Reference: Wacholder A and Carvunis AR (2023) Biological factors and statistical limitations prevent detection of most noncanonical proteins by mass spectrometry. PLoS Biol 21(12):e3002409

Reference Help

Abstract


Ribosome profiling experiments indicate pervasive translation of short open reading frames (ORFs) outside of annotated protein-coding genes. However, shotgun mass spectrometry (MS) experiments typically detect only a small fraction of the predicted protein products of this noncanonical translation. The rarity of detection could indicate that most predicted noncanonical proteins are rapidly degraded and not present in the cell; alternatively, it could reflect technical limitations. Here, we leveraged recent advances in ribosome profiling and MS to investigate the factors limiting detection of noncanonical proteins in yeast. We show that the low detection rate of noncanonical ORF products can largely be explained by small size and low translation levels and does not indicate that they are unstable or biologically insignificant. In particular, proteins encoded by evolutionarily young genes, including those with well-characterized biological roles, are too short and too lowly expressed to be detected by shotgun MS at current detection sensitivities. Additionally, we find that decoy biases can give misleading estimates of noncanonical protein false discovery rates, potentially leading to false detections. After accounting for these issues, we found strong evidence for 4 noncanonical proteins in MS data, which were also supported by evolution and translation data. These results illustrate the power of MS to validate unannotated genes predicted by ribosome profiling, but also its substantial limitations in finding many biologically relevant lowly expressed proteins.

Reference Type
Journal Article
Authors
Wacholder A, Carvunis AR
Primary Lit For
Additional Lit For
Review For

Gene Ontology Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene/Complex Qualifier Gene Ontology Term Aspect Annotation Extension Evidence Method Source Assigned On Reference

Phenotype Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details.

Gene Phenotype Experiment Type Mutant Information Strain Background Chemical Details Reference

Disease Annotations


Increase the total number of rows showing on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene Disease Ontology Term Qualifier Evidence Method Source Assigned On Reference

Regulation Annotations


Increase the total number of rows displayed on this page using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; to filter the table by a specific experiment type, type a keyword into the Filter box (for example, “microarray”); download this table as a .txt file using the Download button or click Analyze to further view and analyze the list of target genes using GO Term Finder, GO Slim Mapper, or SPELL.

Regulator Target Direction Regulation Of Happens During Method Evidence

Post-translational Modifications


Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through its pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Site Modification Modifier Reference

Interaction Annotations


Genetic Interactions

Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details about experiment type and any other genes involved in the interaction.

Interactor Interactor Allele Assay Annotation Action Phenotype SGA score P-value Source Reference

Physical Interactions

Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through the table's pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table; click on the small "i" buttons located within a cell for an annotation to view further details about experiment type and any other genes involved in the interaction.

Interactor Interactor Assay Annotation Action Modification Source Reference

Functional Complementation Annotations


Increase the total number of rows showing on this page by using the pull-down located below the table, or use the page scroll at the table's top right to browse through its pages; use the arrows to the right of a column header to sort by that column; filter the table using the "Filter" box at the top of the table.

Gene Species Gene ID Strain background Direction Details Source Reference