Components of the intraflagellar transport (IFT) system that regulates the assembly of the primary cilium are co-opted by the non-ciliated T cell to orchestrate polarized endosome recycling and to sustain signaling during immune synapse formation. Here, we investigated the potential role of Bardet-Biedl syndrome 1 protein (BBS1), an essential core component of the BBS complex that cooperates with the IFT system in ciliary protein trafficking, in the assembly of the T cell synapse. We demonstrated that BBS1 allows for centrosome polarization towards the immune synapse. This function is achieved through the clearance of centrosomal F-actin and its positive regulator WASH1 (also known as WASHC1), a process that we demonstrated to be dependent on the proteasome. We show that BBS1 regulates this process by coupling the 19S proteasome regulatory subunit to the microtubule motor dynein for its transport to the centrosome. Our data identify the ciliopathy-related protein BBS1 as a new player in T cell synapse assembly that functions upstream of the IFT system to set the stage for polarized vesicular trafficking and sustained signaling. This article has an associated First Person interview with the first author of the paper.

Department of Life Sciences University of Siena 53100 Siena Italy

Department of Medical Biotechnologies University of Siena 53100 Siena Italy

Institute of Molecular Genetics of the Czech Academy of Sciences 142 20 Prague Czech Republic

Kennedy Institute of Rheumatology University of Oxford OX3 7FY Oxford UK

Zobrazit více v PubMed

Alcover A, Alarcon B, Di Bartolo V. Cell Biology of T Cell Receptor Expression and Regulation. Annu Rev Immunol. 2018;36:103–125. PubMed

Baldari CT, Macchia G, Telford JL. Interleukin-2 promoter activation in T-cells expressing activated Ha-ras. J Biol Chem. 1992;267:4289–91. PubMed

Baldari CT, Rosenbaum J. Intraflagellar transport: it’s not just for cilia anymore. Curr Opin Cell Biol. 2010;22:75–80. PubMed PMC

Bard JAM, Goodall EA, Greene ER, Jonsson E, Dong KC, Martin A. Structure and Function of the 26S Proteasome. Annu Rev Biochem. 2018;87:697–724. PubMed PMC

Bello-Gamboa A, Velasco M, Moreno S, Herranz G, Ilie R, Huetos S, Davila S, Sancheza A, De La Serna JB, Calvo V, et al. Actin reorganization at the centrosomal area and the immune synapse regulates polarized secretory traffic of multivesicular bodies in T lymphocytes. Journal of Extracellular Vesicles. 2020;9 PubMed PMC

Bjornson-Hooper ZB, Fragiadakis GK, Spitzer MH, Madhireddy D, McIlwain D, Nolan GP. A comprehensive atlas of immunological differences between humans, mice and non-human primates. bioRxiv. 2019:574160. doi: 10.1101/574160. PubMed DOI PMC

Bonello G, Blanchard N, Montoya MC, Aguado E, Langlet C, He HT, Nunez-Cruz S, Malissen M, Sanchez-Madrid F, Olive D, et al. Dynamic recruitment of the adaptor protein LAT: LAT exists in two distinct intracellular pools and controls its own recruitment. J Cell Sci. 2004;117:1009–16. PubMed

Bouchet J, Del Rio-Iniguez I, Lasserre R, Aguera-Gonzalez S, Cuche C, Danckaert A, McCaffrey MW, Di Bartolo V, Alcover A. Rac1-Rab11-FIP3 regulatory hub coordinates vesicle traffic with actin remodeling and T-cell activation. EMBO J. 2016;35:1160–74. PubMed PMC

Burakov AV, Nadezhdina ES. Association of nucleus and centrosome: magnet or velcro? Cell Biol Int. 2013;37:95–104. PubMed

Bustos-Moran E, Blas-Rus N, Martin-Cofreces NB, Sanchez-Madrid F. Orchestrating Lymphocyte Polarity in Cognate Immune Cell-Cell Interactions. Int Rev Cell Mol Biol. 2016;327:195–261. PubMed PMC

Cassioli C, Baldari CT. A Ciliary View of the Immunological Synapse. Cells. 2019;8 PubMed PMC

