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 4

Differential actin structure in hCTLs from ARPC1B-deficient patient.

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Differential actin structure in hCTLs from ARPC1B-deficient patient. (A)...
(A) Immunoblot analysis of calnexin, ARPC1B, and ARPC1A in hCTL lysates from 4 different HD (lanes 1 to 4) and ARPC1B patient samples from day 20 (lanes 4 to 6) and day 12 (lanes 7 and 8) after expansion. HD 4 (lanes 4 and 7) was used for the rest of the study. MW markers in kD. Lanes were run on the same gel, but were noncontiguous. (B) Single plane of a Z-stack across hCTLs from HD or ARPC1B patient plated on ICAM-1– (0.5 μg/mL) or anti-CD3–coated (10 μg/mL; right panel) slide, fixed, stained with phalloidin and CD8 (not shown), and imaged using Structural Illumination Microscopy. Scale bars: 3 μm. (C) Quantitation of the mean intensity of phalloidin staining (F-actin) in HD or ARPC1B-deficient patient hCTLs generated from images as exemplified in B (right panel) (see Methods for description). HD n = 10 cells ARPC1B-patient n = 13 cells. ***P = 0.0053, unpaired t test. Data representative of 3 independent experiments.