The vimentin intermediate filament network restrains regulatory T cell suppression of graft-versus-host disease
Cameron McDonald-Hyman, James T. Muller, Michael Loschi, Govindarajan Thangavelu, Asim Saha, Sudha Kumari, Dawn K. Reichenbach, Michelle J. Smith, Guoan Zhang, Brent H. Koehn, Jiqiang Lin, Jason S. Mitchell, Brian T. Fife, Angela Panoskaltsis-Mortari, Colby J. Feser, Andrew Kemal Kirchmeier, Mark J. Osborn, Keli L. Hippen, Ameeta Kelekar, Jonathan S. Serody, Laurence A. Turka, David H. Munn, Hongbo Chi, Thomas A. Neubert, Michael L. Dustin, Bruce R. Blazar
Cameron McDonald-Hyman, James T. Muller, Michael Loschi, Govindarajan Thangavelu, Asim Saha, Sudha Kumari, Dawn K. Reichenbach, Michelle J. Smith, Guoan Zhang, Brent H. Koehn, Jiqiang Lin, Jason S. Mitchell, Brian T. Fife, Angela Panoskaltsis-Mortari, Colby J. Feser, Andrew Kemal Kirchmeier, Mark J. Osborn, Keli L. Hippen, Ameeta Kelekar, Jonathan S. Serody, Laurence A. Turka, David H. Munn, Hongbo Chi, Thomas A. Neubert, Michael L. Dustin, Bruce R. Blazar
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Research Article Immunology

The vimentin intermediate filament network restrains regulatory T cell suppression of graft-versus-host disease

Abstract

Regulatory T cells (Tregs) are critical for maintaining immune homeostasis. However, current Treg immunotherapies do not optimally treat inflammatory diseases in patients. Understanding the cellular processes that control Treg function may allow for the augmentation of therapeutic efficacy. In contrast to activated conventional T cells, in which protein kinase C-θ (PKC-θ) localizes to the contact point between T cells and antigen-presenting cells, in human and mouse Tregs, PKC-θ localizes to the opposite end of the cell in the distal pole complex (DPC). Here, using a phosphoproteomic screen, we identified the intermediate filament vimentin as a PKC-θ phospho target and show that vimentin forms a DPC superstructure on which PKC-θ accumulates. Treatment of mouse Tregs with either a clinically relevant PKC-θ inhibitor or vimentin siRNA disrupted vimentin and enhanced Treg metabolic and suppressive activity. Moreover, vimentin-disrupted mouse Tregs were significantly better than controls at suppressing alloreactive T cell priming in graft-versus-host disease (GVHD) and GVHD lethality, using a complete MHC-mismatch mouse model of acute GVHD (C57BL/6 donor into BALB/c host). Interestingly, vimentin disruption augmented the suppressor function of PKC-θ–deficient mouse Tregs. This suggests that enhanced Treg activity after PKC-θ inhibition is secondary to effects on vimentin, not just PKC-θ kinase activity inhibition. Our data demonstrate that vimentin is a key metabolic and functional controller of Treg activity and provide proof of principle that disruption of vimentin is a feasible, translationally relevant method to enhance Treg potency.

Authors

Cameron McDonald-Hyman, James T. Muller, Michael Loschi, Govindarajan Thangavelu, Asim Saha, Sudha Kumari, Dawn K. Reichenbach, Michelle J. Smith, Guoan Zhang, Brent H. Koehn, Jiqiang Lin, Jason S. Mitchell, Brian T. Fife, Angela Panoskaltsis-Mortari, Colby J. Feser, Andrew Kemal Kirchmeier, Mark J. Osborn, Keli L. Hippen, Ameeta Kelekar, Jonathan S. Serody, Laurence A. Turka, David H. Munn, Hongbo Chi, Thomas A. Neubert, Michael L. Dustin, Bruce R. Blazar

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Figure 2

Vimentin disruption augments Treg function.

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Vimentin disruption augments Treg function. (A–C) Suppression of (A) CD4...
(A–C) Suppression of (A) CD4+ and CD8+ Tcon proliferation by WT Tregs transfected with control or vimentin siRNA, (B) CD8+ Tcon proliferation by DMSO- or AEB071-pretreated WT Tregs, and (C) CD4+ and CD8+ Tcon proliferation by PKC-θ–KO Tregs transfected with either control or vimentin siRNA in classical in vitro Treg suppression assays. 1:1 to 1:9 Treg/Tcon ratio. (D) Survival and (E) clinical GVHD scores (0 = no disease, 10 = severe disease) for recipient mice after receiving BM, BM plus Tcons (BM+T), or BM plus Tcons plus Tregs pretreated with DMSO or AEB071 (DMSO or AEB071). Data were pooled from 4 independent experiments. BM, n = 25; BM+T, n = 29; DMSO, n = 29; AEB071, n = 31. (F) Survival and (G) clinical GVHD scores for recipient mice after receiving BM only, BM plus Tcons, or BM plus Tcons plus Tregs transfected with control or vimentin siRNA. Data were pooled from 2 independent experiments. BM, n = 10; BM plus Tcons, n = 12; control siRNA, n = 12; vimentin siRNA, n = 12. Statistical comparisons in E and G represent DMSO versus AEB071 and control versus vimentin siRNA, respectively. Data in A–C are results from 1 representative experiment of 4 independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by 1-way ANOVA with multiple comparisons analysis and Tukey’s post test (A–C), log-rank test for survival analysis (D and F), and unpaired Student’s t test (E and G). Error bars indicate the SEM.