Structural insights into a regulatory mechanism of FIR RRM1–FUSE interaction

Ni, Xiaoqi; Joerger, A.C.; Chaikuad, A.; Knapp, Stefan

doi:10.1098/rsob.230031

Cited by 3 publications

References 31 publications

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The Arabidopsis SR45 splicing factor bridges the splicing machinery and the exon–exon junction complex

Fanara,

Schloesser,

Joris

et al. 2024

Journal of Experimental Botany

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The Arabidopsis splicing factor serine/arginine-rich 45 (SR45) contributes to several biological processes. The sr45-1 loss-of-function mutant exhibits delayed root development, late flowering, unusual numbers of floral organs, shorter siliques with decreased seed sets, narrower leaves and petals, and altered metal distribution. SR45 bears a unique RNA recognition motif (RRM) flanked by one serine/arginine-rich (RS) domain on both sides. Here, we studied the function of each of SR45 domains by examining their involvement in: (i) the spatial distribution of SR45, (ii) the establishment of a protein-protein interaction network including spliceosomal and exon-exon junction complex (EJC) components, and (iii) the RNA binding specificity. We report that the endogenous SR45 promoter is active during vegetative and reproductive growth, and that the SR45 protein localizes in the nucleus. We demonstrate that the C-terminal arginine/serine-rich domain is a determinant of nuclear localization. We show that the SR45 RNA recognition motif (RRM) domain specifically binds purine-rich RNA motifs via three residues (H101, H141, Y143), and is also involved in protein-protein interactions. We further show that SR45 bridges both mRNA splicing and surveillance machineries as a partner of EJC core components and peripheral factors, which requires phosphoresidues likely phosphorylated by kinases from both CLK and SRPK families. Our findings provide insights into the contribution of each SR45 domain to both spliceosome and EJC assemblies.

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The Arabidopsis SR45 splicing factor bridges the splicing machinery and the exon–exon junction complex

Fanara,

Schloesser,

Joris

et al. 2024

Journal of Experimental Botany

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Structural insights into a regulatory mechanism of FIR RRM1–FUSE interaction

Joerger

Chaikuad

et al. 2023

Open Biol.

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FUBP-interacting repressor (FIR) is a suppressor of transcription of the proto-oncogene MYC. FIR binds to the far upstream element (FUSE) of the MYC promoter. Competition of FIR with FUSE-binding protein 1 (FUBP1) is a key mechanism of MYC transcriptional regulation. To gain insights into the structural mechanisms regulating FIR DNA interaction, we determined the crystal structure of two FIR RRM domains (RRM1-2) with single-stranded FUSE DNA sequences. These structures revealed an ability of the RRM domain to recognize diverse FUSE regions through distinct intermolecular interactions and binding modes. Comparative structural analyses against available RRM-ssDNA/RNA complexes showed that the nucleotide configurations in FIR were similar to those in other RRMs that harbour a tyrosine at the conserved aromatic position in the RNP2 motif (Y-type RRM), but not those with a phenylalanine (F-type RRM). Site-directed mutagenesis experiments demonstrated that a single substitution, Y115F, altered the binding affinities of oligonucleotides to FIR RRM, suggesting an important role of this conserved aromatic residue in ssDNA/RNA interactions. Our study provides the structural basis for further mechanistic studies on this important protein–DNA interaction.

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TheArabidopsisSR45 splicing factor bridges the splicing machinery and the exon-exon junction complex

Fanara,

Schloesser,

Joris

et al. 2023

Preprint

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The Arabidopsis splicing factor serine/arginine-rich 45 (SR45) contributes to several biological processes. The sr45-1 loss-of-function mutant exhibits delayed root development, late flowering, unusual numbers of floral organs, shorter siliques with decreased seed sets, narrower leaves and petals, and altered metal distribution. Here, we studied the function of each of SR45 domains by examining their involvement in: (i) the spatial distribution of SR45, (ii) the establishment of a protein-protein interaction network including spliceosomal and exon-exon junction complex (EJC) components, and (iii) the RNA binding specificity. We report that the endogenous SR45 promoter is active during vegetative and reproductive growth, and that the SR45 protein localizes in the nucleus. We demonstrate that the C-terminal arginine/serine-rich domain is a determinant of nuclear localization. We show that the SR45 RNA recognition motif (RRM) domain specifically binds purine-rich RNA motifs via three residues (H101, H141, Y143), and is also involved in protein-protein interactions. We further show that SR45 bridges both mRNA splicing and surveillance machineries as a partner of EJC core components and peripheral factors, which requires phosphoresidues likely phosphorylated by kinases from both CLK and SRPK families. Our findings provide insights into the contribution of each SR45 domain to both spliceosome and EJC assemblies.

show abstract

Structural insights into a regulatory mechanism of FIR RRM1–FUSE interaction

Cited by 3 publications

References 31 publications

The Arabidopsis SR45 splicing factor bridges the splicing machinery and the exon–exon junction complex

The Arabidopsis SR45 splicing factor bridges the splicing machinery and the exon–exon junction complex

Structural insights into a regulatory mechanism of FIR RRM1–FUSE interaction

TheArabidopsisSR45 splicing factor bridges the splicing machinery and the exon-exon junction complex

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