title: Identification of a signature motif for the eIF4a3-SECIS interaction.
abstract: eIF4a3, a DEAD-box protein family member, is a component of the exon junction complex which assembles on spliced mRNAs. The protein also acts as a transcript-selective translational repressor of selenoprotein synthesis during selenium deficiency. Selenocysteine (Sec) incorporation into selenoproteins requires a Sec Insertion Sequence (SECIS) element in the 3' untranslated region. During selenium deficiency, eIF4a3 binds SECIS elements from non-essential selenoproteins, preventing Sec insertion. We identified a molecular signature for the eIF4a3-SECIS interaction using RNA gel shifts, surface plasmon resonance and enzymatic foot printing. Our results support a two-site interaction model, where eIF4a3 binds the internal and apical loops of the SECIS. Additionally, the stability of the complex requires uridine in the SECIS core. In terms of protein requirements, the two globular domains of eIF4a3, which are connected by a linker, are both critical for SECIS binding. Compared to full-length eIF4a3, the two domains in trans bind with a lower association rate but notably, the uridine is no longer important for complex stability. These results provide insight into how eIF4a3 discriminates among SECIS elements and represses translation.
authors.affiliation: Department of Cell Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA.
authors.forename: Michael E
authors.initials: ME
authors.lastname: Budiman
authors.forename: Jodi L
authors.initials: JL
authors.lastname: Bubenik
authors.forename: Donna M
authors.initials: DM
authors.lastname: Driscoll
mesh_terms: D020413:3' Untranslated Regions; D000818:Animals; D001665:Binding Sites; D053487:DEAD-box RNA Helicases / Q000737:chemistry / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D010448:Peptide Initiation Factors / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D051140:Selenoproteins / Q000096:biosynthesis / Q000235:genetics*; D014529:Uridine / Q000737:chemistry
chemical_list: D020413:3' Untranslated Regions; D010448:Peptide Initiation Factors; D012333:RNA, Messenger; D051140:Selenoproteins; C568403:eIF4a3 protein, rat; D053487:DEAD-box RNA Helicases; D014529:Uridine