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Updated: Jun 4, 2025
A Streamlined Approach for Mass Spectrometry-Based Proteomics Using Selected Tissue Regions
Published on: April 18, 2025
Mass Spectrometry-Based Workflow for the Identification and Quantification of Alternative and Canonical Proteins in
Clémence Guillon1, Carole Pichereaux2,3,4, Ikrame Lazar5
1Laboratoire de Recherche en Sciences Végétales (LRSV), CNRS/UT3/INPT, 31320 Auzeville-Tolosane, France.
Abstract:
The identification of small proteins and proteins produced from unannotated open reading frames (called alternative proteins or AltProts) has changed our vision of the proteome and has attracted more and more attention from the scientific community. Despite several studies investigating particular AltProts in diseases and demonstrating their importance in such context, we are still missing data on their expression and functions in many pathologies. Among these, pancreatic ductal adenocarcinoma (PDAC) is a particularly relevant case to study alternative proteins. Indeed, late detection of this disease, notably due to the lack of reliable biomarkers of early-stage PDAC, and the fact that tumors rapidly develop resistance to most of the treatments used in the clinics warrant the exploration of new repertoires of molecules. In the present article, we aim to investigate the alternative proteome of pancreatic cancer cell lines as a first attempt to decipher the expression of AltProts in PDAC. Thanks to a combined data-dependent and data-independent acquisition mass spectrometry workflow, we were able to identify tryptic peptides matching 113 AltProts in a panel of 6 cell lines. In addition, we identified AltProts differentially expressed between pancreatic cancer cell lines and other cells (HeLa and HEK293T). Finally, mining the TCGA and Gtex databases showed that the corresponding transcripts encoding several AltProts we identified are differentially expressed between PDAC tumors and normal tissues and are correlated with the patient's survival.
Insights
Alternative proteins (AltProts) from unannotated genes are increasingly recognized. This study identifies AltProts in pancreatic cancer cell lines, revealing potential diagnostic biomarkers and therapeutic targets for pancreatic ductal adenocarcinoma.
Area of Science:
- Proteomics
- Cancer Biology
- Biochemistry
Background:
- Alternative proteins (AltProts) from unannotated open reading frames are gaining attention in the scientific community.
- Despite their known roles in some diseases, comprehensive data on AltProt expression and function in pancreatic ductal adenocarcinoma (PDAC) is lacking.
- PDAC's late detection and treatment resistance highlight the need for novel molecular targets.
Purpose of the Study:
- To investigate the alternative proteome of pancreatic cancer cell lines.
- To identify novel AltProts expressed in PDAC.
- To explore the potential of AltProts as biomarkers for PDAC.
Main Methods:
- Utilized a combined data-dependent and data-independent acquisition mass spectrometry workflow.
- Analyzed a panel of 6 pancreatic cancer cell lines.
- Compared AltProt expression between cancer and non-cancer cell lines (HeLa, HEK293T).
- Mined TCGA and Gtex databases for transcript expression and patient survival correlations.
Main Results:
- Identified 113 AltProts in pancreatic cancer cell lines.
- Discovered differentially expressed AltProts between pancreatic cancer cells and other cell types.
- Found that transcripts encoding several identified AltProts are differentially expressed in PDAC tumors versus normal tissues.
- Observed correlations between AltProt transcript expression and patient survival.
Conclusions:
- This study provides a first comprehensive look at the alternative proteome in PDAC cell lines.
- Identified AltProts represent potential novel biomarkers for early PDAC detection.
- AltProts may offer new therapeutic avenues for pancreatic cancer treatment.
- Further research into AltProt functions in PDAC is warranted.
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