Understanding how animals and plants defend themselves from infection is critically important to basic biology, agriculture, and medicine. Central to defense against infection is the immune system. In insects and other arthropods, a key component of the immune system is a pathway called the phenoloxidase cascade, which consists of a complex network of proteins that includes the enzyme phenoloxidase. Phenoloxidase in turn catalyzes the formation of toxic immune defense molecules and a type of pigment called melanin. Many studies make clear that the phenoloxidase cascade is extremely important in immune defense but how this biochemical pathway is regulated is only partially understood. This proposal addresses the question of what factors must interact with phenoloxidase to produce an enzyme that can convert the starting substrate for this reaction into melanin? To address this question, we will conduct a series of detailed biochemical studies that will identify the proteins that interact with phenoloxidase, and functional studies that will identify the components required to produce a functional enzyme complex. The molecules produced by phenoloxidase are critical to many biological processes in addition to immunity. These include wounding responses in plants, pigmentation of animals, and the onset of diseases like Parkinson's and melanotic cancers. Results of this study will thus contribute information of importance that extends beyond enhancing our understanding of immunity. The proposed research also provides an ideal platform for promoting scientific training and education. The project will provide independent research opportunities for postdoctoral, graduate and undergraduate trainees in immunology and biochemistry. Results from the study will also be used to enhance classroom instruction at the high school, undergraduate, and graduate levels.