component splicing factor (SF)3a p120 as a coactivator specific for human ER␣ (hER␣) activation function-1 that physically associated with ER␣ dependent on the phosphorylation state of Ser-118. SF3a p120 potentiated hER␣-mediated RNA splicing, and notably, the potentiation of RNA splicing by SF3a p120 depended on hER Ser-118 phosphorylation. Thus, our findings suggest a mechanism by which growth factor signaling can regulate gene expression through the modulation of RNA splicing efficiency via phosphorylation of sequencespecific activators, after association between such activators and the spliceosome.nuclear receptor ͉ estrogen ͉ coactivator ͉ mitogen-activated protein kinase ͉ RNA splicing M ost of the actions of estrogen are thought to be mediated via the transcriptional control of target genes by nuclear estrogen receptors (ER) ␣ and ␤, members of the steroid hormone receptor gene superfamily that act as ligand-inducible transcription factors. ERs bind as dimers to specific estrogen response elements in the promoters of some target genes (1). However, most ER target promoters appear to recruit ERs without specific DNA binding, presumably through associations with sequence-specific factors bound to the promoters (2). ERs contain two transactivation functions (AFs), AF-1 in the Nterminal A͞B domain and AF-2 in the C-terminal ligand-binding E͞F domain. Ligand-induced transactivation by ERs requires multiple distinct classes of coactivator complexes, as well as a number of coregulators. The best-characterized complex contains p160͞SRC family proteins (3, 4) and CBP͞p300 histone acetyltransferases (5, 6), along with the RNA coactivator steroid receptor RNA activator (7) and presumably other known and unknown coactivators (8, 9). Another histone acetyltransferasescontaining complex, the TBP-free TAF II -containing (TFTC)-like complex (10), can also coactivate ER transactivation, as can the nonhistone acetyltransferases DRIP (VDR interacting protein)͞TRAP (thyroid hormone receptor-associated protein)͞ SMCC (SRB͞MED cofactor complexes)͞ARC (activatorrecruited cofactor) complex (11-13).ER-mediated estrogen signaling is known to involve cross-talk with other signaling pathways (14). For instance, growth factors potentiate estrogen-induced cellular proliferation in female reproductive tissues (15). Phosphorylation of the Ser-118 residue in the human ER␣ (hER␣) A͞B domain by mitogen-activated protein kinase (MAPK) activated by growth factors (16, 17) and cyclin-dependent kinase-7 (18) results in the potentiation of AF-1 function (16). However, the molecular basis of ER␣ AF-1 potentiation by MAPK-mediated phosphorylation remains unclear. In a previous study, we identified the DEAD-box RNA helicase subfamily member p68͞p72...

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Section: Phosphorylated Her␣ Associates With the Spliceosome Through mentioning

confidence: 94%

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Masuhiro

Mezaki²,

Sakari³

et al. 2005

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

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“…Phosphorylation of SF3b155 and CDC5L occurs concomittant with spliceosome activation and step I of splicing, respectively Numerous studies have shown that reversible phosphorylation plays an important role in pre-mRNA splicing (for review, see Soret and Tazi 2003). Previously, it was shown that the SF3b155 protein is phosphorylated prior to or during the first step of splicing (Wang et al 1998).…”

Section: Protein Dynamics During the B Act To C Complex Transitionmentioning

confidence: 99%

Characterization of purified human B^act spliceosomal complexes reveals compositional and morphological changes during spliceosome activation and first step catalysis

