Functional role and ribosomal position of the unique N-terminal region of DHX29, a factor required for initiation on structured mammalian mRNAs. Issue 22 (9th December 2021)

• Record Type: Journal Article
• Authors: Sweeney, Trevor R; Dhote, Vidya; Guca, Ewelina; Hellen, Christopher U T; Hashem, Yaser; Pestova, Tatyana V
• Is Part Of: Nucleic acids research. Volume 49:Issue 22(2021)
• Page Start: 12955
• Page End: 12969
• Publication Date: 2021-12-09
• Subjects: Nucleic acids -- Periodicals; Molecular biology -- Periodicals; 572.805
• DOI: 10.1093/nar/gkab1192
• Languages: English
• ISSNs: 0305-1048

Abstract

Abstract: Translation initiation on structured mammalian mRNAs requires DHX29, a DExH protein that comprises a unique 534-aa-long N-terminal region (NTR) and a common catalytic DExH core. DHX29 binds to 40S subunits and possesses 40S-stimulated NTPase activity essential for its function. In the cryo-EM structure of DHX29-bound 43S preinitiation complexes, the main DHX29 density resides around the tip of helix 16 of 18S rRNA, from which it extends through a linker to the subunit interface forming an intersubunit domain next to the eIF1A binding site. Although a DExH core model can be fitted to the main density, the correlation between the remaining density and the NTR is unknown. Here, we present a model of 40S-bound DHX29, supported by directed hydroxyl radical cleavage data, showing that the intersubunit domain comprises a dsRNA-binding domain (dsRBD, aa 377–448) whereas linker corresponds to the long α-helix (aa 460–512) that follows the dsRBD. We also demonstrate that the N-terminal α-helix and the following UBA-like domain form a four-helix bundle (aa 90–166) that constitutes a previously unassigned section of the main density and resides between DHX29's C-terminal α-helix and the linker. In vitro reconstitution experiments revealed the critical and specific roles of these NTR elements for DHX29's function.