Chicaybam L, Sodre AL, Curzio BA, Bonamino MH. An efficient low cost method for gene transfer to T lymphocytes. PLoS One. 2013;8:e60298. PubMed PMC

Choudhuri K, Llodra J, Roth EW, Tsai J, Gordo S, Wucherpfennig KW, Kam LC, Stokes DL, Dustin ML. Polarized release of T-cell-receptor-enriched microvesicles at the immunological synapse. Nature. 2014;507:118–23. PubMed PMC

Compeer EB, Kraus F, Ecker M, Redpath G, Amiezer M, Rother N, Nicovich PR, Kapoor-Kaushik N, Deng Q, Samson GPB, et al. A mobile endocytic network connects clathrin-independent receptor endocytosis to recycling and promotes T cell activation. Nat Commun. 2018;9:1597. PubMed PMC

Das V, Nal B, Dujeancourt A, Thoulouze MI, Galli T, Roux P, Dautry-Varsat A, Alcover A. Activation-induced polarized recycling targets T cell antigen receptors to the immunological synapse; involvement of SNARE complexes. Immunity. 2004;20:577–88. PubMed

de la Roche M, Asano Y, Griffiths GM. Origins of the cytolytic synapse. Nat Rev Immunol. 2016;16:421–32. PubMed

de la Roche M, Ritter AT, Angus KL, Dinsmore C, Earnshaw CH, Reiter JF, Griffiths GM. Hedgehog signaling controls T cell killing at the immunological synapse. Science. 2013;342:1247–50. PubMed PMC

Desai PB, Stuck MW, Lv B, Pazour GJ. Ubiquitin links smoothened to intraflagellar transport to regulate Hedgehog signaling. J Cell Biol. 2020;219 PubMed PMC

Didier C, Merdes A, Gairin JE, Jabrane-Ferrat N. Inhibition of proteasome activity impairs centrosome-dependent microtubule nucleation and organization. Mol Biol Cell. 2008;19:1220–9. PubMed PMC

Dustin ML. The immunological synapse. Cancer Immunol Res. 2014;2:1023–33. PubMed PMC

Ehrlich LI, Ebert PJ, Krummel MF, Weiss A, Davis MM. Dynamics of p56lck translocation to the T cell immunological synapse following agonist and antagonist stimulation. Immunity. 2002;17:809–22. PubMed

Farina F, Gaillard J, Guerin C, Coute Y, Sillibourne J, Blanchoin L, Thery M. The centrosome is an actin-organizing centre. Nat Cell Biol. 2016;18:65–75. PubMed PMC

Finetti F, Cassioli C, Baldari CT. Transcellular communication at the immunological synapse: a vesicular traffic-mediated mutual exchange. F1000Res. 2017;6:1880. PubMed PMC

Finetti F, Paccani SR, Riparbelli MG, Giacomello E, Perinetti G, Pazour GJ, Rosenbaum JL, Baldari CT. Intraflagellar transport is required for polarized recycling of the TCR/CD3 complex to the immune synapse. Nat Cell Biol. 2009;11:1332–9. PubMed PMC

Finetti F, Patrussi L, Galgano D, Cassioli C, Perinetti G, Pazour GJ, Baldari CT. The small GTPase Rab8 interacts with VAMP-3 to regulate the delivery of recycling T-cell receptors to the immune synapse. J Cell Sci. 2015;128:2541–52. PubMed PMC

Finetti F, Patrussi L, Masi G, Onnis A, Galgano D, Lucherini OM, Pazour GJ, Baldari CT. Specific recycling receptors are targeted to the immune synapse by the intraflagellar transport system. J Cell Sci. 2014;127:1924–37. PubMed PMC

Follit JA, Tuft RA, Fogarty KE, Pazour GJ. The intraflagellar transport protein IFT20 is associated with the Golgi complex and is required for cilia assembly. Mol Biol Cell. 2006;17:3781–92. PubMed PMC

Forsythe E, Beales PL. Bardet-Biedl syndrome. Eur J Hum Genet. 2013;21:8–13. PubMed PMC

Galgano D, Onnis A, Pappalardo E, Galvagni F, Acuto O, Baldari CT. The T cell IFT20 interactome reveals new players in immune synapse assembly. J Cell Sci. 2017;130:1120–1121. PubMed PMC

