Allergenicity resulting from functional mimicry of a Toll-like receptor complex protein - PubMed (original) (raw)

Allergenicity resulting from functional mimicry of a Toll-like receptor complex protein

Aurelien Trompette et al. Nature. 2009.

Abstract

Aeroallergy results from maladaptive immune responses to ubiquitous, otherwise innocuous environmental proteins. Although the proteins targeted by aeroallergic responses represent a tiny fraction of the airborne proteins humans are exposed to, allergenicity is a quite public phenomenon-the same proteins typically behave as aeroallergens across the human population. Why particular proteins tend to act as allergens in susceptible hosts is a fundamental mechanistic question that remains largely unanswered. The main house-dust-mite allergen, Der p 2, has structural homology with MD-2 (also known as LY96), the lipopolysaccharide (LPS)-binding component of the Toll-like receptor (TLR) 4 signalling complex. Here we show that Der p 2 also has functional homology, facilitating signalling through direct interactions with the TLR4 complex, and reconstituting LPS-driven TLR4 signalling in the absence of MD-2. Mirroring this, airway sensitization and challenge with Der p 2 led to experimental allergic asthma in wild type and MD-2-deficient, but not TLR4-deficient, mice. Our results indicate that Der p 2 tends to be targeted by adaptive immune responses because of its auto-adjuvant properties. The fact that other members of the MD-2-like lipid-binding family are allergens, and that most defined major allergens are thought to be lipid-binding proteins, suggests that intrinsic adjuvant activity by such proteins and their accompanying lipid cargo may have some generality as a mechanism underlying the phenomenon of allergenicity.

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Figures

Figure 1

Figure 1. Der p 2 reconstitutes and amplifies TLR4 signalling in the absence and presence of MD-2, respectively

a, HEK293 cells expressing CD14 and TLR4 (HEK293/CD14/TLR4) were transfected with Der p 2 (black bars) or empty vector (EV, white bars) and stimulated (or mock stimulated) with LPS (10 ng/ml). b, HEK293/CD14/TLR4 cells transfected with MD-2 along with Der p 2 (black bars) or EV (white bars) were stimulated (or mock stimulated) with LPS or IL-1β (100 ng/ml). c and d, HEK293/CD14/TLR4 cells transfected with Der p 2 (black bars), Der p 2-Y91A (grey bars) or EV (white bars), in the absence (c) or presence (d) of MD-2, were stimulated (or mock stimulated) with LPS. Data represent means ± S.E. of cultures (stimulated - mock-stimulated) in a single experiment, representative of an experimental n of 3–8. *P < 0.05; ‡P < 0.005; §P < 0.001; unpaired _t_-test.

Figure 2

Figure 2. Der p 2 interacts directly with the TLR4 complex and with LPS

a, Co-immunoprecipitation (co-IP) of Der p 2 with TLR4 in lysates of HEK293FT cells transfected with the indicated constructs or EV controls (−). b, Co-IP of Der p 2 with MD-2. c, Co-IP of Der p 2 with CD14. d, Effect of Der p 2 on co-IP of MD-2 with TLR4. e, Supernatants from HEK293FT cells transfected with HA-tagged Der p 2 (or EV) underwent incubation with biotinylated LPS, precipitation (P) with streptavidin beads, and immunoblotting (IB) with HA Ab. Similar results were seen with immunoaffinity-purified Der p 2 and Der p 2 mAb. Data are representative of an experimental n = 2–8.

Figure 3

Figure 3. TLR4-dependent induction of experimental allergic asthma by Der p 2

Wild type and TLR4-deficient mice were sensitized intranasally on d 0, 1 and 2 with rDer p 2 (0.1 µg)/LPS (0.026 pg) in PBS, or PBS/LPS (0.026 pg), challenged intranasally with 1/4 of the sensitization dose on d 14, 15, 18 and 19, and analyzed on d 21. a, b Inflammatory cells in bronchoalveolar lavage (BAL) fluids. a, Total cell numbers (white bars, PBS/LPS; black bars, Der p 2/LPS). b, Differential cell counts. c, Total serum IgE (bars as in a). d, Representative lung sections stained with periodic acid-Schiff. Data (a–c) represent means ± S.E. of 5–8 animals/group, and are representative of 2 independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001; 2-way ANOVA on log-transformed data.

Figure 4

Figure 4. Der p 2 induces experimental allergic asthma in the absence of MD-2

Wild type and MD-2-deficient mice were sensitized, challenged and analyzed as in Figure 3, except for the use of 10-fold higher doses of Der p 2, given the less robust stimulation of TLR4 by Der p 2 observed in the absence of MD-2. a, b Inflammatory cells in BAL fluids. a, Total cell numbers (white bars, PBS/LPS; black bars, Der p 2/LPS). b, Differential cell counts. c, Total serum IgE (bars as in a). d, Representative lung sections stained with periodic acid-Schiff. Data (a–c) represent means ± S.E. of 5–9 animals/group. *P < 0.05, **P < 0.01, ***P < 0.001; 2-way ANOVA on log-transformed data.

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