Mass spectrometry-based proteomics revealed Glypican-1 as a novel ADAM17 substrate

Autor(es)

Kawahara, Rebeca ; Granato, Daniela Campos ; Yokoo, Sami ; Domingues, Romenia Ramos ; Trindade, Daniel Maragno ; Paes Leme, Adriana Franco

• Tipo de documento: Artigo Científico
• Fonte: JOURNAL OF PROTEOMICS; v. 151, p. 13-pg., 2017-01-16.

Resumo

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. (C) 2016 Elsevier B.V. All rights reserved. (AU)

• Processo FAPESP: 09/54067-3 - EMU: aquisição de um espectrômetro de massas acoplado a cromatografia líquida para permitir ampliar a capacidade de atendimento de usuários e disponibilizar novas tecnologias no Laboratório de Espectrometria de Massas do Centro de Biologia Molecular Estrutural (ABTLUS)
• Beneficiário: Adriana Franco Paes Leme
• Modalidade de apoio: Auxílio à Pesquisa - Programa Equipamentos Multiusuários
• Processo FAPESP: 11/22421-2 - Determinação dos sítios de clivagens de alvos de ADAM-17 recombinante em cultura de células humanas
• Beneficiário: Rebeca Kawahara Sakuma
• Modalidade de apoio: Bolsas no Brasil - Doutorado Direto
• Processo FAPESP: 10/19278-0 - Estudo da regulação de ADAMs em câncer oral
• Beneficiário: Adriana Franco Paes Leme
• Modalidade de apoio: Auxílio à Pesquisa - Jovens Pesquisadores