Bessonov¹,

Anokhina²,

Krasauskas³

et al. 2010

RNA

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195

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To better understand the compositional and structural dynamics of the human spliceosome during its activation, we set out to isolate spliceosomal complexes formed after precatalytic B but prior to catalytically active C complexes. By shortening the polypyrimidine tract of the PM5 pre-mRNA, which lacks a 39 splice site and 39 exon, we stalled spliceosome assembly at the activation stage. We subsequently affinity purified human B act complexes under the same conditions previously used to isolate B and C complexes, and analyzed their protein composition by mass spectrometry. A comparison of the protein composition of these complexes allowed a fine dissection of compositional changes during the B to B act and B act to C transitions, and comparisons with the Saccharomyces cerevisiae B act complex revealed that the compositional dynamics of the spliceosome during activation are largely conserved between lower and higher eukaryotes. Human SF3b155 and CDC5L were shown to be phosphorylated specifically during the B to B act and B act to C transition, respectively, suggesting these modifications function at these stages of splicing. The two-dimensional structure of the human B act complex was determined by electron microscopy, and a comparison with the B complex revealed that the morphology of the human spliceosome changes significantly during its activation. The overall architecture of the human and S. cerevisiae B act complex is similar, suggesting that many of the higher order interactions among spliceosomal components, as well as their dynamics, are also largely conserved.

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“…ATP is required not only for the ATPases/RNA helicases that drive the conformational rearrangements in the spliceosome, but also for the many kinases and phosphatases involved in the phosphorylation/dephosphorylation of numerous spliceosomal proteins. For example, SR protein phosphorylation/dephosphorylation is essential for premRNA splicing (for review, see Misteli 1999;Soret and Tazi 2003), with SR protein phosphorylation shown to be required during spliceosome assembly (Mermoud et al 1994;Xiao and Manley 1997). Dephosphorylation of several spliceosomal phosphoproteins is required for the catalytic steps of splicing, with PP1/PP2A phosphatases playing key roles at this stage (Mermoud et al 1992;Shi et al 2006).…”

Section: Introductionmentioning

confidence: 99%

ATPγS stalls splicing after B complex formation but prior to spliceosome activation

Agafonov

Santen

Kastner

et al. 2016

RNA

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The ATP analog ATPγS inhibits pre-mRNA splicing in vitro, but there have been conflicting reports as to which step of splicing is inhibited by this small molecule and its inhibitory mechanism remains unclear. Here we have dissected the effect of ATPγS on pre-mRNA splicing in vitro. Addition of ATPγS to splicing extracts depleted of ATP inhibited both catalytic steps of splicing. At ATPγS concentrations ≥0.5 mM, precatalytic B complexes accumulate, demonstrating a block prior to or during the spliceosome activation stage. Affinity purification of the ATPγS-stalled B complexes (B ATPγS ) and subsequent characterization of their abundant protein components by 2D gel electrophoresis revealed that B ATPγS complexes are compositionally more homogeneous than B complexes previously isolated in the presence of ATP. In particular, they contain little or no Prp19/CDC5L complex proteins, indicating that these proteins are recruited after assembly of the precatalytic spliceosome. Under the electron microscope, B ATPγS complexes exhibit a morphology highly similar to B complexes, indicating that the ATPγS-induced block in the transformation of the B to B act complex is not due to a major structural defect. Likely mechanisms whereby ATPγS blocks spliceosome assembly at the activation stage, including inhibition of the RNA helicase Brr2, are discussed. Given their more homogeneous composition, B complexes stalled by ATPγS may prove highly useful for both functional and structural analyses of the precatalytic spliceosome and its conversion into an activated B act spliceosomal complex.

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Phosphorylation-Dependent Control of the Pre-mRNA Splicing Machinery

Cited by 31 publications

References 265 publications

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Characterization of purified human B^act spliceosomal complexes reveals compositional and morphological changes during spliceosome activation and first step catalysis

ATPγS stalls splicing after B complex formation but prior to spliceosome activation

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Phosphorylation-Dependent Control of the Pre-mRNA Splicing Machinery

Cited by 31 publications

References 265 publications

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation

Characterization of purified human Bact spliceosomal complexes reveals compositional and morphological changes during spliceosome activation and first step catalysis

ATPγS stalls splicing after B complex formation but prior to spliceosome activation

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Characterization of purified human B^act spliceosomal complexes reveals compositional and morphological changes during spliceosome activation and first step catalysis