title: Mass spectrometry-based proteomics revealed Glypican-1 as a novel ADAM17 substrate.
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.
  
  
  
  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.
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
authors: Rebeca
authors: R
authors: Kawahara
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
authors: Daniela Campos
authors: DC
authors: Granato
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
authors: Sami
authors: S
authors: Yokoo
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
authors: Romênia Ramos
authors: RR
authors: Domingues
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil.
authors: Daniel Maragno
authors: DM
authors: Trindade
authors: Laboratório Nacional de Biociências, LNBio, CNPEM, Campinas, Brazil. Electronic address: adriana.paesleme@lnbio.cnpem.br.
authors: Adriana Franco
authors: AF
authors: Paes Leme
mesh_terms: D000072198:ADAM17 Protein / Q000378:metabolism*; D002294:Carcinoma, Squamous Cell; D045744:Cell Line, Tumor; D055252:Cell-Derived Microparticles; D053673:Glypicans / Q000378:metabolism*; D006258:Head and Neck Neoplasms; D006801:Humans; D013058:Mass Spectrometry; D045726:Metalloproteases; D011485:Protein Binding; D059748:Proteolysis; D040901:Proteomics / Q000379:methods*; D000077195:Squamous Cell Carcinoma of Head and Neck
chemical_list: D053673:Glypicans; D045726:Metalloproteases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human
keywords: Glypican; Metalloproteinase; Shedding