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Structural basis for IL-33 recognition and its antagonism by the helminth effector protein HpARI2

Jamwal, Abhishek, Colomb, Florent, McSorley, Henry J., Higgins, Matthew K.·2024·Nature Communications
DOI 10.1038/s41467-024-49550-0PMC PMC11189471
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Abstract

IL-33 plays a significant role in inflammation, allergy, and host defence against parasitic helminths. The model gastrointestinal nematode Heligmosomoides polygyrus bakeri secretes the Alarmin Release Inhibitor HpARI2, an effector protein that suppresses protective immune responses and asthma in its host by inhibiting IL-33 signalling. Here we reveal the structure of HpARI2 bound to mouse IL-33. HpARI2 contains three CCP-like domains, and we show that it contacts IL-33 primarily through the second and third of these. A large loop which emerges from CCP3 directly contacts IL-33 and structural comparison shows that this overlaps with the binding site on IL-33 for its receptor, ST2, preventing formation of a signalling complex. Truncations of HpARI2 which lack the large loop from CCP3 are not able to block IL-33-mediated signalling in a cell-based assay and in an in vivo female mouse model of asthma. This shows that direct competition between HpARI2 and ST2 is responsible for suppression of IL-33-dependent responses.

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Fig. 1 The CCP1 domain does not contribute to the effector function of HpARI2. a Schematic representation of the three CCP-like domains of HpARI2 (CCP1-3). b Surface plasmin resonance analysis of the binding of full-length HpARI2 (upper panel), HpARI2_CCP1/2 (middle panel) and HpARI_CCP2/3 (lower panel) (red dashed lines) to mIL-33 were generated by flowing 2-fold serial dilutions of mIL-33 starting from 40 nM (over HpARI2), 200 nM (over HpARI2_CCP2/3), and 1000 nM (over HpARI2_CCP1/2) over surfaces immobilised with HpARI2 truncations. Indicated KD values were deduced from Langmuir fit (red dashed lines) and the experiments were conducted twice (n = 2). c Overview of the cellular assay for assessing effector function of HpARI2s. Mouse bone marrow cells were isolated and cultured with IL-2, IL-7, and IL-33 in the presence of HpARI2 truncations at various concentrations, followed by measurement of mIL-5 levels in cultured supernatants. d Concentration-response curves for the inhibition of signalling by various HpARI2 forms as determined by IL-5 production. Error bars indicate mean +/- standard error of the mean of 4 technical replicates. Data are representative of 3 repeat experiments (n = 3). Source data are provided as a Source Data file.
Fig. 1 The CCP1 domain does not contribute to the effector function of HpARI2. a Schematic representation of the three CCP-like domains of HpARI2 (CCP1-3). b Surface plasmin resonance analysis of the binding of full-length HpARI2 (upper panel), HpARI2_CCP1/2 (middle panel) and HpARI_CCP2/3 (lower panel) (red dashed lines) to mIL-33 were generated by flowing 2-fold serial dilutions of mIL-33 starting from 40 nM (over HpARI2), 200 nM (over HpARI2_CCP2/3), and 1000 nM (over HpARI2_CCP1/2) over surfaces immobilised with HpARI2 truncations. Indicated KD values were deduced from Langmuir fit (red dashed lines) and the experiments were conducted twice (n = 2). c Overview of the cellular assay for assessing effector function of HpARI2s. Mouse bone marrow cells were isolated and cultured with IL-2, IL-7, and IL-33 in the presence of HpARI2 truncations at various concentrations, followed by measurement of mIL-5 levels in cultured supernatants. d Concentration-response curves for the inhibition of signalling by various HpARI2 forms as determined by IL-5 production. Error bars indicate mean +/- standard error of the mean of 4 technical replicates. Data are representative of 3 repeat experiments (n = 3). Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 2 The structure of the HpARI2_CCP2/3:IL-33 complex. a Two views of HpARI2_CCP2/3 (CCP2 in cyan and CCP3 in blue) bound to mIL-33 (green). b The structure of HpARI2_CCP2/3 (CCP2 in cyan and CCP3 in blue) with disulphide bonds (DS1-4) shown as yellow sticks. The domain schematic on the left gives an overview of the structure with precise domain boundaries and patterns of disulphide bonds within each domain derived from the crystal structure. c Two views of the HpARI2_CCP2/3:mIL-33 complex with boxes below showing zoomed views of these interfaces. Interfacial residues are shown as sticks and hydrogen or polar interactions are indicated with a dashed line. d Binding of mIL-33 to HpARI2 variants determined by surface plasmon resonance analysis at a HpARI2 concentration of 100 nM. Bars represent the average and standard deviation for three independent measurements (n = 3). e Surface plasmon resonance analysis of HpARI2-LD binding to mIL-33. Multicycle response curves for two-fold mIL-33 dilutions starting from 2 µM (n = 2). Source data are provided as a Source Data file.
