Loss of ARPC1B impairs cytotoxic T lymphocyte maintenance and cytolytic activity
Lyra O. Randzavola, Katharina Strege, Marie Juzans, Yukako Asano, Jane C. Stinchcombe, Christian M. Gawden-Bone, Matthew N.J. Seaman, Taco W. Kuijpers, Gillian M. Griffiths
Lyra O. Randzavola, Katharina Strege, Marie Juzans, Yukako Asano, Jane C. Stinchcombe, Christian M. Gawden-Bone, Matthew N.J. Seaman, Taco W. Kuijpers, Gillian M. Griffiths
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Research Article Cell biology Immunology

Loss of ARPC1B impairs cytotoxic T lymphocyte maintenance and cytolytic activity

Abstract

CD8 cytotoxic T lymphocytes (CTLs) rely on rapid reorganization of the branched F-actin network to drive the polarized secretion of lytic granules, initiating target cell death during the adaptive immune response. Branched F-actin is generated by the nucleation factor actin-related protein 2/3 (Arp2/3) complex. Patients with mutations in the actin-related protein complex 1B (ARPC1B) subunit of Arp2/3 show combined immunodeficiency, with symptoms of immune dysregulation, including recurrent viral infections and reduced CD8+ T cell count. Here, we show that loss of ARPC1B led to loss of CTL cytotoxicity, with the defect arising at 2 different levels. First, ARPC1B is required for lamellipodia formation, cell migration, and actin reorganization across the immune synapse. Second, we found that ARPC1B is indispensable for the maintenance of TCR, CD8, and GLUT1 membrane proteins at the plasma membrane of CTLs, as recycling via the retromer and WASH complexes was impaired in the absence of ARPC1B. Loss of TCR, CD8, and GLUT1 gave rise to defects in T cell signaling and proliferation upon antigen stimulation of ARPC1B-deficient CTLs, leading to a progressive loss of CD8+ T cells. This triggered an activation-induced immunodeficiency of CTL activity in ARPC1B-deficient patients, which could explain the susceptibility to severe and prolonged viral infections.

Authors

Lyra O. Randzavola, Katharina Strege, Marie Juzans, Yukako Asano, Jane C. Stinchcombe, Christian M. Gawden-Bone, Matthew N.J. Seaman, Taco W. Kuijpers, Gillian M. Griffiths

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Figure 6

Absence of ARPC1B affects CTL effector functions.

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Absence of ARPC1B affects CTL effector functions. (A and B) Percentage l...
(A and B) Percentage lysis of P815 (after 2 hours) by HD or ARPC1B-deficient patient hCTLs at the indicated effector-to-target ratios in the presence (A) or absence (B) of IL-2. (C) Percentage lysis of P815-NucLight Red targets over time measured by IncuCyte killing assay using HD or ARPC1B-deficient patient hCTLs that have been deprived of IL-2 for 16 hours. (D) Western blot analysis of p-ERK1/2 and ERK1/2 expression in HD and ARPC1B-deficient patient hCTLs stimulated for 15 minutes with plate-bound anti-CD3 antibody at the indicated concentrations. Numbers indicate the fold change (ratio) of p-ERK2 expression following stimulation in ARPC1B-deficient patient and HD after normalization to total ERK2 expression. (E) Representative flow cytometry plot and quantitation (F) of LAMP1-PE (CD107a) uptake in HD and ARPC1B-deficient patient hCTLs (gated on CD8+ cells) after 2 hours incubation under the indicated conditions. (G and H) Immunoblot of granzyme B (G) and ARPC1A (H) in HD and ARPC1B-deficient patient in the presence or absence of IL-2. MW markers in kD. (A–C) Data are shown as mean of 3 independent experiments; error bars represent SEM. (D, G, H) Data are representative of 4 independent experiments.