Noncanonical Translation Initiation Comes of Age

ABSTRACT The canonical translation initiation mechanism involves base pairing between the mRNA and 16S rRNA. However, a variety of identified mechanisms deviate from this conventional route. Beck and Janssen (J Bacteriol 199:e00091-17, 2017, https://doi.org/10.1128/JB.00091-17 ) have recently described another noncanonical mode of translation initiation. Here, we describe how this process differs from previously reported mechanisms, with the hope that it will foster increased awareness of the diversity of regulatory mechanisms that await discovery.

70S-scanning initiation is a novel and frequent initiation mode of ribosomal translation in bacteria

According to the standard model of bacterial translation initiation, the small ribosomal 30S subunit binds to the initiation site of an mRNA with the help of three initiation factors (IF1–IF3). Here, we describe a novel type of initiation termed “70S-scanning initiation,” where the 70S ribosome does not necessarily dissociate after translation of a cistron, but rather scans to the initiation site of the downstream cistron. We detailed the mechanism of 70S-scanning initiation by designing unique monocistronic and polycistronic mRNAs harboring translation reporters, and by reconstituting systems to characterize each distinct mode of initiation. Results show that 70S scanning is triggered by fMet-tRNA and does not require energy; the Shine–Dalgarno sequence is an essential recognition element of the initiation site. IF1 and IF3 requirements for the various initiation modes were assessed by the formation of productive initiation complexes leading to synthesis of active proteins. IF3 is essential and IF1 is highly stimulating for the 70S-scanning mode. The task of IF1 appears to be the prevention of untimely interference by ternary aminoacyl (aa)-tRNA•elongation factor thermo unstable (EF-Tu)•GTP complexes. Evidence indicates that at least 50% of bacterial initiation events use the 70S-scanning mode, underscoring the relative importance of this translation initiation mechanism.

Ribosomal protein S19-binding domain provides insights into hantavirus nucleocapsid protein-mediated translation initiation mechanism

The binding domain for the ribosomal protein S19 was mapped to amino acids 151–175 of hantavirus nucleocapsid protein (N). This study provides insights about the mechanism of ribosome loading during the N-mediated translation initiation strategy.

Leucine-tRNA Initiates at CUG Start Codons for Protein Synthesis and Presentation by MHC Class I

Effective immune surveillance by cytotoxic T cells requires newly synthesized polypeptides for presentation by major histocompatibility complex (MHC) class I molecules. These polypeptides are produced not only from conventional AUG-initiated, but also from cryptic non–AUG-initiated, reading frames by distinct translational mechanisms. Biochemical analysis of ribosomal initiation complexes at CUG versus AUG initiation codons revealed that cells use an elongator leucine-bound transfer RNA (Leu-tRNA) to initiate translation at cryptic CUG start codons. CUG/Leu-tRNA initiation was independent of the canonical initiator tRNA (AUG/Met-tRNAiMet) pathway but required expression of eukaryotic initiation factor 2A. Thus, a tRNA-based translation initiation mechanism allows non–AUG-initiated protein synthesis and supplies peptides for presentation by MHC class I molecules.