A Trypsin‐like Protease from Alternaria alternata Allergens Promotes Airway Inflammation through Activation of Protease‐activated Receptor‐2/β‐arrestin Signaling

Abstract

Fungal Alternaria alternata exposure is correlated with increased morbidity and a higher risk of fatal asthma attacks in asthmatic patients. Alternaria allergens contain serine proteases capable of activating proteinase‐activated receptor‐2 (PAR2) and promote airway inflammation that is dependent upon protease activity. We have previously shown that PAR2‐dependent airway inflammation is mediated through the β‐arrestin‐2 activated pathway. Thus, we hypothesized that proteases from Alternaria filtrates will promote activation of the β‐arrestin‐2 (βarr2) dependent cellular signaling pathways that mediate airway inflammation in murine models of Alternaria‐induced asthma. We identify a single alkaline serine protease, with trypsin‐like characteristics, AASP (Alternaria Alkaline Serine Protease) that can activate PAR2, as demonstrated by recruitment of β‐arrestin‐2 to PAR2 and mobilization of intracellular Ca2+ in cultured cells. Using histological and flow cytometric analyses, we demonstrate that Alternaria‐induced airway inflammation requires both PAR2 and βarr2, as demonstrated by the reduced recruitment of leukocytes (particularly eosinophils and CD4+ T‐cells) into the lung, goblet cell hyperplasia, and thickening of the lung epithelium in PAR2−/−or βarr2−/−, compared to wild‐type mice. These inflammation parameters are also reduced by treatment of mice with Soybean Trypsin Inhibitor (SBTI) during the administration of Alternaria allergens, indicating that AASP is likely necessary for the inflammation induced by Alternaria. These are the first experiments demonstrating that the PAR2‐β‐arrestin‐dependent signaling axis is important for asthma induced by household allergens. Thus, targeting this pathway via biased antagonism of PAR2 could be a new avenue of therapeutic intervention for asthma.