THIS IS A FASE-EPSCOR STRENGTHENING STANDARD AWARD THAT WORKS ON PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS (PRRSV). PRRSV IS THE ETIOLOGIC AGENT OF PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME (PRRS), AN ECONOMICALLY DEVASTATING, PANDEMIC DISEASE OF SWINE THAT IS TYPICALLY CHARACTERIZED BY REPRODUCTIVE FAILURE IN BREEDING HERDS AND RESPIRATORY PROBLEMS AND GROWTH RETARDATION IN GROWING PIGS. TWO PRRS OUTBREAKS WERE FIRST REPORTED IN THE LATE 1980S IN NORTH AMERICA AND CENTRAL EUROPE. THE DISEASE IS NOW FOUND IN MOST PIG-PRODUCING COUNTRIES AND AFFECTS THE SWINE INDUSTRY AND FOOD SECURITY WORLDWIDE, CAUSING ENORMOUS ECONOMIC LOSSES EACH YEAR. IN THE US, THE ANNUAL LOSS DUE TO PRRS IS ESTIMATED TO EXCEED $500 MILLION. IN PARTICULAR, THE EMERGENCE OF HIGHLY PATHOGENIC PRRSVS IN CHINA AND VIETNAM IN 2006 AND THEIR RAPID SPREAD TO SEVERAL NEIGHBORING ASIAN COUNTRIES HAVE RAISED A GROWING CONCERN THAT NEW PATHOGENIC PRRSVS CAN SPREAD THROUGHOUT THE WORLD, POSING A SIGNIFICANT THREAT TO THE GLOBAL AGRICULTURAL COMMUNITY, INCLUDING BOTH PRODUCERS AND CONSUMERS.THE GOAL OF THIS PROJECT IS TO UNDERSTAND HOW PRRSV GAINS ACCESS TO THE INTERIOR OF SUSCEPTIBLE CELLS, THE FIRST STEP IN AN INFECTION PROCESS THAT INVOLVES A CASCADE OF MULTIPLE, HIGHLY COORDINATED INTERACTIONS BETWEEN THE VIRUS AND ITS TARGET CELLS. THE WORK PROPOSED HERE INVOLVES THE USE OF TWO COMPLEMENTARY, TECHNOLOGICALLY ADVANCED GENOME-SCALE GENETIC SCREENS FOR GAIN- AND LOSS-OF-FUNCTION OF PRRSV ENTRY TO DISCOVER THE CELLULAR FACTORS IN THE PIG HOST THAT ARE CRITICAL FOR VIRAL ENTRY AND TO DISSECT THE DISCRETE ENTRY STEPS THAT ARE REGULATED BY SPECIFIC HOST FACTORS. TWO INDEPENDENT BUT COMPLEMENTARY AIMS WILL BE CARRIED OUT: IN AIM 1, AN ITERATIVE CDNA LIBRARY SCREENING APPROACH WILL BE USED TO IDENTIFY SPECIFIC CELLULAR GENES IN A HIGHLY PRRSV-SUSCEPTIBLE PORCINE MACROPHAGE CELL LINE THAT CONFER SUSCEPTIBILITY TO PRRSV INFECTION ON A PRRSV-NONSUSCEPTIBLE PORCINE KIDNEY CELL LINE. IN AIM 2, A MULTIPLEXED RNAI SCREENING APPROACH WILL BE EMPLOYED TO IDENTIFY CELLULAR GENES THAT PLAY AN IMPORTANT ROLE IN PRRSV ENTRY INTO A HIGHLY PRRSV-SUSCEPTIBLE PORCINE MACROPHAGE CELL LINE. THE OUTCOMES OF THIS PROJECT WILL (1) PROVIDE A UNIQUE OPPORTUNITY TO GAIN A COMPLETE UNDERSTANDING OF HOW PRRSV-HOST CELL INTERACTIONS OCCUR AT THE LEVEL OF PRRSV ENTRY, (2) SHED NEW LIGHT ON THE CELL/TISSUE TROPISM AND PATHOGENESIS OF PRRSV, AND (3) PROVIDE NEW TARGETS FOR THE DEVELOPMENT OF NOVEL ANTIVIRAL INTERVENTIONS CAPABLE OF INHIBITING THE EARLY STEPS OF PRRSV INFECTION.