Fig. 2 The structure of the HpARI2_CCP2/3:IL-33 complex. a Two views of HpARI2_CCP2/3 (CCP2 in cyan and CCP3 in blue) bound to mIL-33 (green). b The structure of HpARI2_CCP2/3 (CCP2 in cyan and CCP3 in blue) with disulphide bonds (DS1-4) shown as yellow sticks. The domain schematic on the left gives an overview of the structure with precise domain boundaries and patterns of disulphide bonds within each domain derived from the crystal structure. c Two views of the HpARI2_CCP2/3:mIL-33 complex with boxes below showing zoomed views of these interfaces. Interfacial residues are shown as sticks and hydrogen or polar interactions are indicated with a dashed line. d Binding of mIL-33 to HpARI2 variants determined by surface plasmon resonance analysis at a HpARI2 concentration of 100 nM. Bars represent the average and standard deviation for three independent measurements (n = 3). e Surface plasmon resonance analysis of HpARI2-LD binding to mIL-33. Multicycle response curves for two-fold mIL-33 dilutions starting from 2 µM (n = 2). Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 3 HpARI2 competes for binding to IL-33 with the receptor ST2. a Assessment of the binding of the mST2 ectodomain (orange curve) to mIL-33 (green), HpARI2:mIL-33 (blue) and HpARI2_CCP2/3:mIL-33 (cyan) in a size exclusion column. The SEC curves are representative of two independent experiments (n = 2). Coomassie-strained gels show fractions highlighted on the size exclusion traces by dotted blue lines. b Overlay of the HpARI2_CCP2/3:mL33 crystal structure on that of the mIL-33:mST2 complex (5VI4 (IL33:ST2:IL-1RAcP)), revealing a steric clash between CCP3 and ST2. ST2 is orange, mIL-33 is green and the two CCP domains of HpARI2 are cyan (CCP2) and blue (CCP3). The dotted box to the right shows a close view of the complex. c Measurement by surface plasmon resonance of the binding of mIL-33 to immobilised ST2-Fc in the presence of HpARI2 and variants. Bars show the mean from three independent measurements (n = 3) with error bars showing standard deviation. Source data are provided as a Source Data file.
