ATP-binding cassette (ABC) proteins transport a wide variety of substrates, including sugars, amino acids, metal ions, lipids, peptides and proteins, across membranes, and most ABC proteins contain transmembrane domains (ABC transporters). Sequencing of the Schizosaccharomyces pombe genome has allowed identification of all genes encoding ABC transporters in fission yeast. To date, six such genes have been characterized, and an additional five genes encoding ABC transporters were identified from the genome sequence. In an attempt to characterize all of the ABC transporters in fission yeast, all 11 genes were disrupted. While all the genes were found to be dispensable for cell viability, some disruptants lacked apparent phenotypes. GFP-tagged ABC transporters were localized to membranes as follows: plasma membrane (2), vacuolar membrane (4), mitochondrial membrane (2), endoplasmic reticulum membrane (2), and endosome and Golgi membranes (1). Two Cluster II. 1 proteins, Abc2p (SPAC3F10.11c) and Abc4p (SPAC30.04c), were found to be localized to vacuolar membranes, and to be responsible for accumulation of a characteristic red pigment in the vacuole of an adenine biosynthetic mutant. The doubly disrupted mutant abc2 Delta abc4 Delta exhibited drug sensitivity, and a decreased accumulation of monochlorobimane, suggesting that both of the proteins encoded by these genes are involved in detoxification of xenobiotics, and vacuolar sequestration of glutathione S-conjugates.