Phosphomannose isomerase (PMI40) catalyzes the conversion between fructose 6-phosphate and mannose 6-phosphate and thus connects glycolysis, i.e. energy production and GDP-mannose biosynthesis or cell wall synthesis in Saccharomyces cerevisiae. After PMI40 deletion (pmi ؊ ) the cells were viable only if fed with extracellular mannose and glucose. In an attempt to force the GDP-mannose synthesis in the pmi ؊ strain by increasing the extracellular mannose concentrations, the cells showed significantly reduced growth rates without any alterations in the intracellular GDPmannose levels. To reveal the mechanisms resulting in reduced growth rates, we measured genome-wide gene expression levels, several metabolite concentrations, and selected in vitro enzyme activities in central metabolic pathways. The increasing of the initial mannose concentration led to an increase in the mannose 6-phosphate concentration, which inhibited the activity of the second enzyme in glycolysis, i.e. phosphoglucose isomerase converting glucose 6-phosphate to fructose 6-phosphate. As a result of this limitation, the flux through glycolysis was decreased as was the median expression of the genes involved in glycolysis. The expression levels of RAP1, a transcription factor involved in the regulation of the mRNA levels of several enzymes in glycolysis, as well as those of cell cycle regulators CDC28 and CLN3, decreased concomitantly with the growth rates and expression of many genes encoding for enzymes in glycolysis.Phosphomannose isomerase enzyme (PMIe) 1 catalyzes the interconversion of fructose 6-phosphate (Fru-6-P) in glycolysis to mannose 6-phosphate (Man-6-P) through a mannose pathway. In the eukaryotic model organism, yeast Saccharomyces cerevisiae, phosphomannose isomerase is encoded by the PMI40 gene (1). In a PMI40 deletion strain (pmi Ϫ ), the synthesis of Man-6-P from Fru-6-P is not possible, disabling the growth of such a strain on medium without mannose. The inability to grow, caused by defective glycosylation of a temperature-sensitive pmi40 mutant of S. cerevisiae, and repairing the defects by addition of mannose to the growth medium have been described previously (2). In humans PMIe deficiency is the cause of carbohydrate-deficient glycoprotein syndrome type Ib, but the condition can be successfully treated by mannose administration (3). Man-6-P produced either from Fru-6-P or mannose serves as a precursor for the de novo biosynthesis of GDP-mannose. Man-6-P is converted to mannose 1-phosphate (Man-1-P) by phosphomannomutase encoded by SEC53. Subsequently, Man-1-P is ligated with the guanosine 5-triphosphate molecule (GTP) to form GDP-mannose by Man-1-P guanylyltransferase encoded by PSA1 (4). The de novo formation of the purine ring of GTP, required for the biosynthesis of GDPmannose, starts from ribose 5-phosphate in the pentose phosphate pathway and requires also 3-phosphoglycerate in the glycolysis as a precursor. Taken together, the biosynthesis of GTP is more complex than the mannose pathway (4). GDPmannose is needed in ...