title: Proteogenomic Discovery of a Small, Novel Protein in Yeast Reveals a Strategy for the Detection of Unannotated Short Open Reading Frames.
abstract: In recent years, proteomic data have contributed to genome annotation efforts, most notably in humans and mice, and spawned a field termed "proteogenomics". Yeast, in contrast with higher eukaryotes, has a small genome, which has lent itself to simpler ORF prediction. Despite this, continual advances in mass spectrometry suggest that proteomics should be able to improve genome annotation even in this well-characterized species. Here we applied a proteogenomics workflow to yeast to identify novel protein-coding genes. Specific databases were generated, from intergenic regions of the genome, which were then queried with MS/MS data. This suggested the existence of several putative novel ORFs of <100 codons, one of which we chose to validate. Synthetic peptides, RNA-Seq analysis, and evidence of evolutionary conservation allowed for the unequivocal definition of a new protein of 78 amino acids encoded on chromosome X, which we dub YJR107C-A. It encodes a new type of domain, which ab initio modeling suggests as predominantly α-helical. We show that this gene is nonessential for growth; however, deletion increases sensitivity to osmotic stress. Finally, from the above discovery process, we discuss a generalizable strategy for the identification of short ORFs and small proteins, many of which are likely to be undiscovered.
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Daniel
authors.initials: D
authors.lastname: Yagoub
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Aidan P
authors.initials: AP
authors.lastname: Tay
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Zhiliang
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Joshua J
authors.initials: JJ
authors.lastname: Hamey
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Curtis
authors.initials: C
authors.lastname: Cai
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Samantha Z
authors.initials: SZ
authors.lastname: Chia
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Gene
authors.initials: G
authors.lastname: Hart-Smith
authors.affiliation: Systems Biology Initiative, School of Biotechnology and Biomolecular Sciences, University of New South Wales , Sydney, New South Wales 2052, Australia.
authors.forename: Marc R
authors.initials: MR
authors.lastname: Wilkins
mesh_terms: D000595:Amino Acid Sequence; D030541:Databases, Genetic; D055786:Gene Knockout Techniques; D023281:Genomics / Q000379:methods*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D017423:Sequence Analysis, RNA / Q000379:methods; D053719:Tandem Mass Spectrometry
chemical_list: D029701:Saccharomyces cerevisiae Proteins
keywords: proteogenomics; small ORFs; small proteins; yeast