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Received 1 Dec 2015 | Accepted 7 Apr 2016 | Published 23 Jun 2016

Bonnie van Wilgenburg1,*, Iris Scherwitzl2,*, Edward C. Hutchinson3, Tianqi Leng1, Ayako Kurioka1,Corinna Kulicke1, Catherine de Lara1, Suzanne Cole4, Sirijitt Vasanawathana5, Wannee Limpitikul6,Prida Malasit7,8, Duncan Young9, Laura Denney4, STOP-HCV consortiumw, Michael D. Moore3, Paolo Fabris10, Maria Teresa Giordani10, Ye Htun Oo11, Stephen M. Laidlaw12, Lynn B. Dustin12, Ling-Pei Ho4,Fiona M. Thompson1, Narayan Ramamurthy1, Juthathip Mongkolsapaya2,8,**, Christian B. Willberg1,13,**, Gavin R. Screaton2,** & Paul Klenerman1,13,**

Mucosal-associated invariant T (MAIT) cells are abundant in humans and recognize bacterial ligands. Here, we demonstrate that MAIT cells are also activated during human viral infections in vivo. MAIT cells activation was observed during infection with dengue virus, hepatitis C virus and inuenza virus. This activationdriving cytokine release and Granzyme B upregulationis TCR-independent but dependent on IL-18 in synergy with IL-12, IL-15 and/or interferon-a/b. IL-18 levels and MAIT cell activation correlate with disease severity in acute dengue infection. Furthermore, HCV treatment with interferon-a leads to specic MAIT cell activation in vivo in parallel with an enhanced therapeutic response. Moreover,

TCR-independent activation of MAIT cells leads to a reduction of HCV replication in vitro mediated by IFN-g. Together these data demonstrate MAIT cells are activated following viral infections, and suggest a potential role in both host defence and immunopathology.

1 Peter Medawar Building for Pathogen Research and Translational Gastroenterology Unit, Nufeld Department of Clinical Medicine, University of Oxford, Oxford OX1 3SY, UK. 2 Division of Immunology and Inammation, Department of Medicine, Faculty of Medicine, Imperial College, Hammersmith Campus, London W12 0NN, UK. 3 Sir William Dunn School of Pathology, University of Oxford, University of Oxford, South Parks Road, Oxford OX1 3RE, UK. 4 MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, Nufeld Department of Clinical Medicine, University of Oxford, Oxford OX3 9DU, UK.

5 Department of Pediatric, Khon Kaen Hospital, Khon Kaen 40000, Thailand. 6 Department of Pediatric, Songkhla Hospital, Songkhla 90100, Thailand. 7 BIOTEC, NSTDA, Phatumthani 12120, Thailand. 8 Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand. 9 Nufeld Department of Anesthesia, The John Radcliffe Hospital, Oxford OX3 9DU, UK. 10 Infectious Diseases and Tropical Medicine Unit, San Bortolo Hospital, Vicenza, VI 37, Italy. 11 Centre for Liver Research & NIHR Biomedical Research Unit in Liver Disease, University of Birmingham, Birmingham B15 2TT, UK. 12 Nufeld Department of Orthopaedics, Rheumatology, and Musculoskeletal Sciences, Kennedy Institute of Rheumatology, University of Oxford, Oxford OX3 7 LF, UK. 13 Oxford NIHR Biomedical Research Centre, The John Radcliffe Hospital, Oxford OX3 9DU, UK. * These authors contributed equally to this work. ** These authors jointly supervised this work. Correspondence and requests for materials should be addressed to B.v.W. (email: mailto:[email protected]

Web End [email protected] ) or to J.M. (email:mailto:[email protected]

Web End [email protected] ) or to P.K. (email: mailto:[email protected]

Web End [email protected] ). wA full list of consortia members appear at the end of the paper.

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DOI: 10.1038/ncomms11653 OPEN

MAIT cells are activated during human viral infections

ARTICLE NATURE COMMUNICATIONS | DOI: 10.1038/ncomms11653

Mucosal-associated invariant T (MAIT) cells are an abundant and distinctive T cell subset rst identied by Porcelli et al.1 in 1993 in humans on the basis of

the invariant V-alpha chain (Va7.2-Ja33) and later described in mice2. MAIT cells comprise B510% of T-cells within the periphery and are found enriched in both the gut and in the liverwhere in humans they may represent between B12% and 50% of T-cells36. In both mouse and man, MAIT cells are restricted to a bacterial riboavin synthesis pathway-derived ligand presented by the conserved MHC-class I like molecule MR1 (refs 7,8). In vivo challenge models using riboavin synthesizing bacteria, such as Klebsiella species and mycobacteria, have conrmed an anti-bacterial role for MAIT cells9,10. MR1 tetramers loaded with the ligand rRL-6-CH2OH have allowed the specic detection of human and murine MAIT cells11.

In recent years, a number of groups have elucidated the distinct phenotypic and functional prole of MAIT cells, characterized by the high expression of the C-type lectin CD161 (KLRB1 or NKR-P1A), and the capacity to secrete multiple cytokines, including IL-17, interferon (IFN)-g and TNF-a1216. The distinctive phenotype of MAIT cells appears to be driven by two key transcription factors, RORgt and PLZF3,6,13,17. RORgt expression is linked to development of type 17 function and expression of surface receptors such as IL23R and CCR6 (refs 5,18). This is consistent with mucosal defence and anti-bacterial functions and also consistent with the bacterial specicity of the receptor. PLZF is critical for development of invariant natural killer T cell (iNKT) cells and may be responsible for a distinct set of innate phenotypic features, including marked upregulation of the pro-inammatory cytokine receptors IL-18R and IL-12R19,20. This dual transcriptional drive suggests that MAIT cells may possess multiple parallel functionalities or modes of activation.

Given the specicity of the T-cell receptor (TCR), it appears that activation of MAIT cells in vivo is driven by responsiveness to bacteria (and some yeasts)21. However, given their innate phenotype, broad range of effector functions, and tissue distribution, we addressed the question of whether they may also have evolved to respond to viral infections in vivo. If so, this would markedly widen the role of MAIT cells in human infection.

Here, we addressed MAIT cell activation in response to a set of major human viral pathogens both in vivo and in vitro. In vivo, MAIT cells are activated in response to dengue virus (DENV), hepatitis C virus (HCV) and inuenza virus. In vitro, DENV, HCV and inuenza virus readily trigger MAIT cell populations in an IL-18 dependent manner, in concert with other virally-driven pro-inammatory cytokines, including IFN-a/b. Activation of

MAIT cells in acute dengue correlated both with disease outcome and with IL-18 signals. Similarly, in vivo activation of MAIT cells during HCV therapy correlated with specic addition of IFN-a during therapy. Taken together, these data strongly implicate a role for MAIT cells in response to major virus infections of man and provide a mechanism for their virus-responsive nature. Overall, this signicantly expands the pathogen response repertoire of this abundant human T-cell subset.

ResultsMAIT cell activation during acute viral infections in vivo. Whether human MAIT cells respond during acute viral infections remains unclear. To address this we analysed MAIT cell activation in DENV and inuenza virus infection. In this study, MAIT cells were dened as CD161 Va7.2 CD8

CD4 CD3 T cells. DENV infection presents with a rapid onset of a febrile illness, which lasts around a week. After

the virus has been cleared and fever drops, patients follow two distinct courses: an uncomplicated resolution (dengue fever; DF) or a severe course associated with organ dysfunction and haemorrhage (dengue haemorrhagic fever; DHF). The determinants driving outcome are still poorly understood. However, an aberrant immune response, around the day of defervescence, is thought to have an important role in the pathogenesis. We analysed MAIT cells in a longitudinal study of peripheral blood mononuclear cells (PBMC) from a cohort of patients with DHF or DF. Blood samples were collected at different time points during the acute phase of infection and dates were named in relation to the day fever subsided (day of defervescence, day 0). Samples taken 10 or more days after the day of defervescence (day 410) were considered convalescent.

We found a small but signicant reduction in MAIT cell frequencies between the acute phase (day 0) and the convalescent phase (day 410) of infection (Fig. 1a,b), but no differences in

MAIT cell frequency between DHF and DF patients (Fig. 1c).

Analysis of CD38 revealed marked in vivo activation of MAIT cells (Fig. 1d,e), which increased over the course of infection and peaked at a critical moment for DENV infected patientsthe day of defervescence. Interestingly, patients who developed the severe form of dengue had higher levels of MAIT cell activation as judged by CD38 expression compared to DF patients over the course of acute infection (Fig. 1f). MAIT cell activation resolved to healthy control levels in the convalescent sample (Fig. 1d,e).

