Amino Termini of Many Yeast Proteins Map to Downstream Start Codons

Abstract

Comprehensive knowledge of proteome complexity is crucial to understanding cell function. Amino termini of yeast proteins were identified through peptide mass spectrometry on glutaraldehyde-treated cell lysates as well as a parallel assessment of publicly-deposited spectra. An unexpectedly large fraction of detected amino-terminal peptides (35%) mapped to translation initiation at AUG codons downstream of the annotated start codon. Many of the implicated genes have suboptimal sequence contexts for translation initiation near their annotated AUG, and their ribosome profiles show elevated tag densities consistent with translation initiation at downstream AUGs as well as their annotated AUGs. These data suggest that a significant fraction of the yeast proteome derives from initiation at downstream AUGs, increasing significantly the repertoire of encoded proteins and their potential functions and cellular localizations.

Roles for transcript leaders in translation and mRNA decay revealed by transcript leader sequencing

“…Such internal TSSs exhibited characteristic nucleosome signatures similar to canonical TSSs, suggesting they are true TSSs and not merely artifacts of the technique. Furthermore, protein products from internal initiation have recently been identified by mass spectrometry, thus corroborating our findings here that such internal transcripts are translated (Fournier et al 2012). Analysis of transcript 59 ends also yielded a significant number of ORF-internal reads in Drosophila (16%) (Ni et al 2010) and human cells (2%-3%) (Kanamori-Katayama et al 2011).…”

Constitutive and Nitrogen Catabolite Repression-sensitive Production of Gat1 Isoforms

“…The alternative translational initiation phenomena described here may have arisen for any one of several reasons: (i) differential mRNA production over the GAT1 locus (alternative splicing or start site selection); (ii) utilization of several translation start sites, associated with an internal ribosomal entry site(s), caused by leaky ribosome scanning (56,57); or (iii) ribosome shunting (58,59). Such regulatory events have been documented with respect to controlled production of DNA-binding proteins (60) or in response to environmental cues or stresses (61,62).…”

Ribosome Profiling as a Tool to Decipher Viral Complexity

“…This internal initiation, resulting in the production of truncated proteins, can result from leaky scanning past the first AUG codon on a transcript or from production of truncated transcripts beginning past the first AUG. These truncated proteins, many of which can be detected by proteomics (44)(45)(46), may have functions distinct from those initiated at the first, canonical AUG. A truncated form of the antiviral signaling protein MAVS, produced by leaky scanning past the first start codon and initiation at a downstream AUG, actually inhibits the signaling activity of the full-length protein (47). Similar opposition between alternative protein isoforms is seen in C/EBP transcription factors (48) and may reflect a more general model, as short protein isoforms may be able to compete with long isoforms for binding sites but lack other activities once bound.…”