Serine proteases are well known as enzymes involved in digestion of dietary proteins, blood coagulation, and homeostasis. Only recent groundbreaking studies revealed a novel role of serine proteases as signaling molecules acting via protease-activated receptors (PARs). Important effects of PAR activation on leukocyte motility, cytokine production, adhesion molecule expression, and a variety of other physiological or pathophysiological functions have been described in vitro and in vivo. The crucial role of PAR activation during disease progression was revealed in animal models of different gastrointestinal pathologies, neuroinflammatory and neurodegenerative processes, skin, joint and airway inflammation, or allergic responses. This review focuses on the findings related to the impact of PAR deficiency in animal models of inflammatory and allergic diseases. Additionally, we observe the role of PAR activation in the regulation of functional responses of innate and adaptive immune cells in vitro. Understanding the mechanisms by which PARs exert the effects of serine proteases on immune cells may lead to new therapeutic strategies in inflammation, immune defense, and allergy.