Granzyme B expression was also assessed due to its tight regulation in MAIT cells, and its absence in cells from healthy donors3,22. Furthermore, upregulation of Granzyme B is associated with the acquisition of cytolytic function by MAIT cells22,23. We therefore analysed Granzyme B function in acute dengue and found this followed the same time course as that of CD38 (Fig. 1gi).

Given their role in mucosal defence, we next addressed the activation of MAIT cells in response to inuenza virus, a virus with a segmented genome of negative-sense RNA. Again, patients with acute, severe inuenza virus infection had reduced MAIT cell frequencies and an increase in Granzyme B expression on MAIT cells (Fig. 1jm).

Taken together, our results indicate substantial triggering of MAIT cells in vivo during acute viral infection.

MAIT cell activation during chronic viral infection in vivo. MAIT cells are found at high frequencies within the liver in both healthy and diseased states3,5,13,24. Therefore, we analysed MAIT cell activation during chronic infection with HCV, like DENV, a member of the Flaviviridae family of positive-sense RNA viruses. We examined MAIT cell frequency and phenotype in the PBMC of patients with persistent and resolved HCV infection (spontaneously or after therapy). In all HCV patients, regardless of status, we observed a reduction in MAIT cell frequencies compared to healthy controls (Fig. 2a). However, we only observed upregulation of Granzyme B in patients with prolonged HCV infection (including those who had subsequently responded to antiviral therapy; Fig. 2b,c), and not in those patients with prior short-lived viremia at a distant time-point associated with acute resolving infection (thus, more akin to convalescent DENV infection). Our results indicate substantial activation of MAIT cells in vivo both during acute and chronic viral infections.

Viral MAIT cell activation in vitro. Having observed MAIT cell activation in vivo during acute and chronic viral infections, we next established in vitro models for viral infections using PBMCs or human CD8 T cells, co-incubated with infected or

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NATURE COMMUNICATIONS | DOI: 10.1038/ncomms11653 ARTICLE

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Figure 1 | MAIT cell activation during acute viral infections in vivo. PBMCs from healthy controls (HC, n 510), patients suffering from severe dengue

(DHF, n 210), dengue (DF, n 410) or acute, hospitalized patients infected with inuenza A virus (n 79) were analysed by ow cytometry by

gating on live CD3CD8 CD161 Va7.2 (MAIT) cells. (a,d,g) Representative ow cytometry plots. (b,e,h) Comparison between acute (day 0) and convalescent phase (day 410) of infection. (c,f,i) Comparison between DHF and DF patients. (jm) Acute, hospitalized patients infected with inuenza virus. (b,c,j,k) MAIT cell frequency as a proportion of the CD8 T cell population. (e,f) Percentage of CD38 expression by MAIT cells. (h,i,l,m) Percentage of Granzyme B expression by MAIT cells. Statistical signicance was determined with a Wilcoxon matched-paired test (b,e,h) or MannWhitney test (j,k). Bars represent meanss.e.m. ns40.05, *P 0.05, **Pr0.01, ***Pr0.001, ****Pr0.0001. conv, convalescent; Gr B, Granzyme B; HC, healthy control;

ICU, intensive care unit.

virus-treated dendritic cells (DCs) or macrophages as antigen presenting cells (APC).

MAIT cells were readily and specically activated in response to DENV-treated APCs (multiplicity of infection (MOI) 1), as

indicated by production of IFN-g and Granzyme B (Fig. 3a,b), as well as, expression of CD38, with minimal production of TNF-a (Supplementary Fig. 1ac). Activation was dependent on the presence of APCs as DENV alone did not activate MAIT cells (data not shown).

We next examined activation of MAIT cells in vitro in response to inuenza virus. APCs incubated with inuenza virus (MOI 1) promoted strong MAIT cell activation, inducing

upregulation of CD69 and IFN-g (with minimal TNF-a secretion), in a dose-dependent manner (Fig. 3c,d). Also, Granzyme B expression was induced (Fig. 3e). This activation was not restricted to laboratory-adapted inuenza virus strains, but also was seen in response to recently-circulating strains of inuenza A and B (Fig. 3f). Interestingly, MAIT cell activation was also readily induced using PBMCs instead of puried APCs (Supplementary Fig. 1d). Moreover, inuenza virus exposure of puried monocyte, macrophage and DC populations all led to MAIT cell activation (Supplementary Fig. 1e).

Finally, we tested the impact of HCV on MAIT cells in vitro. MAIT cells expressed IFN-g and Granzyme B (Fig. 3g,h), as well

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ARTICLE NATURE COMMUNICATIONS | DOI: 10.1038/ncomms11653

a HC

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Figure 2 | MAIT cell activation during chronic viral infection in vivo. PBMCs from healthy controls (n 2023) or patients (n 1225) with persistent

(treatment naive, REL, NR) and resolved HCV infection (SVR, SC) were analysed by ow cytometry by gating on live CD3 CD8CD161 Va7.2 (MAIT) cells. (a) MAIT cell frequency as a proportion of the CD8 Tcells. (b,c) Granzyme B expression by MAIT cells. (b) Representative ow cytometry plots. Bars represent meanss.e.m. Statistical signicance was determined with the KruskalWallis test followed by the Dunns test. ns40.05,*P 0.05, **Pr0.01, ***Pr0.001, ****Pr0.0001. HC, healthy control; Gr B, Granzyme B; REL/NR, relapse/non-response; SC, spontaneous clearance;

SVR, sustained virological response; tr., treatment.

as CD69 in response APCs treated with HCV (MOI 1), in a

dose-dependent and specic manner (Supplementary Fig. 1fi).

Taken together, we demonstrate that MAIT cells are activated in vitro in response to DENV, inuenza virus and HCV-exposed APCs.

Viral MAIT cell activation is dependent on cytokines. Next, we investigated the mechanisms of MAIT cells activation in response to viruses. The ligand presented by MR1 is derived from the riboavin synthesis pathway, which is not found in host cells or viruses7. Consistent with this, using the models described above, we conrmed by antibody blocking of MR1 that MAIT cell activation was TCR-independent for all the viruses tested (Fig. 4ac).

As activation was TCR-independent, we explored triggering of MAIT cells by cytokines. Previously, we have shown that TLR8 is capable of inducing IFN-g expression in MAIT cells via IL-12 and

IL-18 (ref. 24). In addition, IL-15 can specically activate distinct functions of MAIT cells in synergy with IL-12 and/or IL-18, in a dose-dependent manner (Supplementary Figs 2 and 3)25. We extended this nding by exploring responses to a range of TLR ligands in PBMCs and found endosomal TLR3 was also a potent activator (Supplementary Fig. 4). As with TLR8, TLR3 induced MAIT cell activation via IL-18 and IL-12 and not MR112,13. TLR sensing by APCs can occur in the absence of viral replication26,27. To assess the requirement of viral replication for MAIT cell activation, we used ultraviolet-irradiation of the viruses, which prevents transcription or replication. Ultravioletirradiated DENV was no longer able to activate MAIT cells (Supplementary Fig. 5a). In contrast, ultraviolet-irradiated HCV and inuenza virus were still able to activate MAIT cells, although less efciently compared to untreated virus (Supplementary Fig. 5b,c). Accordingly, DENV productively infects APCs, whereas productive inuenza virus and HCV infection is limited in both primary and stem cell-derived human APCs (Supplementary Fig. 5df)2830. Furthermore, the level of DENV infection correlated with MAIT cell IFN-g expression (Supplementary Fig. 5e).

We next assessed blockade of IL-12, IL-18 and IL-15 in the in vitro virus models. Figure 4d reveals that IL-12 and IL-18 had the most signicant impact on DENV-dependent MAIT cell activation using DCs, with anti-IL-18 blockade alone leading to complete suppression of IFN-g expression. IL-18 was the most abundant cytokine induced in DCs in response to DENV (Fig. 4e).

Anti-IL-18 also inhibited activation of MAIT cells induced by inuenza virus, a result observed using puried APCs (Fig. 4f) or PBMCs (Supplementary Fig. 6a). Blockade of IL-12 and IL-15

alone or in combination had a limited impact in inuenza virus-induced MAIT cell activation using macrophages (Fig. 4f). Accordingly, IL-18, but not IL-12 or IL-15 was secreted by macrophages after incubation with inuenza virus (Fig. 4g).

Similar to DENV and inuenza virus, IL-18 had a dominant role in HCV-induced MAIT cell activation (Fig. 4h). Addition of further blockade with anti-IL-15 was required to achieve maximal suppression. We assessed expression of IL-12, IL-15 and IL-18 on macrophages after incubation with HCV (Fig. 4i). No IL-15 protein was detected, yet IL-15R (responsible for IL-15 trans-presentation31) was upregulated by macrophages in response to HCV, as measured by quantitative PCR (Supplementary Fig. 6b).

