Slp1 and Slp2-a Localize to the Plasma Membrane of CTL and Contribute to Secretion from the Immunological Synapse

• Published in: Traffic (Copenhagen, Denmark) Vol. 9; no. 4; pp. 446 - 457
• Main Authors: Holt, Oliver, Kanno, Eiko, Bossi, Giovanna, Booth, Sarah, Daniele, Tiziana, Santoro, Alessandra, Arico, Maurizio, Saegusa, Chika, Fukuda, Mitsunori, Griffiths, Gillian M
• Format: Journal Article
• Language: English
• Published: Oxford, UK Oxford, UK : Blackwell Publishing Ltd 01.04.2008; Blackwell Publishing Ltd
• Subjects: Animals; Blood Proteins - genetics; Blood Proteins - immunology; Calpain - genetics; Calpain - metabolism; Cell Membrane - metabolism; CTL; Humans; immunological synapse; Membrane Proteins - genetics; Membrane Proteins - immunology; Mice; Mice, Knockout; Nerve Tissue Proteins; Original; Peptide Hydrolases - genetics; Peptide Hydrolases - metabolism; Protein Isoforms - genetics; Protein Isoforms - immunology; rab GTP-Binding Proteins - genetics; rab GTP-Binding Proteins - immunology; rab27 GTP-Binding Proteins; Rab27a; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; secretory lysosome; Slp2-a; T-Lymphocytes, Cytotoxic - immunology; Vesicular Transport Proteins
• ISSN: 1398-9219; 1600-0854
• DOI: 10.1111/j.1600-0854.2008.00714.x

Rab27a is required for polarized secretion of lysosomes from cytotoxic T lymphocytes (CTLs) at the immunological synapse. A series of Rab27a-interacting proteins have been identified; however, only Munc13-4 has been found to be expressed in CTL. In this study, we screened for expression of the synaptotagmin-like proteins (Slps): Slp1/JFC1, Slp2-a/exophilin4, Slp3-a, Slp4/granuphilin, Slp5 and rabphilin in CTL. We found that both Slp1 and Slp2-a are expressed in CTL. Isoforms of Slp2-a in CTL showed variation of the linker region but conserved the C2A and C2B and Slp homology (SHD) domains. Both Slp1 and Slp2-a interact with Rab27a in CTL, and Slp2-a, but not Slp1, is rapidly degraded when Rab27a is absent. Slp2-a contains PEST-like sequences within its linker region, which render it susceptible to degradation. Both Slp1 and Slp2-a localize predominantly to the plasma membrane of both human and mouse CTLs, and we show that Slp2-a can focus tightly at the immunological synapse formed with a target cell. Individual knockouts of either Slp2-a or Slp1 fail to impair CTL-mediated killing of targets; however, overexpression of a dominant-negative construct consisting of the SHD of Slp2-a, which is 56% identical to that of Slp1, reduces target cell death, suggesting that both Slp1 and Slp2-a contribute to secretory lysosome exocytosis from CTL. These results suggest that both Slp1 and Slp2-a may form part of a docking complex, capturing secretory lysosomes at the immunological synapse.