The reducing polyketide synthases found in filamentous fungi are involved in the biosynthesis of many drugs and toxins. Lichens produce bioactive polyketides, but the roles of reducing polyketide synthases in lichens remain to be clearly elucidated. In this study, a reducing polyketide synthase gene (UlPKS3) was isolated and characterized from a cultured mycobiont of Usnea longissima. Complete sequence information regarding UlPKS3 (6,519 bp) was obtained by screening a fosmid genomic library. A UlPKS3 sequence analysis suggested that it contains features of a reducing fungal type I polyketide synthase with β-ketoacyl synthase (KS), acyltransferase (AT), dehydratase (DH), enoyl reductase (ER), ketoacyl reducatse (KR), and acyl carrier protein (ACP) domains. This domain structure was similar to the structure of ccRads1, which is known to be involved in resorcylic acid lactone biosynthesis in Chaetomium chiversii. The results of phylogenetic analysis located UlPKS3 in the clade of reducing polyketide synthases. RT-PCR analysis results demonstrated that UlPKS3 had six intervening introns and that UlPKS3 expression was upregulated by glucose, sorbitol, inositol, and mannitol.