Overall, these experiments indicate a mechanism by which viruses can activate MAIT cells, dependent on viral sensing and cytokines, with a key role for IL-18, and independent of MR1 and TCR.

Virally-triggered IL-18 correlates with MAIT cell activation. Having identied IL-18 as an important cytokine for viral MAIT cell activation in vitro, we further explored its relevance in vivo. In DENV infection, IL-18 levels in plasma were markedly increased during the acute phase compared to the convalescent phase of infection and healthy controls (Fig. 5a). In addition, patients suffering from the severe form of dengue (DHF) had signicantly higher levels of IL-18 over the course of acute infection compared to patient with DF (Fig. 5b). Interestingly, expression of the IL-18 receptor (IL-18R) on MAIT cells was upregulated during the acute phase of infection and resolved to healthy control levels in the convalescent samples (Fig. 5c). As observed for IL-18, patients with DHF revealed higher expression of IL-18R on MAIT cells compared to patient with DF (Fig. 5d,e). Thus, levels of IL-18 in plasma and expression of IL-18R on MAIT cells in vivo are upregulated during the acute phase of infection, especially in those patients suffering from DHF. These ndings show an association with increased activation of MAIT cells and disease severity in dengue patients (Figs 1f and 5f). These results are not restricted to DENV infection since IL-18 protein (Fig. 5g) and mRNA (Fig. 5h) were found readily in liver tissues from patients with HCV infection.

Overall, these data indicate that IL-18 is not only a dominant factor during in vitro viral MAIT cell activation, but also has an important role in vivo.

MAIT cells respond to type I IFNs in vitro and in vivo. Type I IFNs are also important cytokines known for their strong antiviral activity. Previous reports have determined a crucial role

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Figure 3 | Viral MAITcell activation in vitro. MAITcells from healthy individuals were analysed by ow cytometry, gated on live CD3 CD161 Va7.2 cells. (a,b) PBMCs (n 7) were co-cultured with autologous monocyte-derived DCs exposed to DENV (MOI 1) as described in Methods. (ce) CD8

Tcells isolated from PBMCs (n 1112) were co-cultured with IAV-exposed macrophages (MOI 1) as described in Methods, unless indicated otherwise.

(f) CD8 T cells isolated from PBMCs (n 1112) were co-cultured with macrophages exposed to the clinical H3N2 inuenza A strain (A/England/691/

2010 (n 7)) or inuenza B (B/Florida/04/2006 (n 8)) (MOI 1) as described in Methods. (g,h) CD8 T cells isolated from PBMCs (n 712)

co-cultured with macrophages exposed to HCV (MOI 1) as described in Methods. Proportion of MAIT cells producing IFN-g (a,c,d,f,g), TNF-a

(c,d), CD69 (c,d) or Granzyme B (b,e,h). (d) Representative ow cytometry plots. All data are representative from at least two independent experiments. Bars represent meanss.e.m. Statistical signicance was determined with the KruskalWallis test followed by the Dunns test (f) or the MannWhitney test (a,b,e,g,h). ns40.05, *P 0.05, ***Pr0.001, ****Pr0.0001. Eng, England; Flor, Florida; HC, healthy control; Gr B, Granzyme B; MF, macrophage;

REL/NR, relapse/non-response; SC, spontaneous clearance; SVR, sustained virological response; tr., treatment.

of IFN-a/b during DENV32, inuenza33 and HCV34 infections in limiting viral replication and disease severity. We therefore addressed whether IFN-a/b can also inuence MAIT cell activation in vitro and in vivo.

First, we tested whether IFN-a/b could trigger MAIT cells alone or in combination. We found IFN-g secretion by MAIT

cells in response to IFN-a/b in combination with IL-12 and IL-18, but not IL-15 (Fig. 6a), in a dose-dependent manner (Supplementary Fig. 7). Next, we conrmed that these TCR-independent responses to cytokines are shared with other CD161 T-cell subsets, but not CD161 T-cell populations.

CD161 CD8 T cells that lack expression of the Va7.2Ja33

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Figure 5 | Virally triggered IL-18 correlates with MAIT cell activation. (ad) PBMCs from healthy controls (n 10) or patients suffering from

severe dengue (DHF, n 212) or dengue (DF, n 412) were analysed.

(a,b) Plasma levels of IL-18. (c,d) IL-18Ra expression on MAIT cells.(a,c) Comparison between acute (day 0) and convalescent (conv) phase (day 410) of infection. (b,d) Comparison between DHF and DF patients.

Correlation of plasma levels of IL-18 (e) or MAIT cell CD38 expression (f) against MAIT cell IL-18Ra expression by the Spearman-rank correlation test. (g) Parafn-embedded human liver tissues sections from HCV or control patients were stained for IL-18 (brown-coloured). Scale bar, 10 mm. (h) IL-18 mRNA expression in liver biopsies from HCV patients (n 55)

relative to uninfected control liver samples is shown (n 6). Bars

represent meanss.e.m. Statistical signicance was determined with the MannWhitney test (b,d,h) or the Wilcoxon matched-paired test(a,c). ns40.05, *P 0.05, ***Pr0.001, ****Pr0.0001. conv, convalescent;

HC, healthy control.

0.1

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Mock HCV

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Figure 4 | Viral MAIT cell activation is dependent on cytokines. (ad,f,h) Isotype control, anti-MR1, anti-IL-12, anti-IL-15 and/or anti-IL-18 antibodies were added to a co-culture exposed to DENV (a,d, n 36), IAV (b,f, n 5

29) or HCV (c,h, n 716). IFN-g expression by MAIT cells (gated on live

CD3CD8 CD161 Va7.2 cells) was analysed by ow cytometry and is shown relative to the isotype control. (e,g,i) IL-12p70, IL-15 and IL-18 levels secreted by virus-exposed APCs. (e) DENV-exposed DCs (n 4) at

42 h. (g) IAV-exposed macrophages (n 7) at 48 h. (i) HCV-exposed

macrophages (n 7) at 48 h. Data are representative from at least two

independent experiments. Bars represent meanss.e.m. Statistical signicance was determined with the MannWhitney test (ac,e,g,i) or the KruskalWallis test followed by the Dunns test or the MannWhitney test (d,f,h). ns40.05, *P 0.05, ***Pr0.001, ****Pr0.0001. ND, not detected.

0 200 300 400 500 200 300 400 500

TCR (Supplementary Fig. 8a), CD161 Va7.2Ja33 CD4 T cells (Supplementary Fig. 8b) and CD4 CD8 T cells (Supplementary Fig. 8c) could all be triggered in response to cytokines, unlike CD161 T-cell populations (Supplementary

Fig. 8d). This is in agreement with our previous ndings which suggest that CD161-expressing lymphocytes display a shared innate response to cytokines16.

To further explore the role of type I IFNs during viral MAIT cell triggering, we used a vaccinia virus-encoded, soluble type I

IFN receptor (B18R)35. Macrophages treated with HCV in vitro were incubated in the presence of B18R and T cells overnight. Blockade of type I IFNs by B18R inhibited MAIT cell IFN-g secretion (Fig. 6b), CD69 expression (Fig. 6c) and Granzyme B upregulation (Fig. 6d).

IFN-a has been an important component of HCV treatment. This gave us an opportunity to explore whether type I IFNs could contribute to MAIT cell activation in vivo. We measured the activation marker CD69 on MAIT cells from HCV patients taking part in a clinical trial in which they were treated with either sofosbuvir (SOF) ribavirin (RBV) pegylated IFN

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IFN-expression

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IFN-expression

% of MAIT cells

HCV replication

(RLU)

5107

IL-12/18

0 Neat

1:50

1:500

1:5000

IL-12

IL-15

MAIT+ supernatant

Unstimulated

IFN-

IL-18

IFN- + IL-12

IFN- + IL-15

IFN- + IL-18

IL-18 + IL-12

IL-18 + IL-15

IL-12 + IL-15

1.5108

HCV replication

(RLU)

2.0

HCV replication

1.0108

1.5

***

IFN-expression

***

Relative

1.0

Relative

CD69 expression

Relative

Gr B expression

1.0

5.0107

0.5

Relative

0.5

0.0

MAIT MAIT+

0.0

HCV luc MAIT sup Stimulation

+ +

+ + +

Iso

IFN-

Iso

IFN-

mock HCV

PBS

B18R

mock HCV

PBS

B18R

PBS

B18R

CD69 expression

% of MAIT cells

Figure 7 | Activated MAIT cells can limit HCV replication. Sorted MAIT cells (CD8 CD161 Va7.2 ) were rested or activated in a TCR-independent manner (IL-12 IL-18 stimulation, MAIT ) for 24 h. (a) IFN-g

expression by MAIT cells (gated on live CD3 CD8CD161 Va7.2 cells) was analysed by ow cytometry. (b,c) Neat or diluted supernatants were transferred to hepatocyte lines infected with HCV expressing luciferase and viral replication measured 4 days post infection.(d) Supernatants were transferred to hepatocyte lines infected with HCV expressing luciferase repeated in the presence of an isotype control or anti-IFN-g antibody. Data are representative from at least two independent experiments. Bars represent meanss.e.m. Statistical signicance was determined with a Wilcoxon matched-paired test (c) or the MannWhitney test (d). **Pr0.01. , unstimulated; luc, luciferase; RLU, relative light

units; sup, supernatant.

