Discovery of noncanonical translation initiation sites through mass spectrometric analysis of protein N termini

Chan-Hyun Na, Mustafa A. Barbhuiya, Min Sik Kim, Steven Verbruggen, Stephen Eacker, Olga Pletnikova, Juan C Troncoso, Marc K Halushka, Gerben Menschaert, Christopher M. Overall, Akhilesh Pandey

Research output: Contribution to journalArticle

Abstract

Translation initiation generally occurs at AUG codons in eukaryotes, although it has been shown that non-AUG or noncanonical translation initiation can also occur. However, the evidence for noncanonical translation initiation sites (TISs) is largely indirect and based on ribosome profiling (Ribo-seq) studies. Here, using a strategy specifically designed to enrich N termini of proteins, we demonstrate that many human proteins are translated at noncanonical TISs. The large majority of TISs that mapped to 5 untranslated regions were noncanonical and led to N-terminal extension of annotated proteins or translation of upstream small open reading frames (uORF). It has been controversial whether the amino acid corresponding to the start codon is incorporated at the TIS or methionine is still incorporated. We found that methionine was incorporated at almost all noncanonical TISs identified in this study. Comparison of the TISs determined through mass spectrometry with ribosome profiling data revealed that about two-thirds of the novel annotations were indeed supported by the available ribosome profiling data. Sequence conservation across species and a higher abundance of noncanonical TISs than canonical ones in some cases suggests that the noncanonical TISs can have biological functions. Overall, this study provides evidence of protein translation initiation at noncanonical TISs and argues that further studies are required for elucidation of functional implications of such noncanonical translation initiation.

Original languageEnglish (US)
Pages (from-to)25-36
Number of pages12
JournalGenome Research
Volume28
Issue number1
DOIs
StatePublished - Jan 1 2018

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Ribosomes
Methionine
Translational Peptide Chain Initiation
Proteins
Initiator Codon
5' Untranslated Regions
Protein Biosynthesis
Eukaryota
Codon
Open Reading Frames
Mass Spectrometry
Amino Acids

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

Cite this

Discovery of noncanonical translation initiation sites through mass spectrometric analysis of protein N termini. / Na, Chan-Hyun; Barbhuiya, Mustafa A.; Kim, Min Sik; Verbruggen, Steven; Eacker, Stephen; Pletnikova, Olga; Troncoso, Juan C; Halushka, Marc K; Menschaert, Gerben; Overall, Christopher M.; Pandey, Akhilesh.

In: Genome Research, Vol. 28, No. 1, 01.01.2018, p. 25-36.

Research output: Contribution to journalArticle

Na, Chan-Hyun ; Barbhuiya, Mustafa A. ; Kim, Min Sik ; Verbruggen, Steven ; Eacker, Stephen ; Pletnikova, Olga ; Troncoso, Juan C ; Halushka, Marc K ; Menschaert, Gerben ; Overall, Christopher M. ; Pandey, Akhilesh. / Discovery of noncanonical translation initiation sites through mass spectrometric analysis of protein N termini. In: Genome Research. 2018 ; Vol. 28, No. 1. pp. 25-36.
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