Exon‐independent recruitment of SRSF1 is mediated by U1 snRNP stem‐loop 3

Jobbins, Andrew M; Campagne, Sébastien; Weinmeister, Robert; Lucas, Christian M; Gosliga, Alison R; Cléry, Antoine; Chen, Li; Eperon, Lucy P.; Hodson, Mark; Hudson, Andrew J.; Allain, Frédéric H.‐T.; Eperon, Ian C.

doi:10.15252/embj.2021107640

Cited by 40 publications

(46 citation statements)

References 162 publications

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“…ESEs containing GA dinucleotides may bind a subset of SR proteins ( Supplementary Table S2 ) but individual GAY-to-CAC mutations and ESE-to-ESS remodeling of the DADLD site were insufficient to alter exon 4b inclusion levels by SR protein regulators (Figure 5H ). The failure to respond to mutations is consistent with a significant ESE-independent component of SRSF1 action, as proposed ( 124 , 125 ). In the most recent model ( 125 ), SRSF1 could be directly recruited to 5′ss through interactions with stem–loop 3 of U1 small nuclear ribonucleoprotein (snRNP), namely, between SRSF1 RRM1 and the CA motif at the 5′ part of the stem-loop, and between RRM2 and a double-stranded GGA motif, with ESEs showing only low transient SRSF1 occupancy ( 125 , 126 ).…”

Section: Discussionsupporting

confidence: 84%

“…In the most recent model ( 125 ), SRSF1 could be directly recruited to 5′ss through interactions with stem–loop 3 of U1 small nuclear ribonucleoprotein (snRNP), namely, between SRSF1 RRM1 and the CA motif at the 5′ part of the stem-loop, and between RRM2 and a double-stranded GGA motif, with ESEs showing only low transient SRSF1 occupancy ( 125 , 126 ). In other words, the ESE-independent SRSF1 interactions appearred to be sufficient for connecting U1 and U2 snRNPs ( 125 ).…”

Section: Discussionmentioning

confidence: 99%

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Exonic splicing code and protein binding sites for calcium

Pengelly

Bakhtiar

Borovská

et al. 2022

Nucleic Acids Research

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Auxilliary splicing sequences in exons, known as enhancers (ESEs) and silencers (ESSs), have been subject to strong selection pressures at the RNA and protein level. The protein component of this splicing code is substantial, recently estimated at ∼50% of the total information within ESEs, but remains poorly understood. The ESE/ESS profiles were previously associated with the Irving-Williams (I-W) stability series for divalent metals, suggesting that the ESE/ESS evolution was shaped by metal binding sites. Here, we have examined splicing activities of exonic sequences that encode protein binding sites for Ca2+, a weak binder in the I-W affinity order. We found that predicted exon inclusion levels for the EF-hand motifs and for Ca2+-binding residues in nonEF-hand proteins were higher than for average exons. For canonical EF-hands, the increase was centred on the EF-hand chelation loop and, in particular, on Ca2+-coordinating residues, with a 1>12>3∼5>9 hierarchy in the 12-codon loop consensus and usage bias at codons 1 and 12. The same hierarchy but a lower increase was observed for noncanonical EF-hands, except for S100 proteins. EF-hand loops preferentially accumulated exon splits in two clusters, one located in their N-terminal halves and the other around codon 12. Using splicing assays and published crosslinking and immunoprecipitation data, we identify candidate trans-acting factors that preferentially bind conserved GA-rich motifs encoding negatively charged amino acids in the loops. Together, these data provide evidence for the high capacity of codons for Ca2+-coordinating residues to be retained in mature transcripts, facilitating their exon-level expansion during eukaryotic evolution.

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Section: Discussionsupporting

confidence: 84%

Section: Discussionmentioning

confidence: 99%

Exonic splicing code and protein binding sites for calcium

Pengelly

Bakhtiar

Borovská

et al. 2022

Nucleic Acids Research

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“…6). We found that SRSF1 (Jobbins, Campagne et al, 2022) and FUS (Jutzi, Campagne et al, 2020) could interact directly with the SL3 of the human U1 snRNA. The specific interaction between Npl3 and the U2 snRNA reported here implicates a broader role for the U snRNAs.…”

Section: Discussionmentioning

confidence: 88%

“…6). We found that SRSF1 (Jobbins, Campagne et al, 2022) and FUS (Jutzi, Campagne et al, 2020) could interact directly with the SL3 of the human U1 snRNA.…”

Section: Functional Insights Into the Role Of Npl3 During Rna Splicingmentioning

confidence: 81%

The structure of yeast Npl3 bound to RNA reveals a cooperative sequence-specific recognition and an RNA chaperone role in splicing

Moursy

Cléry

Gerhardy

et al. 2022

Preprint

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The conserved SR-like protein Npl3 promotes splicing of diverse pre-mRNAs. However, the RNA sequence(s) recognized by the RNA Recognition Motifs (RRM1 & RRM2) of Npl3 during the splicing reaction remain elusive. Here, we developed a split-iCRAC approach in yeast to uncover the consensus sequence bound to each RRM. High-resolution NMR structures show that RRM2 recognizes a GNGG motif leading to an unusual mille-feuille topology. These structures also reveal how RRM1 preferentially interacts with a CC-dinucleotide upstream of this motif, and how the inter-RRM linker and the region C-terminal to RRM2 contributes to cooperative RNA-binding. Structure-guided functional studies show that Npl3 genetically interacts with U2 snRNP specific factors and we provide evidence that Npl3 melts U2 snRNA stem-loop I, a prerequisite for U2/U6 duplex formation within the catalytic center of the Bact spliceosomal complex. Thus, our findings provide insights into an unanticipated RNA chaperoning role for Npl3 during spliceosome active site formation.

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“…U1 snRNP-mediated SR protein displacement appears to involve a complex and dynamic mechanism. SRSF1 interacts with U1-70k (37) and the stem-loop III of U1 snRNA (43).…”

Section: Discussionmentioning

confidence: 99%

Cooperative engagement and subsequent selective displacement of SR proteins define the pre-mRNA 3D structural scaffold for early spliceosome assembly

Saha

Ghosh

2021

Preprint

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Coordination of different serine-arginine-rich (SR) proteins – a class of critical splicing activators – facilitates recognition of the highly degenerate cognate splice signal sequences against the background sequences. Yet, the mechanistic details of their actions remain unclear. Here we show that cooperative binding of SR proteins to exonic and intronic motifs remodels the pre-mRNA 3D structural scaffold. The scaffold generated by pre-mRNA-specific combinations of different SR proteins in an appropriate stoichiometry is recognized by U1 snRNP. A large excess of U1 snRNP particles displaces the majority of the bound SR protein molecules from the remodeled pre-mRNA. A higher than optimal stoichiometry of SR proteins occludes the binding sites on the pre-mRNA, raising the U1 snRNP levels required for SR protein displacement and potentially impeding spliceosome assembly. This novel step is important for distinguishing the substrate and the non-substrate by U2AF65 – the primary 3′ splice site-recognizing factor. Overall, this work elucidates early regulatory steps of mammalian splicing substrate definition by SR proteins.

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Exon‐independent recruitment of SRSF1 is mediated by U1 snRNP stem‐loop 3

Cited by 40 publications

References 162 publications

Exonic splicing code and protein binding sites for calcium

Exonic splicing code and protein binding sites for calcium

The structure of yeast Npl3 bound to RNA reveals a cooperative sequence-specific recognition and an RNA chaperone role in splicing

Cooperative engagement and subsequent selective displacement of SR proteins define the pre-mRNA 3D structural scaffold for early spliceosome assembly

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