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

Agafonov, Dmitry E.; Santen, Marieke van; Kastner, Berthold; Dube, Prakash; Will, Cindy L.; Urlaub, Henning; Lührmann, Reinhard

doi:10.1261/rna.057810.116

Cited by 10 publications

(14 citation statements)

References 47 publications

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“…The Prp39-Prp42 heterodimer binds Yhc1 to anchor the U2 snRNP 3' domain to the U1 snRNP. Similar interactions were observed biochemically between the human alternative splicing factor PRPF39 homodimer and U1C12 (yeast Yhc1), suggesting that PRPF39 may likewise contact the human U2 3' domain, though it is not an obligate component of the human A complex20 (Fig. 2a).…”

supporting

confidence: 65%

Prespliceosome structure provides insights into spliceosome assembly and regulation

Plaschka

Lin

Charenton

et al. 2018

Nature

136

152

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The spliceosome catalyses the excision of introns from pre-mRNA in two steps, branching and exon ligation, and is assembled from five small nuclear ribonucleoprotein particles (snRNPs; U1, U2, U4, U5, U6) and numerous non-snRNP factors. For branching, the intron 5' splice site and the branch point sequence are selected and brought by the U1 and U2 snRNPs into the prespliceosome, which is a focal point for regulation by alternative splicing factors. The U4/U6.U5 tri-snRNP subsequently joins the prespliceosome to form the complete pre-catalytic spliceosome. Recent studies have revealed the structural basis of the branching and exon-ligation reactions, however, the structural basis of the early events in spliceosome assembly remains poorly understood. Here we report the cryo-electron microscopy structure of the yeast Saccharomyces cerevisiae prespliceosome at near-atomic resolution. The structure reveals an induced stabilization of the 5' splice site in the U1 snRNP, and provides structural insights into the functions of the human alternative splicing factors LUC7-like (yeast Luc7) and TIA-1 (yeast Nam8), both of which have been linked to human disease. In the prespliceosome, the U1 snRNP associates with the U2 snRNP through a stable contact with the U2 3' domain and a transient yeast-specific contact with the U2 SF3b-containing 5' region, leaving its tri-snRNP-binding interface fully exposed. The results suggest mechanisms for 5' splice site transfer to the U6 ACAGAGA region within the assembled spliceosome and for its subsequent conversion to the activation-competent B-complex spliceosome. Taken together, the data provide a working model to investigate the early steps of spliceosome assembly.

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supporting

confidence: 65%

Prespliceosome structure provides insights into spliceosome assembly and regulation

Plaschka

Lin

Charenton

et al. 2018

Nature

136

152

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“…Interestingly, in human Prp2 (DHX16) the most N-terminal tip of the corresponding domain has been proposed to contain a PWI-like domain, which is also present in the spliceosomal helicases Brr2 (U5-200K) and has been predicted for Prp22 (DHX8) (Absmeier et al, 2015;Korneta et al, 2012). In Brr2 the PWI-like domain-containing N-terminal region has been proposed to act as a structural switch autoinhibiting Brr2 via substrate competition and conformational clamping (Absmeier et al, 2015;Agafonov et al, 2016;Nguyen et al, 2016). In the absence of detailed structural and biochemical information on the N-terminal extension of Prp2, one can only speculate about the functional role of this domain.…”

Section: Discussionmentioning

confidence: 99%

Crystal structure of the spliceosomal DEAH-box ATPase Prp2

Schmitt

Hamann

Neumann

et al. 2018

Acta Cryst Sect D Struct Biol

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“…Many splicing regulatory factors and general splicing factors purify with SWI/SNF subunits (Zhao et al, 1998;Dellaire et al, 2002) and purifications of the snRNP U2 spliceosome component also include several SWI/SNF subunits (Makarov et al, 2012;Allemand et al, 2016). We performed an analysis of BRG1 and BRM interacting proteins in the RNP fraction and it revealed that BRG1, in particular, interacted with U2 snRNP and U5-U6 snRNP factors that assemble early in the splicing cycle (Lardelli et al, 2010;Hegele et al, 2012;Agafonov et al, 2016;Haselbach et al, 2018). In addition to general splicing factors, we also found that BRG1 interacted with many regulatory RNA binding factors, such as hnRNPs and RNA helicases.…”

Section: Discussionmentioning

confidence: 90%

The SWI/SNF subunits BRG1 affects alternative splicing by changing RNA binding factor interactions with RNA

Mackowiak

Guo

et al. 2019

Preprint

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BRG1 and BRM are ATPase core subunits of the human SWI/SNF chromatin remodelling complexes. The function of the SWI/SNF complexes in transcriptional initiation has been well studied, while a function in alternative splicing has only been studied for a few cases for BRM-containing SWI/SNF complexes. Here, we have expressed BRG1 in C33A cells, a BRG1 and BRM-deficient cell line, and we have analysed the effects on the transcriptome by RNA sequencing. We have shown that BRG1 expression affects the splicing of a subset of genes. For some, BRG1 expression favours exon inclusion and for others, exon skipping. Some of the changes in alternative splicing induced by BRG1 expression do not require the ATPase activity of BRG1. Among the exons regulated through an ATPase-independent mechanism, the included exons had signatures of high GC-content and lacked a positioned nucleosome at the exon. By investigating three genes in which the expression of either wild-type BRG1 or a BRG1-ATPase-deficient variant favoured exon inclusion, we showed that expression of the ATPases promotes the local recruitment of RNA binding factors to chromatin and RNA in a differential manner. The hnRNPL, hnRNPU and SAM68 proteins associated to chromatin in C33A cells expressing BRG1 or BRM, but their association with RNA varied. We propose that SWI/SNF can regulate alternative splicing by interacting with splicing-RNA binding factor and altering their binding to the nascent pre-mRNA, which changes RNP structure. Author summarySplicing, in particular alternative splicing, is a combinatorial process which involves splicing factor complexes and many RNA binding splicing regulatory proteins in different constellations. Most splicing events occur during transcription, which also makes the DNA sequence, the chromatin state and the transcription rate at the exons important components that influence the splicing outcome. We show here that the ATP-dependent chromatin remodelling complex SWI/SNF influences the interactions of splicing regulatory factors with RNA during transcription on certain exons that have a high GC-content. The splicing on this type of exon rely on the ATPase BRG1 and favour inclusion of alternative exons in an ATP-independent manner. SWI/SNF complexes are known to alter the chromatin structure at promoters in transcription initiation, and have been previously shown to alter the transcription rate or nucleosome position in splicing. Our results suggests a further mechanism for chromatin remodelling proteins in splicing: to change the interaction patterns of RNA binding splicing regulatory factors at alternative exons to alter the splicing outcome. Biegel JA, Busse TM, Weissman BE. SWI/SNF chromatin remodeling complexes and cancer. Am Lynch KW. Global analysis of physical and functional RNA targets of hnRNP L reveals distinct sequence and epigenetic features of repressed and enhanced exons. :52. Custódio N, Carmo-Fonseca M. Co-transcriptional splicing and the CTD code. Crit Rev Biochem Mol Biol. 2016 Sep;51(5):395-411. De Conti L, Baralle M...

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

Cited by 10 publications

References 47 publications

Prespliceosome structure provides insights into spliceosome assembly and regulation

Prespliceosome structure provides insights into spliceosome assembly and regulation

Crystal structure of the spliceosomal DEAH-box ATPase Prp2

The SWI/SNF subunits BRG1 affects alternative splicing by changing RNA binding factor interactions with RNA

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