SOF + RBVSOF + RBV / PEG IFN

*** *

0 HC

HCV patients (BOSON trial)

Baseline

4 Weeks

End

Figure 6 | MAIT cells respond to type I interferons in vitro and in vivo. (a) PBMCs from healthy individuals were directly stimulated for 24 h with IFN-a, IFN-b, IL-12, IL-15, IL-18 or indicated combinations thereof. IFN-g expression by MAIT cells (gated on live CD3 CD8 CD161 Va7.2 cells) was analysed by ow cytometry. (bd) B18R (IFN-a/b neutralizing protein) or PBS control were added to the co-culture (n 611). IFN-g

(b), CD69 (c) and Granzyme B (d) expression is shown relative to the control. Data are representative from at least two independent experiments. (e) PBMCs from healthy controls or HCV patients at baseline, during or end of treatment with either SOF RBV or SOF RBV/PEG-IFN were

analysed by ow cytometry. CD69 expression on MAIT cells (gated on live CD3CD8 CD161 Va7.2 cells) was measured. Bars represent meanss.e.m. Statistical signicance was determined with the

KruskalWallis test followed by the Dunns test (a) or the MannWhitney test (be). ns40.05, *P 0.05, ***Pr0.001, ****Pr0.0001. HC,healthy controls.

(PEG-IFN), or only SOF RBV, without PEG-IFN. Interestingly,

CD69 expression on MAIT cells was upregulated only in the treatment group receiving PEG-IFN (Fig. 6e), which corresponded with a signicantly higher sustained virologic response rate36. High CD69 upregulation was specic to CD161 MAIT cells and not observed on CD161 CD8 or

CD4 T cells (Supplementary Fig. 9). These data demonstrate that type I IFNs can contribute to MAIT cell activation in vitro and in vivo.

Activated MAIT cells can limit HCV replication. Finally, we explored whether viral MAIT cell activation can have a functional impact on virus replication. We stimulated sorted MAIT cells in a TCR-independent manner (IL-12 and IL-18) and conrmed activation by measurement of IFN-g secretion (Fig. 7a). The cell

supernatants of unstimulated and stimulated MAIT cells were transferred to a hepatocyte line infected with HCV expressing luciferase, allowing measurement of HCV replication. The activated MAIT cell supernatants were able to potently suppress HCV replication, while no activity from control supernatants (Fig. 7b) or from IL-12 and IL-18 (data not shown) addition to hepatocytes was seen. Diluting the activated MAIT cell supernatant reduced the suppression of HCV replication in a dose-dependent manner (Fig. 7c). Addition of anti-IFN-g reversed the HCV suppression by activated MAIT cell supernatants (Fig. 7d). In an extension of the experimental set up using virally-induced activation to trigger MAIT cells, supernatants from MAIT cells incubated with HCV-treated macrophages reduced HCV replication in the hepatocyte reporter cell line compared to mock-treated macrophages (Supplementary Fig. 10). These results indicate that activation of MAIT cells by viral triggers induces responses that can impact on viral replication in vitro.

DiscussionMAIT cells are an abundant human T-cell population with a semi-invariant TCR determining bacterial specicity. It has previously been considered that MAIT cells were not responsive to viral infection37. However, in this study we provide data to show MAIT cells are also readily and specically activated by pathogenic viruses in vivo and in vitro. Furthermore, we dene mechanisms that allow this virally-driven activation, and show they are distinct in the different viral settings.

The in vivo data shows that there is evidence of MAIT cell activation in response to DENV, HCV and inuenza virus.

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Of these the most striking is the data from acute DENV infection as it is clear that such activation is profound and rapid in this severe clinical setting. Of note, those patients who went on to develop DHF exhibited signicantly higher levels of activation as judged by CD38 expression. This suggests the MAIT cell response could contribute to the pathophysiology of this condition. The correlation between levels of IL-18, levels of IL-18R and MAIT cell activation suggests an important role for IL-18 in this activation process, consistent with the in vitro mechanisms demonstrated. In DENV, HCV and inuenza virus infection we found clear evidence of activation of the MAIT cells, as judged by upregulation of Granzyme B (which is tightly regulated in MAIT cells22). In addition, we noted a decrease in circulating MAIT cell frequencies. The mechanism for this is not clear. It may result from translocation of some cells from blood to tissues, or it could result from activation-induced cell death as has been suggested for HIV infection38.

The data presented are correlativehowever probing a non-redundant role in animal models is not straightforward. In mice the MAIT cell population is a very small minority compared to the frequencies of iNKT cells, which have overlapping functions and innate cytokine responsiveness39. In humans, the situation is reversed, with MAIT cell frequencies exceeding those of iNKT cells by 12 logs13. Existing data indicate a functional antiviral role for iNKT cells in control of virus infections in tissues such as lung, in mouse models: given similar molecular mechanisms of activation apply, and similar effector functions are induced, it seems reasonable to extrapolate from this that MAIT cell activity would have a parallel role in human infection40. Emerging models with increased MAIT cell frequencies in mice may help address this question using new in vivo approaches41.

MR1 tetramers unequivocally identify MAIT cells, but these reagents are not yet widely available. In this study we used CD161 Va7.2 as a surrogate marker for MAIT cells, which is a commonly used staining approach and includes the majority of tetramer-positive MAIT cells6,42. We further dened MAIT cells as CD8 CD4 CD3 T cells, and for in vitro experiments

PBMCs from healthy individuals were used. CD8 is expressed by the majority of MAIT cells, and CD4 only by a small proportion of MAIT cells42. This approach has been used in recent transcriptional studies and allows ready comparison with related CD161-expressing subsets16. Future studies using MR1 tetramers are needed to further establish the distribution of MAIT cell phenotype in virally infected humans in more detail. As we did not have access to the tetramer, the observed loss of MAIT cells in virally infected patients may be potentially explained by down-regulation of CD161, as previously suggested12,43. However, Fernandez et al.44 showed that in HIV-1 infected patients, MR1 tetramers do not bind CD161 Va7.2 T cells, and the loss of MAIT cells in the periphery may be due to recruitment to the site of infection or activation-induced cell death. Of note, our data are not restricted to CD161 Va7.2 cells but also includes responses from the minority

CD161 Va7.2 cells, which we have previously described as sharing phenotypic and functional properties with MAIT cells and which likely also include some recently described Va7.2

MR1-restricted T cell populations45,46.

The mechanisms we have explored reveal a central role for IL-18 in virus-induced activation12,16. However, the critical co-stimulation was not restricted to a single cytokine. In the case of HCV we repeatedly observed an important role for IL-15. IL-15 levels in these experimental conditions were very low, but upregulation of IL-15R was seen and importantly antibody-blockade of IL-15 revealed a clear role for this cytokine. In addition, we observed a role for type I IFNs in the activation of

MAIT cells in response to HCV. This was clear in both addition and blockade experiments and is consistent with data from natural killer cells, where such cytokine-mediated activation is described47.

The cytokine activation of MAIT cells was mediated by APCs with a role for viral sensing and replication. In the case of DENV, viral replication was required for MAIT cell activation and this was positively correlated with the level of productive infection in the APC. Although we observed a reduction of MAIT cell activation in response to UV-irradiated HCV and IAV, replication does not have a key role in this context, since HCV and inuenza poorly replicate in macrophages28,29. As APC have continual and extensive endocytic activity, viral recognition can occur through endocytosis of viral particles as well as through direct infection26,48. Using TLR agonists, we found that both surface and endocytic TLRs can trigger MAIT cells, in particular viral pathogen-associated molecular patterns.

Using murine bone marrow-derived DCs, Le Bourhis et al.37 has previously shown that MAIT cells are not responsive to viral infection. This discrepancy is likely due to the use of murine, rather than human cells, which may reect differential abilities in secreting, as well as, responding to cytokines.

