title: RBFOX2 promotes protein 4.1R exon 16 selection via U1 snRNP recruitment.
abstract: The erythroid differentiation-specific splicing switch of protein 4.1R exon 16, which encodes a spectrin/actin-binding peptide critical for erythrocyte membrane stability, is modulated by the differentiation-induced splicing factor RBFOX2. We have now characterized the mechanism by which RBFOX2 regulates exon 16 splicing through the downstream intronic element UGCAUG. Exon 16 possesses a weak 5' splice site (GAG/GTTTGT), which when strengthened to a consensus sequence (GAG/GTAAGT) leads to near-total exon 16 inclusion. Impaired RBFOX2 binding reduces exon 16 inclusion in the context of the native weak 5' splice site, but not the engineered strong 5' splice site, implying that RBFOX2 achieves its effect by promoting utilization of the weak 5' splice site. We further demonstrate that RBFOX2 increases U1 snRNP recruitment to the weak 5' splice site through direct interaction between its C-terminal domain (CTD) and the zinc finger region of U1C and that the CTD is required for the effect of RBFOX2 on exon 16 splicing. Our data suggest a novel mechanism for exon 16 5' splice site activation in which the binding of RBFOX2 to downstream intronic splicing enhancers stabilizes the pre-mRNA-U1 snRNP complex through interactions with U1C.
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA. shu-ching_huang@dfci.harvard.edu
authors.forename: Shu-Ching
authors.initials: SC
authors.lastname: Huang
authors.forename: Alexander C
authors.initials: AC
authors.lastname: Ou
authors.forename: Jennie
authors.initials: J
authors.lastname: Park
authors.forename: Faye
authors.initials: F
authors.lastname: Yu
authors.forename: Brian
authors.initials: B
authors.lastname: Yu
authors.forename: Angela
authors.initials: A
authors.lastname: Lee
authors.forename: Guang
authors.initials: G
authors.lastname: Yang
authors.forename: Anyu
authors.initials: A
authors.lastname: Zhou
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Benz
mesh_terms: D001483:Base Sequence; D003598:Cytoskeletal Proteins / Q000235:genetics*; D005091:Exons; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics*; D017434:Protein Structure, Tertiary; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry / Q000378:metabolism; D016335:Zinc Fingers
chemical_list: D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; C454924:RBFOX2 protein, human; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D012097:Repressor Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; D017411:Ribonucleoproteins, Small Nuclear; C028841:erythrocyte membrane band 4.1 protein
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