Gawden-Bone CM, Frazer GL, Richard AC, Ma CY, Strege K, Griffiths GM. PIP5 Kinases Regulate Membrane Phosphoinositide and Actin Composition for Targeted Granule Secretion by Cytotoxic Lymphocytes. Immunity. 2018;49:427–437.:e4. PubMed PMC

Gomez TS, Billadeau DD. A FAM21-containing WASH complex regulates retromer-dependent sorting. Dev Cell. 2009;17:699–711. PubMed PMC

Guo DF, Cui HX, Zhang QH, Morgan DA, Thedens DR, Nishimura D, Grobe JL, Sheffield VC, Rahmouni K. The BBSome Controls Energy Homeostasis by Mediating the Transport of the Leptin Receptor to the Plasma Membrane. Plos Genetics. 2016;12 PubMed PMC

Haeussler M, Schonig K, Eckert H, Eschstruth A, Mianne J, Renaud JB, Schneider-Maunoury S, Shkumatava A, Teboul L, Kent J, et al. Evaluation of off-target and on-target scoring algorithms and integration into the guide RNA selection tool CRISPOR. Genome Biol. 2016;17:148. PubMed PMC

Hao YH, Doyle JM, Ramanathan S, Gomez TS, Jia D, Xu M, Chen ZJ, Billadeau DD, Rosen MK, Potts PR. Regulation of WASH-dependent actin polymerization and protein trafficking by ubiquitination. Cell. 2013;152:1051–64. PubMed PMC

Hsu MT, Guo CL, Liou AY, Chang TY, Ng MC, Florea BI, Overkleeft HS, Wu YL, Liao JC, Cheng PL. Stage-Dependent Axon Transport of Proteasomes Contributes to Axon Development. Dev Cell. 2015;35:418–31. PubMed

Ibanez-Vega J, Del Valle Batalla F, Saez JJ, Soza A, Yuseff MI. Proteasome Dependent Actin Remodeling Facilitates Antigen Extraction at the Immune Synapse of B Cells. Front Immunol. 2019;10:225. PubMed PMC

Jin H, White SR, Shida T, Schulz S, Aguiar M, Gygi SP, Bazan JF, Nachury MV. The conserved Bardet-Biedl syndrome proteins assemble a coat that traffics membrane proteins to cilia. Cell. 2010;141:1208–19. PubMed PMC

Joseph N, Biber G, Fried S, Reicher B, Levy O, Sabag B, Noy E, Barda-Saad M. A conformational change within the WAVE2 complex regulates its degradation following cellular activation. Sci Rep. 2017;7:44863. PubMed PMC

Katsanis N. The oligogenic properties of Bardet-Biedl syndrome. Hum Mol Genet. 2004;13(Spec No 1):R65–71. PubMed

Langousis G, Shimogawa MM, Saada EA, Vashisht AA, Spreafico R, Nager AR, Barshop WD, Nachury MV, Wohlschlegel JA, Hill KL. Loss of the BBSome perturbs endocytic trafficking and disrupts virulence of Trypanosoma brucei. Proc Natl Acad Sci U S A. 2016;113:632–7. PubMed PMC

Larghi P, Williamson DJ, Carpier JM, Dogniaux S, Chemin K, Bohineust A, Danglot L, Gaus K, Galli T, Hivroz C. VAMP7 controls T cell activation by regulating the recruitment and phosphorylation of vesicular Lat at TCR-activation sites. Nat Immunol. 2013;14:723–31. PubMed

Liu YP, Tsai IC, Morleo M, Oh EC, Leitch CC, Massa F, Lee BH, Parker DS, Finley D, Zaghlou NA, et al. Ciliopathy proteins regulate paracrine signaling by modulating proteasomal degradation of mediators. Journal of Clinical Investigation. 2014;124:2059–2070. PubMed PMC

Lui-Roberts WW, Stinchcombe JC, Ritter AT, Akhmanova A, Karakesisoglou I, Griffiths GM. Cytotoxic T lymphocyte effector function is independent of nucleus-centrosome dissociation. Eur J Immunol. 2012;42:2132–41. PubMed PMC