IFN-a-mediated activation of MAIT cells is clinically and functionally relevant, as demonstrated in vivo in the well-controlled BOSON clinical trial. In addition to directly acting antiviral drugs (SOF in combination with RBV), IFN-a was included in one arm only and associated with MAIT cell activation and an enhanced therapy response36. It is possible that MAIT cells act as local ampliers of IFN-a-mediated therapies, contributing to an antiviral effect through secretion of further local antiviral cytokines or potentially via clearance of infected cells. The specic activation by type I IFNs appears to be important evidence in favour of an evolution of MAIT cells in response to viral signals. Although type I IFN signalling is not exclusive to virus infections, a wealth of data links IFN pathways with antiviral defence, and we propose that MAIT cells form an important part of this network in humans. Of note, IFN-a signalling alone was insufcient to fully activate MAIT cells, indicating a potentially fail-safe mechanism to prevent harmful over-activation.

The secondary consequences of MAIT cell activation could be of signicance for both protection and immunopathology. Activation of MAIT cells in tissues could provide both direct and indirect antiviral effects. HCV, for example, is known to be highly sensitive to T-cell-derived IFN-g49,50. Accordingly, we found that activation of MAIT cells by HCV-induced responses that limited HCV replication in vitro, in an IFN-g dependent manner. Thus, the local activation of MAIT cells can be plausibly correlated with a direct antiviral function in vivo.

Loss of MAIT cell frequencies (as we have observed in blood) may contribute to impairment of both viral and bacterial control, whereas excess cytokine secretion by MAIT cells may lead to immunopathologic outcomes. For example, high levels of MAIT cell activation could contribute to the cytokine storm associated with DHF and severe cases of inuenza virus51,52. Activation-induced loss of MAIT cells is potentially relevant in the context of inuenza virus infection, which is epidemiologically associated with secondary bacterial pneumonia due to Staphylococcus aureus or Streptococcus pneumonia53,54. Both of these organisms possess the MAIT cell ligand7,37, and it is plausible that depletion of MAIT cells in situ could lead to transient impairment of local control of these pathogens. Interestingly, the transcription factor PLZF, which likely drives the innate phenotype and functionality of MAIT cells, may also contribute to the induction of activation-induced cell death, through upregulation of specic intracellular caspases55.

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In conclusion, human MAIT cells are activated in response to virus infections (as illustrated by a simplied model depicted in Supplementary Fig. 11), through overlapping mechanisms dependent on the virus and likely the environment and cell types affected. These data extend the likely functions of these abundant human lymphocytes well beyond their traditional connes. Modulation of this cell subset (and related CD161 T-cell subsets16,5658) could provide a novel opportunity to promote antiviral defence or limit immunopathology.

Methods

Patient samples. All samples were collected with appropriate patient consent and local research ethics committee approval. Samples were frozen and stored at

80 C until examined. Healthy donors were supplied by the NHS blood service without any further donor information. The study on DENV patient samples was approved by the Scientic and Ethical Committee of the Khon Kaen Hospital in Thailand and the Riverside Ethics Committee in the UK (06/Q0401/22). Laboratory conrmation of DENV infection was determined by RTPCR detection of DENV nucleic acid (which also conrmed the infecting serotype) or seroconversion in an ELISA of IgM. Secondary infection was dened based on the ratio of DENV-specic IgM to IgG o1.2 on or after day 6 of illness. Disease severity was classied according to 1997 World Health Organization criteria. Of the patients enroled in the study, 10 patients were classied as mildly symptomatic of dengue fever and 10 patients were classied as severely symptomatic of dengue haemorrhagic fever with plasma leakage and bleeding (Supplementary Table 1). The day of defervescence was dened as day 0, the day before defervescence as day

1, the day after defervescence as day 1, and so forth. PBMCs were isolated

from whole blood by Ficoll-Hypaque density-gradient centrifugation at various time points during hospitalization and cryopreserved until further use.

Patients with inuenza virus infection requiring hospitalization were recruited from two hospitals based in Oxford and Glasgow during the 2009 H1N1 pandemic under ethical approval (09/H0606/92). Inuenza virus infection was diagnosed using an in-house generic inuenza A real time PCR assay based on the Matrix gene and conrmed with swine-origin H1N1/09 specic Matrix gene PCR59,60. Patients had concomitant conditions as documented in Supplementary Table 2, but were not on immunosuppressants or steroids at the point of sampling. Two patients were pregnant, and one was 6 days post-partum. PBMC samples from HCV patients (Supplementary Table 3) were collected from the Hepatitis Clinic at the John Radcliffe Hospital, Oxford, UK and consented according to a locally approved protocol (COREC 04.OXA.010). Liver biopsy specimens were obtained from patients with HCV infection (n 55) at S. Bortolo Hospital, Vicenza, Italy

and scored using the Ishak system. Control samples were normal adjacent tissue from six uninfected volunteers undergoing liver resection for other reasons (source Proteogenex, CA, USA). Trials were conducted in accordance with the Declaration of Helsinki, ethical approval was obtained from local ethics committees, and all patients provided written informed consent. Explanted HCV-diseased liver tissues were obtained from QE transplant programme and non-diseased normal liver tissue was obtained from donor liver tissue surplus to clinical requirements. Normal liver was liver tissue from hepatic resection of colorectal metastasis. For studying MAIT cell activation in the context of HCV infection and type I IFNs in vivo, PBMC from the BOSON clinical trial were used, as part of a collaboration between STOP-HCV and Gilead Sciences36,61. BOSON is a randomized, open-label, phase 3 study testing SOF plus RBV with or without pegylated IFN-a in patients with HCV genotype 3 and treatment-experienced cirrhotic patients with HCV genotype 2 (Supplementary Table 4). This study was registered with the European Clinical Trials Database, number 2013-002641-11.

Tissue staining. Parafn-embedded human liver tissues sections were dewaxed using Clearene and IMS. Endogenous peroxidase was blocked by using 5% hydrogen peroxide followed by antigen retrieval using 10% EDTA buffer. Slides were blocked using casein buffer. Primary Rabbit IL-18 antibody LS-B2809, 1:50 dilution was applied for an hour followed by Vector Impress anti-rabbit secondary for 30 min in a staining chamber with rocking. Slides were then washed with PBS Tween and Impress DAB substrate was applied for 2 mins. Slides were

then counterstained in Mayers haematoxylin and mounted in DPX.

Cell culture. Human PBMCs were isolated from leukocyte cones from healthy donors supplied by the NHS blood service. CD8 T-cells were isolated from

PBMCs using positive selection with MACS CD8 microbeads (Miltenyi).

Monocytes were enriched from PBMCs using MACS CD14 microbeads

(Miltenyi). Monocytes were differentiated into macrophages (macrophages or GM-MF), by incubating the CD14 cells for 7 days in X-VIVO15 (Lonza),

Pen/strep (Sigma-Aldrich), L-glutamine (Sigma-Aldrich) with 50 ng ml 1 GM-CSF (Miltenyi). Immature dendritic cells (imDCs or DCs) were obtained by culturing

CD14 cells for 45 days in the presence of 20100 ng ml 1 GM-CSF (Miltenyi/First Link) and 25 ng ml 1 IL-4 (Miltenyi/eBioscience). Where stated,

MAIT cells were sorted using a Beckman Coulter MoFlo XDP.

Viruses. DENV serotype 2, strain 16681, was grown in C6/36 cells in Leibovitzs L-15 medium with L-glutamine and supplemented with 2% fetal calf serum. Culture medium was centrifuged and stored at 80 C. The titres of virus were

determined by a focus-forming assay on Vero cells and expressed as focus-forming units per ml. Briey, virus was serially diluted and incubated with Vero cells for 2 h at 37 C. The monolayers were then overlaid with 1.5% carboxymethylcellulose and incubated at 37 C for 3 days. Virus foci were stained with anti-DENV E antibody (4G2) followed by peroxidase-conjugated anti-mouse Ig and visualized by the addition of DAB substrate. Infection rates of DENV infected DCs were assessed using an antibody detecting the non-structural DENV protein, NS3, by ow cytometry. The mouse monoclonal anti-DENV NS3 (E1D8) and anti-DENV E (4G2) were gifts from E. Harris and AFRIMS.