Fig. 3 HpARI2 competes for binding to IL-33 with the receptor ST2. a Assessment of the binding of the mST2 ectodomain (orange curve) to mIL-33 (green), HpARI2:mIL-33 (blue) and HpARI2_CCP2/3:mIL-33 (cyan) in a size exclusion column. The SEC curves are representative of two independent experiments (n = 2). Coomassie-strained gels show fractions highlighted on the size exclusion traces by dotted blue lines. b Overlay of the HpARI2_CCP2/3:mL33 crystal structure on that of the mIL-33:mST2 complex (5VI4 (IL33:ST2:IL-1RAcP)), revealing a steric clash between CCP3 and ST2. ST2 is orange, mIL-33 is green and the two CCP domains of HpARI2 are cyan (CCP2) and blue (CCP3). The dotted box to the right shows a close view of the complex. c Measurement by surface plasmon resonance of the binding of mIL-33 to immobilised ST2-Fc in the presence of HpARI2 and variants. Bars show the mean from three independent measurements (n = 3) with error bars showing standard deviation. Source data are provided as a Source Data file.CC BY · PubMed Central
Fig. 4 Assessing the effect of HpARI2 mutants on IL-33 signalling and function. a Mouse bone marrow cells were cultured for 5 days with IL-2 (10 ng/ml), IL-7 (10 ng/ml) and IL-33 (1 ng/ml) in the presence of a range of concentrations of HpARI2 or its N69, N70, N152 or LD mutants. Levels of IL-13 (left) and IL-5 (right) in culture supernatants were measured by ELISA. Error bars indicate mean +/- SEM of 6 technical replicates per condition. Data is representative of 3 repeat experiments. b The left panel is a schematic showing that Alternaria allergen (50 μg), together with HpARI2 WT, N70 or LD mutants (10 μg) were intranasally administered to BALB/c female mice. The right depicts calculated cell numbers of SiglecFhiCD11c–CD45+ live cells in right lobes of lung. Data are pooled from 2 repeat experiments for n = 7, except for the PBS control which has n = 3. Error bars indicate mean +/- SEM. One-way ANOVA with Sidak’s multiple comparisons test was used to compare each group to Alt control ****p < 0.0001, ns = p > 0.05. Source data are provided as a Source Data file.
Fig. 4 Assessing the effect of HpARI2 mutants on IL-33 signalling and function. a Mouse bone marrow cells were cultured for 5 days with IL-2 (10 ng/ml), IL-7 (10 ng/ml) and IL-33 (1 ng/ml) in the presence of a range of concentrations of HpARI2 or its N69, N70, N152 or LD mutants. Levels of IL-13 (left) and IL-5 (right) in culture supernatants were measured by ELISA. Error bars indicate mean +/- SEM of 6 technical replicates per condition. Data is representative of 3 repeat experiments. b The left panel is a schematic showing that Alternaria allergen (50 μg), together with HpARI2 WT, N70 or LD mutants (10 μg) were intranasally administered to BALB/c female mice. The right depicts calculated cell numbers of SiglecFhiCD11c–CD45+ live cells in right lobes of lung. Data are pooled from 2 repeat experiments for n = 7, except for the PBS control which has n = 3. Error bars indicate mean +/- SEM. One-way ANOVA with Sidak’s multiple comparisons test was used to compare each group to Alt control ****p < 0.0001, ns = p > 0.05. Source data are provided as a Source Data file.CC BY · PubMed Central

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  1. McSorley, H. J. & Smyth, D. J. IL-33: A central cytokine in helminth infections. Semin Immunol. 53, 101532 (2021). · DOI
  2. Liew, F. Y., Girard, J. P. & Turnquist, H. R. Interleukin-33 in health and disease. Nat. Rev. Immunol. 16, 676–689 (2016). · DOI
  3. Roussel, L., Erard, M., Cayrol, C. & Girard, J. P. Molecular mimicry between IL-33 and KSHV for attachment to chromatin through the H2A-H2B acidic pocket. EMBO Rep. 9, 1006–1012 (2008). · DOI
