Jul 2026· Experimental and molecular pathology (Print)· Vol 147, pp.
105069
· 0 citations· 145 references
Medicine
TL;DR
A concise summary of the origin, discovery methods, and classification of non-canonical ORFs offers insights into the origins and functions of non-canonical ORF-encoded peptides from an evolutionary perspective, while also exploring the biological functions and regulatory mechanisms of these non-canonical ORF-encoded hidden proteins in tumorigenesis and progression.
Abstract
There is increasing evidence that translation is not limited to annotated protein-coding genes. Ribosome profiling sequencing, mass spectrometry-based proteomics, and immunopeptidomics have identified the productive translation of non-canonical open reading frames (ORFs). This suggests that the functional proteome includes not only conserved proteins but also proteins hidden in non-coding RNAs and de novo proteins. Some of these translated products are functional peptides, while others may be non-functional, potentially arising from evolutionary events. Several non-canonical ORF-encoded peptides have been found to regulate multiple physiological and pathological functions, particularly in cancer, immunity, and inflammation, indicating that they have potential as biomarkers and novel therapeutic targets. To better understand the diversity of functional peptides and translated non-canonical ORFs based on existing data, we summarize their classification according to transcriptional features and supporting evidence, including non-canonical ORFs located in ncRNAs and canonical mRNAs. This review provides a concise summary of the origin, discovery methods, and classification of non-canonical ORFs. It offers insights into the origins and functions of non-canonical ORF-encoded peptides from an evolutionary perspective, while also exploring the biological functions and regulatory mechanisms of these non-canonical ORF-encoded hidden proteins in tumorigenesis and progression.
The potential for open modification searching to correct potential mistakes in non-canonical proteins detection by preventing modified canonical peptides or variants from being incorrectly identified as non-canonical peptides is shown.
V. Vasylieva, Enrico Massignani, Tine Claeys et al.· bioRxiv· 0 citations
The pangenome provides unparalleled specificity to understand genetic variation in SD genes allowing us to distinguish functional genes from pseudogenes and highlighting potential gene innovations that arose most recently in human evolution.
Luyao Ren, DongAhn Yoo, Katarina Vlajic et al.· bioRxiv· 0 citations
Quantifying the diversity of RNAs and proteins produced by cells is fundamental to the biological and clinical sciences. However, many RNAs and proteins remain uncharacterised, especially proteins translated from alternate RNA isoforms; untranslated regions of mRNAs and non-coding RNAs, as well as the effects of DNA variation on protein sequences. Proteogenomics aims to characterise the complete proteome by integrating genomics and/or transcriptomics with proteomics, but current tools have limitations in useability, analysis features and visualisation of resulting data. To address these gaps, we developed GenomeProt, a user-friendly GUI-based tool for integrative proteogenomic analysis. We demonstrate its utility by integrating long-read RNA sequencing with mass-spectrometry-based proteomics to pinpoint proteoform expression generated by alternative splicing; discover novel, unannotated proteins in human brain samples; and quantify variant-containing peptides associated with treatment resistance in a melanoma xenograft model. GenomeProt brings the discovery power of proteogenomics to biologists, illuminating the hidden proteome.
Hitesh Kore, Josie Gleeson, C. Wan et al.· bioRxiv· 1 citation
This work systematically quantified the intracellular accumulation of 111 evolutionarily conserved human uORFps and identified a conserved class of human uORFs that encode intrinsic barriers to productive translation and provide a rigorous framework for understanding how noncanonical coding sequences shape the human proteome.
Hinata Hashimoto, Taichi Akase, H. Kurasawa et al.· bioRxiv· 0 citations
Summary Transcripts annotated as noncoding can produce peptides, but detecting translation does not establish peptide function. The central problem is to determine which instances of ribosome engagement lead to productive translation, which cancer phenotypes depend on the peptide rather than the source RNA, and which validated products are clinically relevant. This Review addresses that problem through a graded evidence framework that proceeds from transcript architecture and ribosome engagement to endogenous peptide detection and causal attribution to the peptide. Under this standard, validated peptides act at molecular control points where local concentration or enzymatic amplification can reconcile low abundance with biological effect. On that basis, we define the conditions required for actionability: tumor-enriched expression, reproducible endogenous measurement, a defined mechanism, and an intervention matched to that mechanism. Progress therefore depends on causal attribution, not on longer candidate lists.
Da Zhou, Hui-Ling Li, Jieun Yang et al.· iScience· 0 citations
The human genome contains hundreds of thousands of short open reading frames (sORFs), yet their translational products, hereafter referred to as sORF-encoded microproteins (also historically termed micropeptides), have long remained overlooked in conventional genome annotation. Increasing evidence demonstrates that these molecules play critical regulatory roles in human physiology and disease. This narrative review summarizes current knowledge regarding the classification, discovery, translational regulation, and biological significance of sORF-encoded microproteins. Their expression is tightly controlled by epigenetic mechanisms, including DNA methylation, Polycomb-mediated H3K27 trimethylation, and bivalent histone modifications, as well as epitranscriptomic regulation through N6-methyladenosine-mediated translation. Key microproteins such as Humanin, MOTS-c, Myoregulin, DWORF, HOXB-AS3-p, Mitoregulin, and CASIMO1 illustrate the diverse roles of sORF-derived peptides in cardiometabolic disease, neurodegeneration, and cancer. The review also highlights emerging therapeutic strategies, including epigenome editing, peptide replacement, and immunopeptidome-based neoantigen targeting. Collectively, the sORF-encoded microproteome represents an epigenetically integrated and functionally significant regulatory layer of the human proteome with major implications for fundamental biology and precision medicine.
Ayman M. Mustafa, Rebaz M. Ali, S. Omar et al.· Epigenomics· 0 citations
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