Identification of the cytoplasmic domains of CXCR4 involved in Jak2 and STAT3 phosphorylation.

The chemokine SDF-1alpha transduces G(i)-dependent and -independent signals through CXCR4. Activation of Jak2/STAT3, a G(i)-independent signaling pathway, which plays a major role in survival signals, is known to be activated after SDF-1alpha binding to CXCR4 but the domains of CXCR4 involved in this signaling remain unexplored. Using human embryonic kidney HEK-293 cells stably expressing wild-type or mutated forms of CXCR4, we demonstrated that STAT3 phosphorylation requires the N-terminal part of the third intracellular loop (ICL3) and the tyrosine 157 present at the end of the second intracellular loop (ICL2) of CXCR4. In contrast, neither the conserved Tyr(135) in the DRY motif at the N terminus of ICL2 nor the Tyr(65) and Tyr(76) in the first intracellular loop (ICL1) are involved in this activation. ICL3, which does not contain any tyrosine residues, is needed to activate Jak2. These results demonstrate that two separate domains of CXCR4 are involved in Jak2/STAT3 signaling. The N-terminal part of ICL3 is needed to activate Jak2 after SDF-1alpha binding to CXCR4, leading to phosphorylation of only one cytoplasmic Tyr, present at the C terminus of ICL2, which triggers STAT3 activation. This work has profound implications for the understanding of CXCR4-transduced signaling.

Apoptosis of uninfected cells induced by HIV envelope glycoproteins.

Apoptosis, or programmed cell death, is a key event in biologic homeostasis but is also involved in the pathogenesis of many human diseases including human immunodeficiency virus (HIV) infection. Although multiple mechanisms contribute to the gradual T cell decline that occurs in HIV-infected patients, programmed cell death of uninfected bystander T lymphocytes, including CD4+ and CD8+ T cells, is an important event leading to immunodeficiency. The HIV envelope glycoproteins (Env) play a crucial role in transducing this apoptotic signal after binding to its receptors, the CD4 molecule and a coreceptor, essentially CCR5 and CXCR4. Depending on Env presentation, the receptor involved and the complexity of target cell contact, apoptosis induction is related to death receptor and/or mitochondria-dependent pathways. This review summarizes current knowledge of Env-mediated cell death leading to T cell depletion and clinical complications and covers the sometimes conflicting studies that address the possible mechanisms of T cell death.

Role of the intracellular domains of CXCR4 in SDF-1-mediated signaling.

The CXCR4 chemokine receptor is a G(i) protein-coupled receptor that triggers multiple intracellular signals in response to stromal cell-derived factor 1 (SDF-1), including calcium mobilization and p44/42 extracellular signal-regulated kinases (ERK1/2). Transduced signals lead to cell chemotaxis and are terminated through receptor internalization depending on phosphorylation of the C terminus part of CXCR4. Receptor endocytosis is also required for some receptors to stimulate ERK1/2 and to migrate through a chemokine gradient. In this study, we explored the role played by the 3 intracellular loops (ICL1-3) and the C terminus domain of CXCR4 in SDF-1-mediated signaling by using human embryonic kidney (HEK)-293 cells stably expressing wild-type or mutated forms of CXCR4. ICL3 of CXCR4 is specifically involved in G(i)-dependent signals such as calcium mobilization and ERK activation, but does not trigger CXCR4 internalization after SDF-1 binding, indicating that ERK phosphorylation is independent of CXCR4 endocytosis. Surprisingly, ICL2, with or without the aspartic acid, arginine, and tyrosine (DRY) motif, is dispensable for G(i) signaling. However, ICL2 and ICL3, as well as the C terminus part of CXCR4, are needed to transduce SDF-1-mediated chemotaxis, suggesting that this event involves multiple activation pathways and/or cooperation of several cytoplasmic domains of CXCR4.