  4. Cayrol, C. & Girard, J. P. Interleukin-33 (IL-33): a critical review of its biology and the mechanisms involved in its release as a potent extracellular cytokine. Cytokine 156, 155891 (2022). · DOI
  5. Chan, B. C. L., Lam, C. W. K., Tam, L. S. & Wong, C. K. IL33: Roles in allergic inflammation and therapeutic perspectives. Front. Immunol. 10, 364 (2019). · DOI
  6. Schmitz, J. et al. IL-33, an interleukin-1-like cytokine that signals via the IL-1 receptor-related protein ST2 and induces T helper type 2-associated cytokines. Immunity 23, 479–490 (2005). · DOI
  7. Lefrancais, E. et al. Central domain of IL-33 is cleaved by mast cell proteases for potent activation of group-2 innate lymphoid cells. Proc. Natl Acad. Sci. USA 111, 15502–15507 (2014). · DOI
  8. Gunther, S. et al. IL-1 family cytokines use distinct molecular mechanisms to signal through their shared co-receptor. Immunity 47, 510–523.e4 (2017). · DOI
  9. Cohen, E. S. et al. Oxidation of the alarmin IL-33 regulates ST2-dependent inflammation. Nat. Commun. 6, 8327 (2015). · DOI
  10. Maizels, R. M. & McSorley, H. J. Regulation of the host immune system by helminth parasites. J. Allergy Clin. Immunol. 138, 666–675 (2016). · DOI · Paper page
  11. Maizels, R. M. Regulation of immunity and allergy by helminth parasites. Allergy 75, 524–534 (2020). · DOI · Paper page
  12. Johnston, C. J. C. et al. A structurally distinct TGF-beta mimic from an intestinal helminth parasite potently induces regulatory T cells. Nat. Commun. 8, 1741 (2017). · DOI · Paper page
  13. Vacca, F. et al. A helminth-derived suppressor of ST2 blocks allergic responses. Elife 9, e54017 (2020). · DOI
  14. Colomb, F. et al. IL-33-binding HpARI family homologues with divergent effects in suppressing or enhancing type 2 immune responses. Infect. Immun. 92, e0039523 (2024). · DOI
  15. Chauche, C. et al. A truncated form of HpARI stabilizes IL-33, amplifying responses to the cytokine. Front Immunol. 11, 1363 (2020). · DOI
  16. Curren, B. et al. IL-33-induced neutrophilic inflammation and NETosis underlie rhinovirus-triggered exacerbations of asthma. Mucosal Immunol. 16, 671–684 (2023). · DOI
  17. Meiners, J. et al. IL-33 facilitates rapid expulsion of the parasitic nematode strongyloides ratti from the intestine via ILC2- and IL-9-driven mast cell activation. PLoS Pathog. 16, e1009121 (2020). · DOI
  18. Block, K. E. et al. Physiological microbial exposure transiently inhibits mouse lung ILC2 responses to allergens. Nat. Immunol. 23, 1703–1713 (2022). · DOI
  19. Chen, W. Y., Tsai, T. H., Yang, J. L. & Li, L. C. Therapeutic strategies for targeting IL-33/ST2 signalling for the treatment of Inflammatory Diseases. Cell Physiol. Biochem. 49, 349–358 (2018). · DOI
  20. Kelsen, S. G. et al. Astegolimab (anti-ST2) efficacy and safety in adults with severe asthma: a randomized clinical trial. J. Allergy Clin. Immunol. 148, 790–798 (2021). · DOI
  21. Wechsler, M. E. et al. Efficacy and safety of itepekimab in patients with moderate-to-severe asthma. N. Engl. J. Med. 385, 1656–1668 (2021). · DOI
  22. England, E. et al. Tozorakimab (MEDI3506): an anti-IL-33 antibody that inhibits IL-33 signalling via ST2 and RAGE/EGFR to reduce inflammation and epithelial dysfunction. Sci. Rep. 13, 9825 (2023). · DOI
  23. Okragly, A. J. et al. Generation and characterization of torudokimab (LY3375880): a monoclonal antibody that neutralizes interleukin-33. J. Inflamm. Res. 14, 3823–3835 (2021). · DOI
  24. Holm, L. Dali server: structural unification of protein families. Nucleic Acids Res. 50, W210–W215 (2022). · DOI
