RESEARCH ARTICLE Proteomic analysis of the cellular responses induced in uninfected immune cells by cell-expressed X4 HIV-1 envelope Laurence Molina 1 * , Marina Grimaldi 2 * , Véronique Robert-Hebmann 2 , Lucile Espert 2 , Mihayl Varbanov 2 , Christian Devaux 2 , Claude Granier 1 and Martine Biard-Piechaczyk 2 1 Centre National de la Recherche Scientifique (CNRS) FRE3009, Complex System Modelling and Engineering for Diagnostic, Faculté de Pharmacie, Montpellier, France 2 Centre d’études d’agents Pathogènes et Biotechnologies pour la Santé (CPBS), UMR5236 CNRS/UM1-UM2, Institut de Biologie, Montpellier, France HIV-1 envelope gp120 and gp41 glycoproteins (Env), expressedat the cell surface, induce unin- fected CD4 T-cell death, but the molecular mechanisms leading to this demise are still largely unknown. To better understand these events, we analyzed by a proteomic approach the differential protein expression profile oftwo types of uninfected immune cells after their coculture for 1–3 days with cells that express, or not, Env. First, umbilical cord blood mononuclear cells (UCBMCs) were used to approach the in vivo situation, i.e., blood uninfected naive cells that encounter infected cells. Second, we used the A2.01/CD4.403 T-cell line expressing wild type CXCR4 and a truncated form of CD4 that still undergoes Env-mediated apoptosis, independently of CD4 signaling. After coculture with cells expressing Env, 35 and 39 proteins presenting an altered expression in UCBMCs and the A2.01/CD4.403 T-cell line, respectively, were identified by mass-spectrometry. Whatever the cell type analyzed, the majority of these proteins are involved in degradation processes, redox home- ostasis, metabolism and cytoskeleton dynamics, and linked to mitochondrial functions. This study provides new insights into the events that sequentially occur in bystander T lymphocytes after contact with HIV-infected cells and leading, finally, to apoptotic cell death. Received: March 28, 2007 Revised: May 2, 2007 Accepted: May 21, 2007 Keywords: Cell death / HIV-1 / Mitochondria / Reactive oxygen species 3116 Proteomics 2007, 7, 3116–3130 1 Introduction The development of AIDS in HIV-infected patients is char- acterized by a progressive decline in the number of immune cells [1]. Despite intensive studies, the mechanisms through which HIV-1 infection induces cell death remain largely unknown. Apoptosis, however, is one of the processes likely responsible for T-cell destruction in HIV-infected patients [2– 4]. Indeed, there is a correlation between the extent of apop- tosis and disease progression [5] and highly active anti- retroviral therapy is associated with a lower level of CD4 T- cell apoptosis in HIV-infected patients [6]. Furthermore, bystander uninfected immune cells are the main targets for HIV-induced apoptosis since the degree of cell loss largely exceeds the number of infected cells and the vast majority of Correspondence: Dr. Martine Biard-Piechaczyk, CPBS, UMR5236 CNRS/UM1-UM2, Institut de Biologie, 4, Bd Henri IV, CS 69033, 34965 Montpellier Cedex 2, France E-mail: martine.biard@univ-montp1.fr Fax: 133-4-67-60-44-20 Abbreviations: CA-1, carbonic anhydrase I; CA-2, carbonic anhy- drase II; C1QBP, complement component 1, Q subcomponent binding protein, mitochondrial; DDAH1, NG,NG-dimethylarginine dimethylaminohydrolase 1; DHR, dihydrorhodamine; eIF-3â, eukaryotic translation initiation factor 3 subunit 2; eIF-4A-II, eukar- yotic initiation factor 4A-II; Env, HIV-1 envelope gp120 and gp41 glycoproteins; NPM, nucleophosmin; PEBP-1, phosphatidyl- ethanolamine-binding protein 1; PHGDH, D-3-phosphoglycerate dehydrogenase; PRDX, peroxiredoxin; PSB4, proteasome subunit b Type 4; SLP-2, stomatin-like protein 2; TEM, transmission elec- tron microscopy; UCBMCs, umbilical cord blood mononuclear cells; UCH-L1, ubiquitin carboxy-terminal hydrolase isozyme L1; UQCRC1, ubiquinol-cytochrome C reductase complex core pro- tein 1; X4, CXCR4-dependent HIV-1 strains * Both these authors contributed equally to this work. DOI 10.1002/pmic.200700306  2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.proteomics-journal.com