Stephane A Laporte

Publications (76) View all

• Article: T Cell-Induced Airway Smooth Muscle Cell Proliferation via the Epidermal Growth Factor Receptor.

  Saba Al Heialy, Paul-Andre Risse, Melissa A Zeroual, Horia N Roman, Kimitake Tsuchiya, Sana Siddiqui, Stephane A Laporte, James G Martin
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  ABSTRACT: Allergic asthma is a heterogeneous disease with no curative therapies. T cells infiltrate the airway smooth muscle (ASM) layer and may be implicated in airway remodeling and the increase of ASM mass, a cardinal feature of asthma. The mechanism by which CD4+ T cells drive airway remodeling remains unknown. The objective of this study was to determine the T cell-mediated mechanism of ASM cell proliferation. We hypothesized that CD4+ T cells adhere to ASM cells via CD44 and induce ASM cell proliferation through the activation of the epidermal growth factor receptor (EGFR). A co-culture model showed that contact of antigen-stimulated CD4+ T cells with ASM cells induces high levels of EGFR ligand expression in CD4+ T cells and activation of matrix metalloprotease (MMP)-9, required for the shedding of EGFR ligands. Inhibition of EGFR and MMP-9 prevented the increase of ASM cell proliferation following co-culture. The hyaluronan receptor CD44 is the dominant mediator of tight adherence of T cells to ASM, is colocalized with MMP-9 on the cell surface and neutralization of CD44 prevents ASM cell hyperplasia. These data provide a novel mechanism by which antigen-stimulated CD4+ T cells induce remodeling indicative of a direct trophic role for CD4+ T cells.
  American Journal of Respiratory Cell and Molecular Biology 05/2013; · 5.13 Impact Factor
• Article: GPCR heterodimers- asymmetries in ligand binding and signalling output offer new targets for drug discovery.

  Eugénie Goupil, Stéphane A Laporte, Terence E Hébert
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  ABSTRACT: Dimers of G protein-coupled receptors (GPCRs) have held the imagination of researchers for almost 20 years. However, only recently has their value as potentially novel drug targets been increased significantly, and primarily in the context of GPCR heterodimers. The view of receptor heterodimers as allosteric machines has transformed the way we understand structural and functional asymmetries inherent in their organization. These asymmetries alter both signalling output and how they might be targetted pharmacologically. The article in this issue of BJP by Siddiquee and colleagues highlights our growing understanding of such asymmetries and their implications. They show that heterodimers of the angiotensin II AT1 receptor and the apelin receptor recognize and respond to their respective ligands in distinct ways from the parent receptors expressed alone. Further, they demonstrate asymmetric allosteric effects in the context of the heterodimer that may have significant implications for our understanding of such receptor complexes.
  British Journal of Pharmacology 11/2012; · 4.41 Impact Factor
• Article: C5a- and ASP-mediated C5L2 activation, endocytosis and recycling are lost in S323I-C5L2 mutation.

  Wei Cui, May Simaan, Stephane Laporte, Robert Lodge, Katherine Cianflone
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  ABSTRACT: C5L2, a G-protein-coupled receptor (GPCR), has been identified as an ASP (C3adesArg) and C5a receptor. Controversy exists regarding both ligand binding and functionality. ASP activation of C5L2 is proposed to regulate fat storage. C5L2 is also proposed as a decoy receptor for C5a, an inflammatory mediator, based on absence of Ca(2+) or chemotaxis changes. (i) to evaluate C5L2 receptor activation and recycling using recombinant ASP (rASP) and rC5a and (ii) assess receptor trafficking of S323I-C5L2 mutation previously identified in a family and demonstrated to have altered functionality. stably transfected C5L2-HEK cells were sorted using fluorescent-ASP (Fluos-ASP) binding. Following 2-h serum-free pretreatment, C5L2 was typically localized to the cell-surface. beta-Arrestin-2-GFP transiently transfected C5L2-HEK cells demonstrated rASP and rC5a-dependent beta-arrestin-2-GFP translocation, which showed time-dependent intracellular colocalization with C5L2. Without ligand or C5L2 transfection, no translocation was identified at any time point. Ligand-dependent (rASP and rC5a) C5L2 endocytosis was time-dependent with a 1-h nadir, and was clathrin- and cholesterol-dependent. Transiently transfected Rab-GFP proteins (Rabs 5, 7 and 11) demonstrated time-dependent colocalization of Rab5, Rab7, and Rab11 with C5L2. In contrast to C5L2, a large proportion of stably transfected S323I-C5L2 did not localize to the cell-surface. While S323I-C5L2 was competent for Fluos-ASP and (125)I-ASP binding, although at a reduced level, there was no ligand-mediated receptor phosphorylation. Further, there was no ligand-mediated activation of beta-arrestin-2-GFP translocation, and no downstream functional activation of glucose transport or triglyceride synthesis. C5L2 is a functional metabolic receptor, and serine 323 is important for ASP induced functionality.
  Molecular Immunology 08/2009; 46(15):3086-98. · 2.90 Impact Factor
• Article: The stability of the G protein-coupled receptor-beta-arrestin interaction determines the mechanism and functional consequence of ERK activation.

