title: Cancer-relevant splicing factor CAPERα engages the essential splicing factor SF3b155 in a specific ternary complex.
abstract: U2AF homology motifs (UHMs) mediate protein-protein interactions with U2AF ligand motifs (ULMs) of pre-mRNA splicing factors. The UHM-containing alternative splicing factor CAPERα regulates splicing of tumor-promoting VEGF isoforms, yet the molecular target of the CAPERα UHM is unknown. Here we present structures of the CAPERα UHM bound to a representative SF3b155 ULM at 1.7 Å resolution and, for comparison, in the absence of ligand at 2.2 Å resolution. The prototypical UHM/ULM interactions authenticate CAPERα as a bona fide member of the UHM family of proteins. We identify SF3b155 as the relevant ULM-containing partner of full-length CAPERα in human cell extracts. Isothermal titration calorimetry comparisons of the purified CAPERα UHM binding known ULM-containing proteins demonstrate that high affinity interactions depend on the presence of an intact, intrinsically unstructured SF3b155 domain containing seven ULM-like motifs. The interplay among bound CAPERα molecules gives rise to the appearance of two high affinity sites in the SF3b155 ULM-containing domain. In conjunction with the previously identified, UHM/ULM-mediated complexes of U2AF(65) and SPF45 with SF3b155, this work demonstrates the capacity of SF3b155 to offer a platform for coordinated recruitment of UHM-containing splicing factors.
authors.affiliation: From the Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642 and.
authors.forename: Sarah
authors.initials: S
authors.lastname: Loerch
authors.affiliation: the Howard Hughes Medical Institute and Programs in Gene Function and Expression and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605 alexandre.maucuer@inserm.fr.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Maucuer
authors.affiliation: the Howard Hughes Medical Institute and Programs in Gene Function and Expression and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Valérie
authors.initials: V
authors.lastname: Manceau
authors.affiliation: the Howard Hughes Medical Institute and Programs in Gene Function and Expression and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Green
authors.affiliation: From the Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642 and clara_kielkopf@urmc.rochester.edu.
authors.forename: Clara L
authors.initials: CL
authors.lastname: Kielkopf
mesh_terms: D020816:Amino Acid Motifs; D057809:HEK293 Cells; D006801:Humans; D009363:Neoplasm Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C084105:HCC1 autoantigen; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; C112491:SF3B1 protein, human
keywords: Crystal Structure; Gene Regulation; Protein Domain; Protein Structure and Folding; Protein-Protein Interaction; RNA Splicing