Identification of Novel Interaction between ADAM17 (a Disintegrin and Metalloprotease 17) and Thioredoxin-1

Author(s):Show less -

Aragao, Annelize Z. B. ^[1] ; Nogueira, Maria Luiza C. ^[1] ; Granato, Daniela C. ^[1] ; Simabuco, Fernando M. ^[1] ; Honorato, Rodrigo V. ^[1] ; Hoffman, Zaira ^[2] ; Yokoo, Sami ^[1] ; Laurindo, Francisco R. M. ^[3] ; Squina, Fabio M. ^[2] ; Zeri, Ana Carolina M. ^[1] ; Oliveira, Paulo S. L. ^[1] ; Sherman, Nicholas E. ^[4] ; Paes Leme, Adriana F. ^[1]

Affiliation

^[1] CNPEM, Lab Espectrometria Massas, Lab Nacl Biociencias, LNBio, BR-13083970 Campinas, SP - Brazil

^[2] CNPEM, Lab Nacl Ciencia & Tecnol Bioetanol, CTBE, BR-13083970 Campinas, SP - Brazil

^[3] Univ Sao Paulo, Inst Coracao, BR-05403000 Sao Paulo - Brazil

^[4] Univ Virginia, WM Keck Biomed Mass Spectrometry Lab, Charlottesville, VA 22908 - USA

• Document type: Journal article
• Source: Journal of Biological Chemistry; v. 287, n. 51, p. 43071-43082, DEC 14 2012.
• Web of Science Citations: 22

Abstract

ADAM17, which is also known as TNF alpha-converting enzyme, is the major sheddase for the EGF receptor ligands and is considered to be one of the main proteases responsible for the ectodomain shedding of surface proteins. How a membrane-anchored proteinase with an extracellular catalytic domain can be activated by inside-out regulation is not completely understood. We characterized thioredoxin-1 (Trx-1) as a partner of the ADAM17 cytoplasmic domain that could be involved in the regulation of ADAM17 activity. We induced the overexpression of the ADAM17 cytoplasmic domain in HEK293 cells, and ligands able to bind this domain were identified by MS after protein immunoprecipitation. Trx-1 was also validated as a ligand of the ADAM17 cytoplasmic domain and full-length ADAM17 recombinant proteins by immunoblotting, immunolocalization, and solid phase binding assay. In addition, using nuclear magnetic resonance, it was shown in vitro that the titration of the ADAM17 cytoplasmic domain promotes changes in the conformation of Trx-1. The MS analysis of the cross-linked complexes showed cross-linking between the two proteins by lysine residues. To further evaluate the functional role of Trx-1, we used a heparin-binding EGF shedding cell model and observed that the overexpression of Trx-1 in HEK293 cells could decrease the activity of ADAM17, activated by either phorbol 12-myristate 13-acetate or EGF. This study identifies Trx-1 as a novel interaction partner of the ADAM17 cytoplasmic domain and suggests that Trx-1 is a potential candidate that could be involved in ADAM17 activity regulation. (AU)

• FAPESP's process: 11/02267-9 - Identifying ADAM17 partners and mapping the domains responsible for its interaction with other proteins.
• Grantee: Daniela Campos Granato
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 09/54067-3 - Acquisition of a mass spectrometer coupled to a liquid chromatography system for increasing the capacity to meet the needs of users and for making new technologies available in the Laboratory of Mass Spectrometry
• Grantee: Adriana Franco Paes Leme
• Support Opportunities: Multi-user Equipment Program
• FAPESP's process: 10/19278-0 - Study of regulation of ADAMs in oral cancer
• Grantee: Adriana Franco Paes Leme
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 10/15675-5 - Effects of modulation of the membrane metalloproteinase ADAM-17 in the extracelular subproteome using oral cancer as a model
• Grantee: Fernando Moreira Simabuco
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 09/18301-1 - Study of the regulation of recombinant ADAM-17 by determining phosphorylation sites of the cytoplasmic domain and its ligands in normal and cancer cells.
• Grantee: Annelize Zambon Barbosa Aragão
• Support Opportunities: Scholarships in Brazil - Doctorate