title: Constitutive clathrin-mediated endocytosis of CTLA-4 persists during T cell activation.
abstract: CTLA-4 is one of the most important negative regulators of the T cell immune response. However, the subcellular distribution of CTLA-4 is unusual for a receptor that interacts with cell surface transmembrane ligands in that CTLA-4 is rapidly internalized from the plasma membrane. It has been proposed that T cell activation can lead to stabilization of CTLA-4 expression at the cell surface. Here we have analyzed in detail the internalization, recycling, and degradation of CTLA-4. We demonstrate that CTLA-4 is rapidly internalized from the plasma membrane in a clathrin- and dynamin-dependent manner driven by the well characterized YVKM trafficking motif. Furthermore, we show that once internalized, CTLA-4 co-localizes with markers of recycling endosomes and is recycled to the plasma membrane. Although we observed limited co-localization of CTLA-4 with lysosomal markers, CTLA-4 was nonetheless degraded in a manner inhibited by lysosomal blockade. T cell activation stimulated mobilization of CTLA-4, as judged by an increase in cell surface expression; however, this pool of CTLA-4 continued to endocytose and was not stably retained at the cell surface. These data support a model of trafficking whereby CTLA-4 is constitutively internalized in a ligand-independent manner undergoing both recycling and degradation. Stimulation of T cells increases CTLA-4 turnover at the plasma membrane; however, CTLA-4 endocytosis continues and is not stabilized during activation of human T cells. These findings emphasize the importance of clathrin-mediated endocytosis in regulating CTLA-4 trafficking throughout T cell activation.
authors.affiliation: MRC Centre for Immune Regulation, School of Immunity and Infection, University of Birmingham Medical School, University of Birmingham, Birmingham, United Kingdom.
authors.forename: Omar S
authors.initials: OS
authors.lastname: Qureshi
authors.forename: Satdip
authors.initials: S
authors.lastname: Kaur
authors.forename: Tie Zheng
authors.initials: TZ
authors.lastname: Hou
authors.forename: Louisa E
authors.initials: LE
authors.lastname: Jeffery
authors.forename: Natalie S
authors.initials: NS
authors.lastname: Poulter
authors.forename: Zoe
authors.initials: Z
authors.lastname: Briggs
authors.forename: Rupert
authors.initials: R
authors.lastname: Kenefeck
authors.forename: Anna K
authors.initials: AK
authors.lastname: Willox
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Royle
authors.forename: Joshua Z
authors.initials: JZ
authors.lastname: Rappoport
authors.forename: David M
authors.initials: DM
authors.lastname: Sansom
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism*; D016466:CHO Cells; D060908:CTLA-4 Antigen / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism; D006224:Cricetinae; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D006801:Humans; D008213:Lymphocyte Activation; D021381:Protein Transport
chemical_list: D060908:CTLA-4 Antigen; D002966:Clathrin