  Akira Tohgo, Eric W Choy, Diane Gesty-Palmer, Kristen L Pierce, Stephane Laporte, Robert H Oakley, Marc G Caron, Robert J Lefkowitz, Louis M Luttrell
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  ABSTRACT: By binding to agonist-activated G protein-coupled receptors (GPCRs), beta-arrestins mediate homologous receptor desensitization and endocytosis via clathrin-coated pits. Recent data suggest that beta-arrestins also contribute to GPCR signaling by acting as scaffolds for components of the ERK mitogen-activated protein kinase cascade. Because of these dual functions, we hypothesized that the stability of the receptor-beta-arrestin interaction might affect the mechanism and functional consequences of GPCR-stimulated ERK activation. In transfected COS-7 cells, we found that angiotensin AT1a and vasopressin V2 receptors, which form stable receptor-beta-arrestin complexes, activated a beta-arrestin-bound pool of ERK2 more efficiently than alpha 1b and beta2 adrenergic receptors, which form transient receptor-beta-arrestin complexes. We next studied chimeric receptors in which the pattern of beta-arrestin binding was reversed by exchanging the C-terminal tails of the beta2 and V2 receptors. The ability of the V2 beta 2 and beta 2V2 chimeras to activate beta-arrestin-bound ERK2 corresponded to the pattern of beta-arrestin binding, suggesting that the stability of the receptor-beta-arrestin complex determined the mechanism of ERK2 activation. Analysis of covalently cross-linked detergent lysates and cellular fractionation revealed that wild type V2 receptors generated a larger pool of cytosolic phospho-ERK1/2 and less nuclear phospho-ERK1/2 than the chimeric V2 beta 2 receptor, consistent with the cytosolic retention of beta-arrestin-bound ERK. In stably transfected HEK-293 cells, the V2 beta 2 receptor increased ERK1/2-mediated, Elk-1-driven transcription of a luciferase reporter to a greater extent than the wild type V2 receptor. Furthermore, the V2 beta 2, but not the V2 receptor, was capable of eliciting a mitogenic response. These data suggest that the C-terminal tail of a GPCR, by determining the stability of the receptor-beta-arrestin complex, controls the extent of beta-arrestin-bound ERK activation, and influences both the subcellular localization of activated ERK and the physiologic consequences of ERK activation.
  Journal of Biological Chemistry 03/2003; 278(8):6258-67. · 4.77 Impact Factor
• Article: N-terminal tyrosine modulation of the endocytic adaptor function of the beta-arrestins.

  Sébastien Marion, Gregory B Fralish, Stéphane Laporte, Marc G Caron, Larry S Barak
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  ABSTRACT: The highly homologous beta-arrestin1 and -2 adaptor proteins play important roles in the function of G protein-coupled receptors. Either beta-arrestin variant can function as a molecular chaperone for clathrin-mediated receptor internalization. This role depends primarily upon two distinct, contiguous C-terminal beta-arrestin motifs recognizing clathrin and the beta-adaptin subunit of AP2. However, a molecular basis is lacking to explain the different endocytic efficacies of the two beta-arrestin isoforms and the observation that beta-arrestin N-terminal substitution mutants can act as dominant negative inhibitors of receptor endocytosis. Despite the near identity of the beta-arrestins throughout their N termini, sequence variability is present at a small number of residues and includes tyrosine to phenylalanine substitutions. Here we show that corresponding N-terminal (Y/F)VTL sequences in beta-arrestin1 and -2 differentially regulate mu-adaptin binding. Our results indicate that the beta-arrestin1 Tyr-54 lessens the interaction with mu-adaptin and moreover is a Src phosphorylation site. A gain of endocytic function is obtained with the beta-arrestin1 Y54F substitution, which improves both the beta-arrestin1 interaction with mu-adaptin and the ability to enhance beta2-adrenergic receptor internalization. These data indicate that beta-arrestin2 utilizes mu-adaptin as an endocytic partner, and that the inability of beta-arrestin1 to sustain a similar degree of interaction with mu-adaptin may result from coordination of Tyr-54 by neighboring residues or its modification by Src kinase. Additionally, these naturally occurring variations in beta-arrestins may also differentially regulate the composition of the signaling complexes organized on the receptor.
  Journal of Biological Chemistry 07/2007; 282(26):18937-44. · 4.77 Impact Factor