Manders EM, Stap J, Brakenhoff GJ, van Driel R, Aten JA. Dynamics of three-dimensional replication patterns during the S-phase, analysed by double labelling of DNA and confocal microscopy. J Cell Sci. 1992;103(Pt 3):857–62. PubMed

Martin-Cofreces NB, Baixauli F, Lopez MJ, Gil D, Monjas A, Alarcon B, Sanchez-Madrid F. End-binding protein 1 controls signal propagation from the T cell receptor. EMBO J. 2012;31:4140–52. PubMed PMC

Martin-Cofreces NB, Robles-Valero J, Cabrero JR, Mittelbrunn M, Gordon-Alonso M, Sung CH, Alarcon B, Vazquez J, Sanchez-Madrid F. MTOC translocation modulates IS formation and controls sustained T cell signaling. J Cell Biol. 2008;182:951–62. PubMed PMC

Martin-Cofreces NB, Chichon FJ, Calvo E, Torralba D, Bustos-Moran E, Dosil SG, Rojas-Gomez A, Bonzon-Kulichenko E, Lopez JA, Otón J, et al. The chaperonin CCT controls T cell receptor-driven 3D configuration of centrioles. Sci Adv. 2020;6:eabb7242. PubMed PMC

Mestas J, Hughes CCW. Of mice and not men: Differences between mouse and human immunology. Journal of Immunology. 2004;172:2731–2738. PubMed

Mourao A, Christensen ST, Lorentzen E. The intraflagellar transport machinery in ciliary signaling (vol 41, pg 98, 2016) Current Opinion in Structural Biology. 2016;41:255. PubMed

Mourao A, Nager AR, Nachury MV, Lorentzen E. Structural basis for membrane targeting of the BBSome by ARL6. Nature Structural & Molecular Biology. 2014;21:1035–1041. PubMed PMC

Nachury MV. The molecular machines that traffic signaling receptors into and out of cilia. Curr Opin Cell Biol. 2018;51:124–131. PubMed PMC

Nakayama K, Katoh Y. Ciliary protein trafficking mediated by IFT and BBSome complexes with the aid of kinesin-2 and dynein-2 motors. J Biochem. 2018;163:155–164. PubMed

Nath S, Christian L, Tan SY, Ki S, Ehrlich LI, Poenie M. Dynein Separately Partners with NDE1 and Dynactin To Orchestrate T Cell Focused Secretion. J Immunol. 2016;197:2090–2101. PubMed PMC

Nozaki S, Katoh Y, Kobayashi T, Nakayama K. BBS1 is involved in retrograde trafficking of ciliary GPCRs in the context of the BBSome complex. PLoS One. 2018;13:e0195005. PubMed PMC

Obino D, Farina F, Malbec O, Saez PJ, Maurin M, Gaillard J, Dingli F, Loew D, Gautreau A, Yuseff MI, et al. Actin nucleation at the centrosome controls lymphocyte polarity. Nat Commun. 2016;7:10969. PubMed PMC

Onnis A, Baldari CT. Orchestration of Immunological Synapse Assembly by Vesicular Trafficking. Front Cell Dev Biol. 2019;7:110. PubMed PMC

Onnis A, Finetti F, Patrussi L, Gottardo M, Cassioli C, Spano S, Baldari CT. The small GTPase Rab29 is a common regulator of immune synapse assembly and ciliogenesis. Cell Death Differ. 2015;22:1687–99. PubMed PMC

Pedersen LB, Rosenbaum JL. Intraflagellar transport (IFT) role in ciliary assembly, resorption and signalling. Curr Top Dev Biol. 2008;85:23–61. PubMed

Prasai A, Schmidt Cernohorska M, Ruppova K, Niederlova V, Andelova M, Draber P, Stepanek O, Huranova M. The BBSome assembly is spatially controlled by BBS1 and BBS4 in human cells. J Biol Chem. 2020;295:14279–14290. PubMed PMC

Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Genome engineering using the CRISPR-Cas9 system. Nat Protoc. 2013;8:2281–2308. PubMed PMC