A total of 10 ng RNA transcribed from genotype 2a HCV strain J6CF-JFH1 (obtained from Prof. Bartenschlager62) was electroporated into Huh-7.5 cells (obtained from Apath) and were cultured for up to 3 weeks. Cell culture supernatants were collected for up to 20 days post electroporation centrifuged and concentrated using an Amicon Ultra-15 (Millipore). This inoculum was used to infect huh-7.5 cells. The HCV titre was determined by immunouorescence. Huh-7.5 cells were xed with methanolacetone, blocked with BSA/phosphate-buffered saline (PBS) solution, washed with PBS, stained with anti-HCV core primary antibody (Cambridge Biosciences), washed with PBS and Alexa Fluor 488 donkey polyclonal secondary antibody to Mouse IgG (ab150105, Abcam). To measure HCV replication a HCV luciferase reporter Jc1FLAGp7-nsGluc2A29 virus was used. Jc1FLAG(p7-nsGluc2A) was grown in nave Huh-7.5 cells63 and the infectivity titre was determined by limiting dilution titration on naive Huh-7.5 cells as median tissue culture infective dose (TCID50). To measure HCV replication, Huh-7.5 cells were plated in a 96-well plate and infected at an MOI of 0.1 for 6 h. After washing the Huh-7.5 cells, supernatants from (un)stimulated MAIT cells were added in the presence or absence of a neutralizing IFN-g (clone MD-1, 14-7317-85 eBioscience). Supernatants were collected 4 days post infection and luciferase expression determined using the Renilla Luciferase Assay System (Promega) and a Berthold TriStar2 multimode reader LB 942.

Inuenza A/WSN/33 (H1N1) virus (WSN) was grown in Madin Darby Bovine Kidney (MDBK, obtained from the European Collection of Cell Cultures) cells in Minimum Essential Medium Eagle (Sigma) with 2 mM L-glutamine and 0.5% fetal calf serum. To produce virus stocks, six 175 cm2 tissue culture asks containing sub-conuent MDBK cells were infected at low-multiplicity. After 48 h of culture in a 37 C humidied incubator, supernatants were collected and claried by low-speed centrifugation (30 min, 2,000 g and 30 min, 18,000 g, 4 C). Next, WSN was concentrated through a 30% sucrose cushion by ultracentrifugation (90 min, 112,000 g, 4 C). Finally, WSN was puried on a 3060% sucrose gradient by ultracentrifugation (150 min, 209,000 g, 4 C). The visible band of virus was drawn off with a needle. (Hutchinson, E. and Fodor, E., Nature ProtocolExchange, Purication of inuenza virions by haemadsorption and ultracentrifugation, http://www.nature.com/protocolexchange/protocols/3315

Web End =http://www.nature.com/protocolexchange/protocols/3315) 64,65. Plaque assays were performed on MDBK cells using standard techniques. Inuenza A (H3N2) virus A/England/691/2010 is a clinical strain isolated by Public Health England (PHE), as part of the MOSAIC project. Inuenza B virus, B/Florida/04/2006 was derived from reverse genetics systems by de novo synthesis (GeneArt).

In vitro virus experiments. Unless specied differently, for co-culture experiments using viruses, APCs were treated with virus at a MOI of 1, as determined based on the viral titre and number of cells plated, for 90 to 120 min at 37 C. Virus was washed off and isolated CD8 T-cells were added for an additional 1024 h,

unless specied differently. In the case of the DENV experiments, PBMCs were added instead of isolated CD8 T-cells. In the case of inuenza virus, WSN virus

was used, unless specied differently.

In vitro cytokine stimulations. For cytokine stimulation experiments, isolated CD8 T-cells or PBMCs were stimulated for 24 h with 50 ng ml 1 IL-12

(Miltenyi Biotech), 50 ng ml 1 IL-18 (R&D Systems Europe), 50 ng ml 1 IL-15 (Miltenyi Biotech) or 2,000 U ml 1 IFN-a (Sigma-Aldrich) or 50 ng ml 1 IFN-b (Miltenyi Biotech) or combinations thereof. For blocking experiments, neutralizing agents were added into the culture together with the CD8 T-cells or PBMCs.

Blocking antibodies against IL-12p40/70 (508804, eBioscience), IL-12p70 (MAB219-100, R&D), IL-15 (MAB2471, R&D) or IL-18 (D044-3, MBL International, USA), IFN-g (14-7317-85, eBioscience) or MR1 (361102, Biolegend)

were used at 510 mg ml 1. Type I IFN was blocked using 1 mg ml 1 B18R (34-8185-81, eBioscience).

Flow cytometry. Antibodies/dyes and dilutions used were: viability dye live/dead xable-violet (L34955, Invitrogen, 1:1250), CD3-eFluor450 (48-0038, eBioscience, 1:100), CD3-PECy7 (25-0038, eBioscience, 1:100), CD4-VioGreen (130-096-900, Miltenyi Biotech, 1:50), CD8-VioGreen (130-098-062, Miltenyi Biotech, 1:50), CD8-V450 (560347, BD, 1:50), CD8-PerCP.Cy5.5/PerCP (301032, Biolegend, 1:100), CD38-APC (555462, BD, 1:50), CD69-FITC (11-0699, eBioscience, 1:40), CD161-APC (130-098-908, Miltenyi Biotech, 1:100), CD161-PE (130-099-193, Miltenyi Biotech, 1:100), IFN-g-FITC (130-091-641, Miltenyi Biotech, 1:50),

IFN-g-APCCy7 (502529 Biolegend, 1:50), Va7.2-PE/PeCy7/APC/FITC

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(351705/351711/351707/351703, Biolegend, 1:50). Granzyme B-APC (MHGB05, Invitrogen), IL-18Ra-APC (17-7183-41, eBioscience, 1:50), TNF-a-PeCy7 (502929, Biolegend, 1:100). All data was acquired on a MACSQuant (Miltenyi Biotech) or a BD FACSVerse (BD) and analyzed on FlowJo (Tree Star Inc.). Gating strategy is shown in Supplementary Fig. 12.

Cytokine measurements. Cytokines levels in this study were measured from cell culture supernatants or heparin plasma samples. Cell culture supernatants from macrophages treated with mock, HCV or inuenza A were collected at 48 h. Cell culture supernatants from DCs treated with mock or DENV were collected at 42 h. Heparin plasma samples were collected from DENV infected patients at different phases of illness or from healthy controls. Cell culture supernatants and heparin plasma samples were frozen at 80

C until further use. Cytokine levels were analysed using Bio-Plex human cytokine kits (BioRad) and acquired on a Luminex 100 (Luminex) or a Bio-Plex 200 reader (BioRad) according to the manufacturers instructions.

Statistical analysis. Statistical analysis was carried out with GraphPad Prism software. Statistical signicance was reported as ns P40.05; *Pr0.05; **Pr0.01;

***Pr0.001; ****Pr0.0001. Error bars on graphs represent s.e.m. MannWhitney test was used to calculate signicance levels between two groups. Wilcoxon matched-paired test was used to calculate signicance levels between two paired groups. For comparisons of means from multiple groups against one control group the KruskalWallis with Dunns multiple comparison post-test analysis was performed. Spearman-rank correlation analysis was used to calculate correlations. Sample sizes were adequate to detect large effects between groups, as determined by the reproducibility and variability of each particular experiment and limited by the availability of patient samples. No randomization or blinding was used.

References

1. Porcelli, S., Yockey, C. E., Brenner, M. B. & Balk, S. P. Analysis of T cell antigen receptor (TCR) expression by human peripheral blood CD4-8- alpha/betaT cells demonstrates preferential use of several V beta genes and an invariant TCR alpha chain. J. Exp. Med. 178, 116 (1993).

2. Tilloy, F. et al. An invariant T cell receptor alpha chain denes a novel TAP-independent major histocompatibility complex class Ib-restricted alpha/ beta T cell subpopulation in mammals. J. Exp. Med. 189, 19071921 (1999).

3. Dusseaux, M. et al. Human MAIT cells are xenobiotic-resistant, tissue-targeted, CD161hi IL-17-secreting T cells. Blood 117, 12501259 (2011).

4. Treiner, E. et al. Selection of evolutionarily conserved mucosal-associated invariant T cells by MR1. Nature 422, 164169 (2003).

5. Tang, X.-Z. et al. IL-7 licenses activation of human liver intrasinusoidal mucosal-associated invariant T cells. J. Immunol. 190, 31423152 (2013).

6. Martin, E. et al. Stepwise development of MAIT cells in mouse and human. PLoS Biol. 7, e54 (2009).

7. Kjer-Nielsen, L. et al. MR1 presents microbial vitamin B metabolites to MAIT cells. Nature 491, 717723 (2012).

8. Corbett, A. J. et al. T-cell activation by transitory neo-antigens derived from distinct microbial pathways. Nature 509, 361365 (2014).

9. Georgel, P., Radosavljevic, M., Macquin, C. & Bahram, S. The non-conventional MHC class I MR1 molecule controls infection by Klebsiella pneumoniae in mice. Mol. Immunol. 48, 769775 (2011).

10. Chua, W.-J. et al. Polyclonal MAIT Cells have unique innate functions in bacterial infection. Infect. Immun. 80, 32563267 (2012).

11. Rahimpour, A. et al. Identication of phenotypically and functionally heterogeneous mouse mucosal-associated invariant T cells using MR1 tetramers. J. Exp. Med. 212, 10951108 (2015).

