title: Identification of novel proteins binding the AU-rich element of α-prothymosin mRNA through the selection of open reading frames (RIDome).
abstract: We describe here a platform for high-throughput protein expression and interaction analysis aimed at identifying the RNA-interacting domainome. This approach combines the selection of a phage library displaying "filtered" open reading frames with next-generation DNA sequencing. The method was validated using an RNA bait corresponding to the AU-rich element of α-prothymosin, an RNA motif that promotes mRNA stability and translation through its interaction with the RNA-binding protein ELAVL1. With this strategy, we not only confirmed known RNA-binding proteins that specifically interact with the target RNA (such as ELAVL1/HuR and RBM38) but also identified proteins not previously known to be ARE-binding (R3HDM2 and RALY). We propose this technology as a novel approach for studying the RNA-binding proteome.
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Patrucco
authors.affiliation: b Institute of Biomedical Technologies; National Research Council (ITB CNR) ; Milan , Italy.
authors.forename: Clelia
authors.initials: C
authors.lastname: Peano
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Chiesa
authors.affiliation: c Department of Life Sciences ; University of Trieste ; Italy.
authors.forename: Filomena
authors.initials: F
authors.lastname: Guida
authors.affiliation: c Department of Life Sciences ; University of Trieste ; Italy.
authors.forename: Imma
authors.initials: I
authors.lastname: Luisi
authors.affiliation: d Department of Chemistry ; University of Milan ; Italy.
authors.forename: Ilenia
authors.initials: I
authors.lastname: Boria
authors.affiliation: e Department of Sciences and Innovation ; Università del Piemonte Orientale ; Alessandria , Italy.
authors.forename: Flavio
authors.initials: F
authors.lastname: Mignone
authors.affiliation: b Institute of Biomedical Technologies; National Research Council (ITB CNR) ; Milan , Italy.
authors.forename: Gianluca
authors.initials: G
authors.lastname: De Bellis
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.|f Area of Neuroscience; SISSA ; Trieste , Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Zucchelli
authors.affiliation: f Area of Neuroscience; SISSA ; Trieste , Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Gustincich
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Santoro
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.|c Department of Life Sciences ; University of Trieste ; Italy.
authors.forename: Daniele
authors.initials: D
authors.lastname: Sblattero
authors.affiliation: a Department of Health Sciences and Interdisciplinary Research Center on Autoimmune Diseases (IRCAD) ; Università del Piemonte Orientale ; Novara , Italy.
authors.forename: Diego
authors.initials: D
authors.lastname: Cotella
mesh_terms: D063307:AU Rich Elements / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D016366:Open Reading Frames / Q000235:genetics*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*; D011498:Protein Precursors / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015203:Reproducibility of Results; D013947:Thymosin / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism
chemical_list: D011498:Protein Precursors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C040782:prothymosin alpha; D013947:Thymosin
keywords: AU-rich element; ELAVL1; R3HDM2; RALY; RBM38; RBPome; RNA-binding protein; next-generation DNA sequencing; open reading frame; phage display