  25. Liu, X. et al. Structural insights into the interaction of IL-33 with its receptors. Proc. Natl Acad. Sci. USA 110, 14918–14923 (2013). · DOI
  26. Strickson, S. et al. Oxidised IL-33 drives COPD epithelial pathogenesis via ST2-independent RAGE/EGFR signalling complex. Eur. Respir. J. 62, 2202210 (2023). · DOI
  27. Drurey, C. et al. Intestinal epithelial tuft cell induction is negated by a murine helminth and its secreted products. J. Exp. Med. 219, e20211140 (2022). · DOI
  28. de Oliveira, M. F. A., Talvani, A. & Rocha-Vieira, E. IL-33 in obesity: where do we go from here? Inflamm. Res. 68, 185–194 (2019). · DOI
  29. Winter, G. et al. DIALS: implementation and evaluation of a new integration package. Acta Crystallogr. D. Struct. Biol. 74, 85–97 (2018). · DOI
  30. McCoy, A. J. et al. Phaser crystallographic software. J. Appl Crystallogr 40, 658–674 (2007). · DOI · Paper page
  31. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr D. Biol. Crystallogr 66, 486–501 (2010). · DOI · Paper page
  32. Afonine, P. V. et al. Towards automated crystallographic structure refinement with phenix.refine. Acta Crystallogr D. Biol. Crystallogr 68, 352–367 (2012). · DOI · Paper page
  33. Bricogne G. et al BUSTER version 2.10.4. Cambridge, United Kingdom: Global Phasing Ltd. https://www.globalphasing.com/buster/wiki/index.cgi?BusterCite (2017).
  34. Adams, P. D. et al. PHENIX: a comprehensive python-based system for macromolecular structure solution. Acta Crystallogr D. Biol. Crystallogr. 66, 213–221 (2010). · DOI · Paper page
  35. Hopkins, J. B., Gillilan, R. E. & Skou, S. BioXTAS RAW: improvements to a free open-source program for small-angle X-ray scattering data reduction and analysis. J. Appl. Crystallogr. 50, 1545–1553 (2017). · DOI
  36. Panjkovich, A.S.D.I. CHROMIXS: automatic and interactive analysis of chromatography-coupled small angle X-ray scattering data. Bioinformatics 34, 1944–1946 (2017). · DOI
  37. Franke, D. et al. ATSAS 2.8: a comprehensive data analysis suite for small-angle scattering from macromolecular solutions. J. Appl. Crystallogr. 50, 1212–1225 (2017). · DOI
  38. Svergun, D. I. Determination of the regularization parameter in indirect-transform methods using perceptual criteria. J. Appl. Crystallogr. 25, 495–503 (1992). · DOI
  39. Franke, D. & Svergun, D. I. DAMMIF, a program for rapid ab-initio shape determination in small-angle scattering. J. Appl. Crystallogr. 42, 342–346 (2009). · DOI · Paper page
  40. Grawert, T. W. & Svergun, D. I. Structural modeling using solution small-angle x-ray scattering (SAXS). J. Mol. Biol. 432, 3078–3092 (2020). · DOI
  41. Svergun, D. I. Restoring low resolution structure of biological macromolecules from solution scattering using simulated annealing. Biophys. J. 76, 2879–2886 (1999). · DOI
  42. Wriggers, W. Conventions and workflows for using situs. Acta Crystallogr. D. Biol. Crystallogr 68, 344–351 (2012). · DOI
  43. Pettersen, E. F. et al. UCSF chimera-a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004). · DOI
  44. Putnam, D. K., Lowe, E. W. Jr. & Meiler, J. Reconstruction of SAXS profiles from protein structures. Comput Struct. Biotechnol. J. 8, e201308006 (2013). · DOI

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Metadata

Authors
Jamwal, A., Colomb, F., McSorley, H. J., & Higgins, M. K.
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2024-06-19
Volume
15
Issue
1
ISSN
2041-1723
DOI
10.1038/s41467-024-49550-0
PMC
PMC11189471
Cited by
15
References
5
Language
en
Licence
Funders
Wellcome Trust (221914/Z/20/Z)

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