12. Ussher, J. E. et al. CD161 CD8 T cells, including the MAIT cell subset, are specically activated by IL-12 IL-18 in a TCR-independent manner.

Eur. J. Immunol. 44, 195203 (2014).13. Billerbeck, E. et al. Analysis of CD161 expression on human CD8 T cells

denes a distinct functional subset with tissue-homing properties. Proc. Natl Acad. Sci. USA 107, 30063011 (2010).14. Takahashi, T., Dejbakhsh-Jones, S. & Strober, S. Expression of CD161 (NKR-P1A) denes subsets of human CD4 and CD8 T cells with different functional activities. J. Immunol. 176, 211216 (2006).

15. Lanier, L. L., Chang, C. & Phillips, J. H. Human NKRP1A. A disulde-linked homodimer of the C-type lectin superfamily expressed by a subset of NK and T lymphocytes. J. Immunol. 153, 24172428 (1994).

16. Fergusson, J. R. et al. CD161 denes a transcriptional and functional phenotype shared across distinct human T cell lineages. Cell Rep. 9, 10751088 (2014).

17. Ivanov, I. I. et al. The orphan nuclear receptor RORgammat directs the differentiation program of proinammatory IL-17 T helper cells. Cell 126,

11211133 (2006).18. Singh, S. P. et al. PLZF regulates CCR6 and is critical for the acquisition and maintenance of the Th17 phenotype in human cells. J. Immunol. 194, 43504361 (2015).

19. Savage, A. K. et al. The transcription factor PLZF directs the effector program of the NKT cell lineage. Immunity 29, 391403 (2008).

20. Gleimer, M., von Boehmer, H. & Kreslavsky, T. PLZF controls the expression of a limited number of genes essential for NKT cell function. Front. Immunol. 3, 374 (2012).

21. Patel, O. et al. Recognition of vitamin B metabolites by mucosal-associated invariant T cells. Nat. Commun. 4, 2142 (2013).

22. Kurioka, A. et al. MAIT cells are licensed through granzyme exchange to kill bacterially sensitized targets. Mucosal Immunol. 8, 429440 (2015).

23. Le Bourhis, L., Dusseaux, M. & Bohineust, A. MAIT cells detect and efciently lyse bacterially-infected epithelial cells. PLoS Pathog. 9, e1003681 (2013).

24. Jo, J. et al. Toll-like receptor 8 agonist and bacteria trigger potent activation of innate immune cells in human liver. PLoS Pathog. 10, e1004210 (2014).

25. Sattler, A., Dang-Heine, C., Reinke, P. & Babel, N. IL-15 dependent induction of IL-18 secretion as a feedback mechanism controlling human MAIT-cell effector functions. Eur. J. Immunol. 45, 22862298 (2015).

26. Chattergoon, M. A. et al. HIV and HCV activate the inammasome in monocytes and macrophages via endosomal Toll-like receptors without induction of type 1 interferon. PLoS Pathog. 10, e1004082 (2014).

27. Hidmark, A. S. et al. Early alpha/beta interferon production by myeloid dendritic cells in response to UV-inactivated virus requires viral entry and interferon regulatory factor 3 but not MyD88. J. Virol. 79, 1037610385 (2005).

28. Short, K. R., Brooks, A. G., Reading, P. C. & Londrigan, S. L. The fate of inuenza A virus after infection of human macrophages and dendritic cells.J. Gen. Virol. 93, 23152325 (2012).29. Marukian, S. et al. Cell culture-produced hepatitis C virus does not infect peripheral blood mononuclear cells. Hepatology 48, 18431850 (2008).

30. van Wilgenburg, B., Browne, C., Vowles, J. & Cowley, S. A. Efcient, long term production of monocyte-derived macrophages from human pluripotent stem cells under partly-dened and fully-dened conditions. PLoS ONE 8, e71098 (2013).

31. Stonier, S. W. & Schluns, K. S. Trans-presentation: a novel mechanism regulating IL-15 delivery and responses. Immunol. Lett. 127, 8592 (2010).

32. Liang, Z. et al. Activation of Toll-like receptor 3 impairs the dengue virus serotype 2 replication through induction of IFN-b in cultured hepatoma cells.

PLoS ONE 6, e23346 (2011).33. Garca-Sastre, A. Induction and evasion of type I interferon responses by inuenza viruses. Virus Res. 162, 1218 (2011).

34. Castet, V. et al. Alpha interferon inhibits hepatitis C virus replication in primary human hepatocytes infected in vitro. J. Virol. 76, 81898199 (2002).

35. Symons, J. A., Alcam, A. & Smith, G. L. Vaccinia virus encodes a soluble type I interferon receptor of novel structure and broad species specicity. Cell 81, 551560 (1995).

36. Foster, G. R. et al. Efcacy of sofosbuvir plus ribavirin with or without peginterferon-alfa in patients with HCV genotype 3 infection and treatment-experienced patients with cirrhosis and HCV genotype 2 infection. Gastroenterology 149, 14621470 (2015).

37. Le Bourhis, L. et al. Antimicrobial activity of mucosal-associated invariant T cells. Nat. Immunol. 11, 701708 (2010).

38. Cosgrove, C. et al. Early and nonreversible decrease of CD161 /MAIT

cells in HIV infection. Blood 121, 951961 (2013).39. Reilly, E. C., Wands, J. R. & Brossay, L. Cytokine dependent and independent iNKT cell activation. Cytokine 51, 227231 (2010).

40. Paget, C. et al. Potential role of invariant NKT cells in the control of pulmonary inammation and CD8 T cell response during acute inuenza A virus H3N2

pneumonia. J. Immunol. 186, 55905602 (2011).41. Cui, Y. et al. Mucosal-associated invariant T cell-rich congenic mouse strain allows functional evaluation. J. Clin. Invest. 125, 41714185 (2015).

42. Reantragoon, R. et al. Antigen-loaded MR1 tetramers dene T cell receptor heterogeneity in mucosal-associated invariant T cells. J. Exp. Med. 210, 23052320 (2013).

43. Leeansyah, E. et al. Activation, exhaustion, and persistent decline of the antimicrobial MR1-restricted MAIT-cell population in chronic HIV-1 infection. Blood 121, 11241135 (2013).

44. Fernandez, C. S. et al. MAIT cells are depleted early but retain functional cytokine expression in HIV infection. Immunol. Cell Biol. 93, 177188 (2015).

45. Gherardin, N. A. et al. Diversity of T cells restricted by the MHC class i-related molecule MR1 facilitates differential antigen recognition. Immunity 44, 3245 (2016).

46. Fergusson, J. R. et al. CD161 denes a transcriptional and functional phenotype across distinct human T cell lineages. Cell Rep. 9, 10751088 (2014).

47. Matikainen, S. et al. IFN-alpha and IL-18 synergistically enhance IFN-gamma production in human NK cells: differential regulation of Stat4 activation and IFN-gamma gene expression by IFN-alpha and IL-12. Eur. J. Immunol. 31, 22362245 (2001).

48. Jian, Y.-R., Chang, S.-Y., Lin, P.-Y., Yang, Y.-H. & Chuang, Y.-H. Inactivated inuenza virus vaccine is efcient and reduces IL-4 and IL-6 in allergic asthma mice. Inuenza Other Respi. Viruses 7, 12101217 (2013).

10 NATURE COMMUNICATIONS | 7:11653 | DOI: 10.1038/ncomms11653 | http://www.nature.com/naturecommunications

Web End =www.nature.com/naturecommunications

NATURE COMMUNICATIONS | DOI: 10.1038/ncomms11653 ARTICLE

49. Thimme, R. Determinants of viral clearance and persistence during acute hepatitis C virus infection. J. Exp. Med. 194, 13951406 (2001).

50. Guidotti, L. G. Viral clearance without destruction of infected cells during acute HBV infection. Science 284, 825829 (1999).

51. Pang, T., Cardosa, M. J. & Guzman, M. G. Of cascades and perfect storms: the immunopathogenesis of dengue haemorrhagic fever-dengue shock syndrome (DHF/DSS). Immunol. Cell Biol. 85, 4345 (2007).

52. Wang, S. et al. Inuenza virus-cytokine-protease cycle in the pathogenesis of vascular hyperpermeability in severe inuenza. J. Infect. Dis. 202, 9911001 (2010).

53. Murray, R. J. et al. Community-acquired pneumonia due to pandemic A(H1N1)2009 inuenzavirus and methicillin resistant Staphylococcus aureus co-infection. PLoS ONE 5, e8705 (2010).

54. Morens, D. M., Taubenberger, J. K. & Fauci, A. S. Predominant role of bacterial pneumonia as a cause of death in pandemic inuenza: implications for pandemic inuenza preparedness. J. Infect. Dis. 198, 962970 (2008).55. Grart, S. et al. Human iNKT and MAIT cells exhibit a PLZF-dependent proapoptotic propensity that is counterbalanced by XIAP. Blood 121, 614623 (2013).

