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Sustained signalling at the immune synapse (IS) requires the synaptic delivery of recycling endosome-associated T cell antigen receptors (TCRs). IFT20, a component of the intraflagellar transport system, controls TCR recycling to the IS as a complex with IFT57 and IFT88. Here, we used quantitative mass spectrometry to identify additional interaction partners of IFT20 in Jurkat T cells. In addition to IFT57 and IFT88, the analysis revealed new binding partners, including IFT54 (also known as TRAF3IP1), GMAP-210 (also known as TRIP11), Arp2/3 complex subunit-3 (ARPC3), COP9 signalosome subunit-1 (CSN1, also known as GPS1) and ERGIC-53 (also known as LMAN1). A direct interaction between IFT20 and both IFT54 and GMAP-210 was confirmed in pulldown assays. Confocal imaging of antigen-specific conjugates using T cells depleted of these proteins by RNA interference showed that TCR accumulation and phosphotyrosine signalling at the IS were impaired in the absence of IFT54, ARPC3 or ERGIC-53. Similar to in IFT20-deficient T cells, this defect resulted from a reduced ability of endosomal TCRs to polarize to the IS despite a correct translocation of the centrosome towards the antigen-presenting cell contact. Our data underscore the traffic-related role of an IFT20 complex that includes components of the intracellular trafficking machinery in IS assembly.

Original publication

DOI

10.1242/jcs.200006

Type

Journal article

Journal

Journal of cell science

Publication Date

03/2017

Volume

130

Pages

1110 - 1121

Addresses

Department of Life Sciences, University of Siena, Siena 53100, Italy.

Keywords

T-Lymphocytes, Jurkat Cells, Microtubule-Organizing Center, Humans, Carrier Proteins, Receptors, Transferrin, Microtubule-Associated Proteins, Mannose-Binding Lectins, Membrane Proteins, Receptors, Antigen, T-Cell, Nuclear Proteins, Lymphocyte Activation, Endocytosis, Protein Binding, Actin-Related Protein 2-3 Complex, Mass Spectrometry, Immunological Synapses, HEK293 Cells, Protein Interaction Maps