Five lipase genes have been identified and sequenced from Candida rugosa. However, as the sequences of LIP multigene family are extremely closely related, it is difficult to characterize the expression spectrum of LIP genes. In the present work we have cloned, sequenced, and analyzed the promoters of these five LIP isoform genes, and several putative transcriptional elements including oleate response element (ORE) and upstream activation sequence 1 (UAS1) were identified. A quantitative real-time RT-PCR method was developed for determining the differential expression of C. rugosa lipase family genes in response to various environmental and nutritional factors. While all five LIP genes display significant changes in mRNA expression under oleic acid and/or olive oil culture conditions, LIP2 showed the strongest induction (456-fold) in response to oleic acid. LIP transcription and promoter regulation were studied by assaying the beta-galactosidase activities of promoter-lacZ fusions in Saccharomyces cerevisiae. Three of the LIP genes, LIP3, LIP4, and LIP5, showed significant induction by oleic acid, and their ORE and UAS1 elements are essential for induction by oleic acid. Together, this suggests that the multiple lipase expression profiles may be due to differential transcriptional regulation of the LIP genes in response to environment or nutritional factors.