56. Fergusson, J. R., Fleming, V. M. & Klenerman, P. CD161-expressing human T cells. Front. Immunol. 2, 36 (2011).

57. Tsai, C.-Y. et al. Type I IFNs and IL-18 regulate the antiviral response of primary human gd T cells against dendritic cells infected with Dengue virus.

J. Immunol. 194, 38903900 (2015).58. Fergusson, J. R. et al. CD161(int)CD8 T cells: a novel population of highly

functional, memory CD8 T cells enriched within the gut. Mucosal Immunol.

9, 401413 (2015).59. Denney, L. et al. Reduction of natural killer but not effector CD8 T lymphocytes in three consecutive cases of severe/lethal H1N1/09 inuenza A virus infection. PLoS ONE 5, e10675 (2010).

60. Zhao, Y. et al. High levels of virus-specic CD4 T cells predict severe

pandemic inuenza A virus infection. Am. J. Respir. Crit. Care Med. 186, 12921297 (2012).61. Foster, G. et al. Treatment of decompensated HCV cirrhosis in patients with diverse genotypes: 12 weeks sofosbuvir and NS5A inhibitors with/without ribavirin is effective in HCV genotypes 1 and 3. J. Hepatol. 62 (2015).

62. Pietschmann, T. et al. Construction and characterization of infectious intragenotypic and intergenotypic hepatitis C virus chimeras. Proc. Natl Acad. Sci. USA 103, 74087413 (2006).

63. Lindenbach, B. D. et al. Complete replication of hepatitis C virus in cell culture. Science 309, 623626 (2005).

64. Hutchinson, E. C. et al. Conserved and host-specic features of inuenza virion architecture. Nat. Commun. 5, 4816 (2014).

65. Hutchinson, E. C. et al. Mapping the phosphoproteome of inuenza A and B viruses by mass spectrometry. PLoS Pathog. 8, e1002993 (2012).

Acknowledgements

This research was supported by the NIHR Biomedical Research Program (Oxford), the Wellcome Trust (WT091663MA) and National Institutes for Health Research Biomedical Research Centre (Imperial College), the NIH (U19AI082630) and the Oxford Martin School. I.S. contribution was supported by the Wellcome Trust programme Grant PS2186_WMII. The STOP-HCV consortium is funded by a grant from the Medical Research Council. E.C.H. contribution was supported by MRC programme Grant MR/K000241/1 awarded to Ervin Fodor. L.H. contribution was supported by MRC Project Grant G0600371. Y.H.O. contribution was supported by MRC intermediate fellowship programme. The authors wish to acknowledge the role of the HCV Research UK Biobank in collecting and making available the samples/data used in the generation of this publication. We thank Wendy Barclay for providing Inuenza A (H3N2) virus A/England/691/2010 and Inuenza B virus, B/Florida/04/2006. We thank Mark Ainsworth for overseeing the HCV patients at the JR hospital. We thank Amin Moghaddam, Jacob Hurst and Leo Swadling for their scientic advice. We also want to acknowledge Nattaya Tangthawornchaikul and Thaneeya Duangchinda.

Author contributions

Conceived and designed the experiments: B.W, I.S., P.K., C.W., G.S., J.M.; Performed the experiments: B.W, I.S, C.L, T.L, A.K., C.K., P.F, F.T.; Analyzed the data: B.W., I.S., A.K., C.W.; Contributed to the collection of clinical samples: S.C., S.V., W.L., P.M., D.Y., L.D., STOP-HCV, P.F., M.G., Y.O., L.H.; Contributed other reagents/materials/analysis tools: E.H., N.R., S.M.L., L.B.D., M.D.; Contributed to the writing of this manuscript: P.K., B.W., C.W., G.S., I.S., J.M., L.B.D.

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STOP-HCV consortiumEleanor Barnes14, Jonathan Ball15, Gary Burgess16, Graham Cooke17, John Dillon18, Charles Gore19, Graham Foster20, Neil Guha15, Rachel Halford19, Cham Herath21, Chris Holmes22, Anita Howe23, Emma Hudson14, William Irving15, Salim Khakoo24, Diana Koletzki25, Natasha Martin26, Tamyo Mbisa27, Jane McKeating28, John McLauchlan29, Alec Miners30, Andrea Murray31, Peter Shaw32, Peter Simmonds14, Chris Spencer33, Paul Targett-Adams34, Emma Thomson29, Peter Vickerman35 & Nicole Zitzmann36

14University of Oxford, Peter Medawar Building for Pathogen Research, South Parks Road, Oxford OX1 3SY, UK; 15University of Nottingham, Queens Medical Centre, Nottingham, NG7 2UH; 16Conatus Pharmaceuticals, 16745 West Bernardo Drive, Suite 200, San Diego, California 92127, USA; 17Imperial College London, Wright Fleming Institute, London, UK; 18University of Dundee, Ninewells Hospital & Medical School, Dundee, DD1 9SY, UK; 19Hepatitis C Trust, 27 Crosby Row, London SE1 3YD, UK; 20Queen Marys University of London, 4 Newark Street, London E1 4AT, UK; 21Gilead Sciences, Stockley Park, 2 Roundwood Avenue, Middlesex UB11 1AF, UK; 22University of Oxford, 2429 St Giles, Oxford OX1 3LB, UK; 23BC Centre for Excellence in HIV/AIDS, St Pauls Hospital, 6081081 Burrard Street, Vancouver, British Columbia, Canada V6Z 1Y6; 24University of Southampton, University Road, Southampton, SO17 1BJ, UK; 25Janssen Diagnostics, Turnhoutseweg, 30, 2340 Beerse, Belgium; 26UC San Diego, La Jolla, California 92093-0507, USA; 27Public Health England, 61 Colindale Avenue, London NW9 5EQ, UK; 28University of Birmingham, Centre for Human Virology, Edgbaston, Birmingham, B15 2TT, UK; 29University of Glasgow, MRC-CVR, 464 Bearsden Road, Glasgow, G61 1QH, UK; 30London School of Hygiene & Tropical Medicine, 1517 Tavistock Place, London, WC1H 9SH, UK; 31OncImmune Limited, Clinical Sciences Building, Nottingham City Hospital, Hucknall Road, Nottingham NG5 1PB, UK; 32Merck & Co., Inc., Kenilworth, New Jersey 07033, USA; 33University of Oxford, Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford OX3 7BN, UK; 34Medivir AB, Box 1086, 141 22 Huddinge, Sweden; 35University of Bristol, Oakeld House, Oakeld Grove, Clifton BS8 2BN, UK; 36University of Oxford, South Parks Road, Oxford OX1 3QU, UK.

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Mucosal-associated invariant T (MAIT) cells are abundant in humans and recognize bacterial ligands. Here, we demonstrate that MAIT cells are also activated during human viral infections in vivo. MAIT cells activation was observed during infection with dengue virus, hepatitis C virus and influenza virus. This activation--driving cytokine release and Granzyme B upregulation--is TCR-independent but dependent on IL-18 in synergy with IL-12, IL-15 and/or interferon-α/β. IL-18 levels and MAIT cell activation correlate with disease severity in acute dengue infection. Furthermore, HCV treatment with interferon-α leads to specific MAIT cell activation in vivo in parallel with an enhanced therapeutic response. Moreover, TCR-independent activation of MAIT cells leads to a reduction of HCV replication in vitro mediated by IFN-γ. Together these data demonstrate MAIT cells are activated following viral infections, and suggest a potential role in both host defence and immunopathology.

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Title

MAIT cells are activated during human viral infections

Author

Van Wilgenburg, Bonnie; Scherwitzl, Iris; Hutchinson, Edward C; Leng, Tianqi; Kurioka, Ayako; Kulicke, Corinna; De Lara, Catherine; Cole, Suzanne; Vasanawathana, Sirijitt; Limpitikul, Wannee; Malasit, Prida; Young, Duncan; Denney, Laura; Moore, Michael D; Fabris, Paolo; Giordani, Maria Teresa; Oo, Ye Htun; Laidlaw, Stephen M; Dustin, Lynn B; Ho, Ling-pei; Thompson, Fiona M; Ramamurthy, Narayan; Mongkolsapaya, Juthathip; Willberg, Christian B; Screaton, Gavin R; Klenerman, Paul

Pages

11653

Publication year

2016

Publication date

Jun 2016

Publisher

Nature Publishing Group

e-ISSN

20411723

Source type

Scholarly Journal

Language of publication

English

DOI

https://doi.org/10.1038/ncomms11653

ProQuest document ID

1799027546

MAIT cells are activated during human viral infections

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