Roy NH, Burkhardt JK. The Actin Cytoskeleton: A Mechanical Intermediate for Signal Integration at the Immunological Synapse. Front Cell Dev Biol. 2018;6:116. PubMed PMC

Seaman MN, Gautreau A, Billadeau DD. Retromer-mediated endosomal protein sorting: all WASHed up! Trends Cell Biol. 2013;23:522–8. PubMed PMC

Soares H, Lasserre R, Alcover A. Orchestrating cytoskeleton and intracellular vesicle traffic to build functional immunological synapses. Immunol Rev. 2013;256:118–32. PubMed

Starks RD, Beyer AM, Guo DF, Boland L, Zhang QH, Sheffield VC, Rahmouni K. Regulation of Insulin Receptor Trafficking by Bardet Biedl Syndrome Proteins. Plos Genetics. 2015;11 PubMed PMC

Stephen LA, ElMaghloob Y, McIlwraith MJ, Yelland T, Castro Sanchez P, Roda-Navarro P, Ismail S. The Ciliary Machinery Is Repurposed for T Cell Immune Synapse Trafficking of LCK. Dev Cell. 2018;47:122–132.:e4. PubMed PMC

Stinchcombe JC, Griffiths GM. Communication, the centrosome and the immunological synapse. Philos Trans R Soc Lond B Biol Sci. 2014;369 PubMed PMC

Tobin JL, Beales PL. The nonmotile ciliopathies. Genet Med. 2009;11:386–402. PubMed

Tsang SH, Aycinena ARP, Sharma T. Ciliopathy: Bardet-Biedl Syndrome. Adv Exp Med Biol. 2018;1085:171–174. PubMed

Tsyklauri O, Niederlova V, Forsythe E, Prasai A, Drobek A, Kasparek P, Sparks K, Trachtulec Z, Prochazka J, Sedlacek R, et al. Bardet-Biedl Syndrome ciliopathy is linked to altered hematopoiesis and dysregulated self-tolerance. EMBO Rep. 2021:e50785. PubMed PMC

Vardhana S, Choudhuri K, Varma R, Dustin ML. Essential role of ubiquitin and TSG101 protein in formation and function of the central supramolecular activation cluster. Immunity. 2010;32:531–40. PubMed PMC

Vora SM, Phillips BT. The benefits of local depletion: The centrosome as a scaffold for ubiquitin-proteasome-mediated degradation. Cell Cycle. 2016;15:2124–2134. PubMed PMC

Walczak H, Iwai K, Dikic I. Generation and physiological roles of linear ubiquitin chains. BMC Biol. 2012;10:23. PubMed PMC

Wang J, Fedoseienko F, Chen B, Burstein E, Jia D, Billadeau DD. Endosomal receptor trafficking: Retromer and beyond. Traffic. 2018;19:578–590. PubMed PMC

Watanabe Y, Sasahara Y, Ramesh N, Massaad MJ, Yeng Looi C, Kumaki S, Kure S, Geha RS, Tsuchiya S. T-cell receptor ligation causes Wiskott-Aldrich syndrome protein degradation and F-actin assembly downregulation. J Allergy Clin Immunol. 2013;132:648–655.:e1. PubMed

Wingfield JL, Lechtreck KF, Lorentzen E. Trafficking of ciliary membrane proteins by the intraflagellar transport/BBSome machinery. Essays Biochem. 2018;62:753–763. PubMed PMC

Xu Q, Zhang Y, Wei Q, Huang Y, Li Y, Ling K, Hu J. BBS4 and BBS5 show functional redundancy in the BBSome to regulate the degradative sorting of ciliary sensory receptors. Sci Rep. 2015;5:11855. PubMed PMC

Ye F, Nager AR, Nachury MV. BBSome trains remove activated GPCRs from cilia by enabling passage through the transition zone. J Cell Biol. 2018;217:1847–1868. PubMed PMC

Zucchetti AE, Bataille L, Carpier JM, Dogniaux S, San Roman-Jouve M, Maurin M, Stuck MW, Rios RM, Baldari CT, Pazour GJ, et al. Tethering of vesicles to the Golgi by GMAP210 controls LAT delivery to the immune synapse. Nat Commun. 2019;10:2864. PubMed PMC