We have investigated the thiamine metabolism in Schizosaccharomyces pombe and shown that: (1) Thiamine-repressible acid phosphate, coded for by the gene pho4, dephosphorylates thiamine phosphates indicating that the enzyme acts as a thiamine phosphate phosphatase. (2) In vivo synthesized thiamine is present intracellularly mainly as thiamine diphosphate. Starving cells for glucose decreases the intracellular thiamine pool. (3) The genes thi2, thi3 and thi4 control thiamine biosynthesis and probably code for thiamine biosynthetic enzymes. Thi3, which is involved in the synthesis of the pyrimidine moiety of the thiamine molecule, is allelic to the thiamine repressible gene nmt1. (4) Thiamine uptake is a thiamine regulated process, probably occurs by active transport and is controlled by the gene ptr1.