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Mass spectrometry-based proteomics revealed Glypican-1 as a novel ADAM17 substrate
Rebeca Kawahara1, Daniela Campos Granato1, Sami Yokoo1
1Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
Abstract:
ADAM17 (a disintegrin and metalloproteinase 17) is a plasma membrane metalloprotease involved in proteolytic release of the extracellular domain of many cell surface molecules, a process known as ectodomain shedding. Through this process, ADAM17 is implicated in several aspects of tumor growth and metastasis in a broad range of tumors, including head and neck squamous cell carcinomas (HNSCC). In this study, mass spectrometry-based proteomics approaches revealed glypican-1 (GPC1) as a new substrate for ADAM17, and its shedding was confirmed to be metalloprotease-dependent, induced by a pleiotropic agent (PMA) and physiologic ligand (EGF), and inhibited by marimastat. In addition, immunoblotting analysis of GPC1 in the extracellular media from control and ADAM17shRNA pointed to a direct involvement of ADAM17 in the cleavage of GPC1. Moreover, mass spectrometry-based interactome analysis of GPC1 revealed biological functions and pathways related mainly to cellular movement, adhesion and proliferation, which were events also modulated by up regulation of full length and cleavage GPC1. Altogether, we showed that GPC1 is a novel ADAM17 substrate, thus the function of GPC1 may be modulated by proteolysis signaling.
Biological Significance:
Inhibition of metalloproteases as a therapeutic approach has failed because there is limited knowledge of the degradome of individual proteases as well as the cellular function of cleaved substrates. Using different proteomic techniques, this study uncovered novel substrates that can be modulated by ADAM17 in oral squamous cell carcinoma cell line. Glypican-1 was validated as a novel substrate for ADAM17, with important function in adhesion, proliferation and migration of carcinoma cells. Therefore, this study opens new avenues regarding the proteolysis-mediated function of GPC1 by ADAM17.
Insights
ADAM17 (a disintegrin and metalloproteinase 17) cleaves glypican-1 (GPC1), a novel substrate involved in cancer cell movement and proliferation. This discovery offers new insights into proteolysis signaling in oral squamous cell carcinoma.
Area of Science:
- Biochemistry and Molecular Biology
- Cancer Research
- Proteomics
Background:
- ADAM17 (a disintegrin and metalloproteinase 17) is a metalloprotease crucial for ectodomain shedding of cell surface proteins.
- ADAM17 plays a role in tumor growth and metastasis, particularly in head and neck squamous cell carcinomas (HNSCC).
- Understanding protease substrates and their functions is vital for developing effective metalloprotease-targeted therapies.
Purpose of the Study:
- To identify novel substrates of ADAM17 using mass spectrometry-based proteomics.
- To investigate the role of ADAM17 in the shedding and function of glypican-1 (GPC1).
- To explore the implications of GPC1 modulation by ADAM17 in oral squamous cell carcinoma.
Main Methods:
- Mass spectrometry-based proteomics and interactome analysis.
- Validation of substrate shedding using immunoblotting.
- Functional studies on GPC1 in oral squamous cell carcinoma cell lines.
Main Results:
- Glypican-1 (GPC1) was identified as a novel substrate of ADAM17.
- GPC1 shedding is dependent on ADAM17 activity, induced by PMA and EGF, and inhibited by marimastat.
- GPC1 is involved in cellular adhesion, proliferation, and migration, processes modulated by its shedding.
Conclusions:
- GPC1 is a novel substrate of ADAM17, and its function is modulated by ADAM17-mediated proteolysis.
- This study highlights a new proteolytic signaling pathway involving GPC1 and ADAM17 in cancer.
- Findings open new therapeutic avenues targeting the ADAM17-GPC1 axis in oral squamous cell carcinoma.
