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Abstract:
The eukaryotic initiation factor 4F (eIF4F) complex recruits the 43S preinitiation complex to capped mRNAs. eIF4F comprises the cap-binding protein eIF4E, the RNA helicase eIF4A and the scaffolding protein eIF4G, which binds to both eIF4E and eIF4A. Assembly of eIF4F is regulated by eIF4E-binding pro- teins (4E-BPs), which repress translation by competing with eIF4G for binding to eIF4E. Metazoan eIF4G and 4E-BPs inter- act with eIF4E using a conserved bipartite binding mode involv- ing canonical and non-canonical eIF4E-binding motifs that bind to the dorsal and lateral surface of eIF4E, respectively. In S. cerevisiae (Sc), the two 4E-BPs p20 and Eap1p regulate cap- dependent translation. However, structural information on the assembly of yeast eIF4E-containing complexes is limited to the complex with eIF4G. Upon binding to eIF4E, Sc eIF4G forms a bracelet-like structure remarkably different from the bipartite binding mode seen in metazoan eIF4E–eIF4G complexes. This finding suggested that distinct interaction modes are used in yeast eIF4E-complexes, raising the question of how 4E-BPs function in translation control in fungi. Here, we present crystal structures of Sc eIF4E bound to p20 or Eap1p and the structure of the eIF4E–eIF4G complex from C. thermophilum. In contrast to prior studies, these structures show that in all eIF4E complexes, fungal eIF4E partners share conserved, core eIF4E-binding ele- ments present in metazoans and contain previously unknown non-canonical motifs that engage the lateral surface of eIF4E. Moreover, fungal eIF4E partners exhibit molecular adaptations that stabilize binding to eIF4E: p20 possesses a high-affinity canonical motif; Eap1p harbors a unique sequence stabilizing the interaction with the eIF4E dorsal surface; and eIF4G uses a fungi-specific bracelet-like structure to enhance binding to eIF4E. Our structural and biophysical studies uncover conserved princi- ples governing the assembly of the eIF4F complex and its regulation by 4E-BPs.
