Aquaculture is the fastest growing food producing sector in the world, and catfish represent the major fresh water farmed species in the US. Because of the economic importance of catfish to the U.S. aquaculture industry, it has been a long-term focus of this laboratory to functionally characterize the catfish immune system. In humans, white blood cells play important roles in fighting infection. However, very little is known about these cells in fish. The goal of the current study is to better understand how white blood cells in the catfish recognize and kill bacteria and virus-infected cells. Recent studies have shown that a family of proteins termed Leukocyte Immune Type Receptors (LITRs) is expressed on the surface of catfish white blood cells. These LITR expressing cells are capable of killing virus infected cells. This project will determine how the expression patterns of different LITR genes change in immune organs and white blood cells during the course of an infection and also provide information on the types of proteins or molecular structures LITRs recognize. This study will also provide unique research training opportunities for graduate, undergraduate and high school students, including students from groups under-represented in science. The students will learn basic immunology and gain expertise in a variety of molecular and cellular techniques. Graduate and undergraduate students will have their own research projects, learn how to design experiments, interpret data and present their findings at regional or national meetings.
Studies of teleost immune responses are important because they provide information about the various immune strategies adapted by different animal groups. This project focuses on the LITR immunoregulatory receptor family that is found in channel catfish and other bony fish. LITRs comprise a highly diverse multigene family and many of the LITRs that have been sequenced are structurally related to mammalian NK receptors encoded within the leukocyte receptor complex. Additionally, certain LITRs, as defined by their reactivity with monoclonal antibody CC41, are expressed on catfish cytotoxic cells that recognize and kill virus infected cells via the secretory-perforin granzyme pathway. This project will first define the complete LITR gene repertoire in catfish and examine LITR expression after viral and bacterial challenge using PacBio single molecule long-read sequencing protocols. Second, the ligands for selected CC41-reactive LITRs will be determined using recombinant fusion proteins either in the form of tetramers or immunoglobulin-fusion proteins. The LITR-ligand complexes will be isolated and immunoselected proteins will be analyzed by LS-MS/MS. Third, since preliminary sequencing has identified a small number of LITRs that resemble mammalian Fc-receptors, representatives from this group will be analyzed for immunoglobulin binding. The successful completion of this project will define the functional properties of LITRs and provide insights into the evolution of innate immune regulatory receptors. Moreover, this project will provide novel information about how ectothermic vertebrates maintain self-tolerance and recognize altered-self.
