Transcription factor BZR2 activates chitinase Cht20.2 transcription to confer resistance to wheat stripe rust.

The brassinosteroid pathway promotes a variety of physiological processes in plants and the brassinosteroid insensitive1-ethylmethane sulfonate suppressor (BES)/brassinazole-resistant (BZR) functions as one of its key regulators. We previously showed that the BES/BZR-type transcription factor TaBZR2 mediates the drought stress response in wheat (Triticum aestivum) by directly upregulating the transcriptional activity of glutathione S-transferase 1. However, the function of TaBZR2 in plants under biotic stresses is unknown. In this study, we found that transcript levels of TaBZR2 were upregulated in response to inoculation with wheat stripe rust fungus (Puccinia striiformis f. sp. tritici, Pst) and treatment with flg22 or an elicitor-like protein of Pst, Pst322. Wheat lines overexpressing TaBZR2 conferred increased resistance, whereas TaBZR2-RNAi lines exhibited decreased resistance to multiple races of Pst. TaBZR2 targeted the promoter of the chitinase gene TaCht20.2, activating its transcription. Knockdown of TaCht20.2 in wheat resulted in enhanced susceptibility to Pst, indicating the positive role of TaCht20.2 in wheat resistance. Upon Pst infection in vivo, the overexpression of TaBZR2 increased total chitinase activity, whereas RNAi-mediated silencing of TaBZR2 reduced total chitinase activity. Taken together, our results suggest that TaBZR2 confers broad-spectrum resistance to the stripe rust fungus by increasing total chitinase activity in wheat.

Chitinase-Induced Airway Hyperreactivity and Inflammation in a Mouse Model of Nonallergic Asthma.

INTRODUCTION: Environmental exposure to mites and fungi has been proposed to critically contribute to the development of IgE-mediated asthma. A common denominator of such organisms is chitin. Human chitinases have been reported to be upregulated by interleukin-13 secreted in the context of Th2-type immune responses and to induce asthma. We assessed whether chitin-containing components induced chitinases in an innate immune-dependent way and whether this results in bronchial hyperresponsiveness. MATERIALS AND METHODS: Monocyte/macrophage cell lines were stimulated with chitin-containing or bacterial components in vitro. Chitinase activity in the supernatant and the expression of the chitotriosidase gene were measured by enzyme assay and quantitative PCR, respectively. Non-sensitized mice were stimulated with chitin-containing components intranasally, and a chitinase inhibitor was administered intraperitoneally. As markers for inflammation leukocytes were counted in the bronchoalveolar lavage (BAL) fluid, and airway hyperresponsiveness was assessed via methacholine challenge. RESULTS: We found both whole chitin-containing dust mites as well as the fungal cell wall component zymosan A but not endotoxin-induced chitinase activity and chitotriosidase gene expression in vitro. The intranasal application of zymosan A into mice led to the induction of chitinase activity in the BAL fluid and to bronchial hyperresponsiveness, which could be reduced by applying the chitinase inhibitor allosamidin. DISCUSSION: We propose that environmental exposure to mites and fungi leads to the induction of chitinase, which in turn favors the development of bronchial hyperreactivity in an IgE-independent manner.

In vitro assessment of allergenicity features and localization of probable IgE binding regions.

Rice is cultivated as a staple grain crop in many countries, especially in Asia. In the present study, recombinant rice chitinase was expressed, purified and characterized by in silico and immunobiochemical methods. Rice chitinase was affinity purified and it resolved at 24 kDa on SDS-PAGE. Purified protein was analyzed for pepsin resistance, heat stability, and IgE binding using atopic patients' sera. Chitinase was resistant to pepsin digestion and heat treatment at 90 °C for 1 h. It showed significant IgE binding with 7 of 110 patients' sera positive to different food allergens. Homology modeled 3D structure of rice chitinase was used for B cell epitope prediction. In silico predicted B cell peptides were assessed for IgE binding by ELISA using food allergic patients' sera, epitope RC2 showed IgE binding comparable to chitinase. In conclusion, chitinase was identified as a potential allergen and may share cross reactive epitopes with food allergens.

Class I chitinases as potential panallergens involved in the latex-fruit syndrome.

BACKGROUND: Latex-fruit cross-sensitization has been fully demonstrated. However, the antigens responsible for this "latex-fruit syndrome" have not been identified. We have recently shown that class I chitinases are relevant chestnut and avocado allergens. OBJECTIVE: We sought to evaluate the in vivo and in vitro reactions of purified chestnut and avocado chitinases in relation to the latex-fruit syndrome. METHODS: From a latex-allergic population, eighteen patients allergic to chestnut, avocado, or both were selected. Skin prick tests (SPTs) were performed with crude chestnut and avocado extracts, chitinase-enriched preparations, and purified class I and II chitinases from both fruits. CAP-inhibition assays with the crude extracts and purified proteins were carried out. Immunodetection with sera from patients with latex-fruit allergy and immunoblot inhibition tests with a latex extract were also performed. Eighteen subjects paired with our patients and 15 patients allergic to latex but not food were used as control groups. RESULTS: The chestnut class I chitinase elicited positive SPT responses in 13 of 18 patients with latex-fruit allergy (72%), and the avocado class I chitinase elicited positive responses in 12 of 18 (67%) similarly allergic patients. By contrast, class II enzymes without a hevein-like domain did not show SPT responses in the same patient group. Each isolated class I chitinase reached inhibition values higher than 85% in CAP inhibition assays against the corresponding food extract in solid phase. Immunodetection of the crude extracts and the purified class I chitinases revealed a single 32-kd band for both chestnut and avocado. Preincubation with a natural latex extract fully inhibited the IgE binding to the crude extracts, as well as to the purified chestnut and avocado class I chitinases. CONCLUSION: Chestnut and avocado class I chitinases with an N-terminal hevein-like domain are major allergens that cross-react with latex. Therefore they are probably the panallergens responsible for the latex-fruit syndrome.