C57.BL/6 and Adamts5^-/- mice were intranasally (i.n.) infected (10⁴ pfu/mouse) with X31 (H3N2) influenza virus. Spleens were removed at day 7 and day 10 p.i., and CD8⁺ T cell responses determined. Total CD4⁺ and CD8⁺ T cells numbers were calculated at days (A) 7 and (B) 10 p.i. Influenza-specific D^bNP_366-374 CD8⁺ and D^bPA_224-233 CD8⁺ tetramer-positive T cell numbers were measured at days (C) 7 and (D) 10 p.i. Functional influenza-specific D^bNP_366-374⁺IFNγ⁺CD8⁺ and D^bPA_224-233⁺IFNγ⁺CD8⁺ T cell activity was determined by ICS, and total IFNγ⁺ T cells enumerated at days (E) 7 and (F) 10 after infection. WT denotes C57.BL/6. The results are expressed as means ± SD, and statistical significance (relative to C57.BL/6) was determined by Student’s t test (*p ≤ 0.05, **p ≤ 0.01, n = 5 representing three experiments). Underlying data are provided in S1 Data.

C57.BL/6 and Adamts5^-/- mice were infected i.n. with 10⁴pfu X31 (H3N2) influenza virus. Lungs were removed at days 7 and 10 p.i., and CD8⁺ T cell immunity characterised. Total CD4⁺ and CD8⁺ T cell numbers are shown at days (A) 7 and (B) 10 p.i. Influenza-specific tetramer⁺ D^bNP_366-374⁺ CD8⁺ and D^bPA_224-233⁺ CD8⁺ T cell responses were enumerated at days (C) 7 and (D) 10 p.i. CD8⁺ T cell functionality was assessed by ICS and D^bNP_366-374⁺IFNγ⁺CD8⁺ and D^bPA_224-233⁺IFNγ⁺CD8⁺ T cell responses enumerated at days (E) 7 and (F) 10 p.i. Wildtype denotes C57.BL/6 mice. The results are expressed as means ± SD, and statistical significance (relative to C57.BL/6) was determined by Student’s t test (* = p ≤ 0.05, ** = p ≤ 0.01, n = 5 representing three experiments). Underlying data are provided in S1 Data.

C57.BL/6 and Adamts5^-/- mice were infected i.n. with 10⁴ pfu/mouse X31 (H3N2) influenza virus. MLNs were removed, pooled, and processed at days 7 and 10 p.i., and single-cell suspensions analysed for influenza-specific immunity. Total CD4⁺ and CD8⁺ T cell numbers were determined at days (A) 7 and (B) 10 p.i. Influenza-specific D^bNP_366-374⁺ CD8⁺ and D^bPA_224-233⁺ CD8⁺ tetramer positive T cells were enumerated at days (C) 7 and (D) 10 p.i. CD8⁺ T cell functionality was measured using ICS. Influenza-specific D^bNP_366-374⁺IFNγ⁺CD8⁺ and D^bPA_224-233⁺IFNγ⁺CD8⁺ T cell responses were characterised at days (E) 7 and (F) 10 p.i. Results are expressed as total pooled means from five mice repeated three times. Wildtype denotes C57.BL/6 mice. Underlying data are provided in S1 Data.

Spleen and MLNs were removed from C57.BL/6, Adamts5^-/-Vcan^+/+, and Adamts5^-/-Vcan^+/hdf mice and processed at day 10 p.i., and single cell suspensions were analysed for influenza-specific immunity. (A) Total CD8⁺ T cell numbers were determined at day 10 p.i. in the spleen. (B) Influenza-specific D^bNP_366-374⁺ CD8⁺ and D^bPA_224-233⁺ CD8⁺ tetramer positive T cells in the spleen were enumerated at day 10 p.i. CD8⁺ T cell functionality was measured using ICS. (C) Influenza specific D^bNP_366-374⁺IFNγ⁺CD8⁺ and D^bPA_224-233⁺IFNγ⁺CD8⁺ T cell responses were characterised in the spleen at days 10 p.i. (D) Total CD8⁺ T cell numbers were assessed at day 10 p.i. in the pooled MLN. (E) Influenza-specific D^bNP_366-374⁺ CD8⁺ and D^bPA_224-233⁺ CD8⁺ tetramer positive T cells in the pooled MLN were enumerated at day 10 p.i. CD8⁺ T cell functionality was measured using ICS. (F) Influenza-specific D^bNP_366-374⁺IFNγ⁺CD8⁺ and D^bPA_224-233⁺IFNγ⁺CD8⁺ T cell responses were characterised in the pooled MLN at day 10 p.i. The results are expressed as means ± SD (spleen data) or as pooled means (MLN data), and statistical significance (relative to C57.BL/6 mice) was determined by one-way ANOVA (*p ≤ 0.05, ***p ≤ 0.005 relative to C57.BL/6, n = 5 representing three individual experiments). WT denotes C57.BL/6 mice and ts5^-/- denotes Adamts5^-/-. (G) Our model for ADAMTS5 enzyme activity and T cell migration proposes that versican can inhibit T cell effector function by acting as a physical block. Cleavage of versican by ADAMTS5 removes the ECM blockade, allowing migration (top panel). Moreover, versican accumulation in the absence of ADAMTS enzyme activity results in T cell clustering (bottom panel). Underlying data are provided in S1 Data.