Interferon regulatory factor-1 (IRF-1) is a transcriptional activator that is involved in interferon response, regulation of cell growth and oncogenesis. To try to identify the molecules that regulate the function of IRF-1, we performed yeast two-hybrid screening and isolated protein inhibitor of activated STAT3 (PIAS3) as an IRF-1-binding protein. This protein was also found to bind with small ubiquitin-related modifier-1 (SUMO-1) and ubiquitin-conjugating enzyme 9, an E2 in the SUMO-1-conjugating system. Co-expression of PIAS3 induced SUMO-1 modification of IRF-1 in a RING finger domain-dependent manner and also repressed transcriptional activity of IRF-1. Thus, PIAS3 functions as a SUMO-1 ligase for IRF-1 and also as a repressor of IRF-1 transcriptional activity.