This study examined the relationship between alterations in cellular metabolism and induction of proliferation in renal proximal tubular cells (RPTC) after epidermal growth factor (EGF) exposure. EGF treatment (10 ng/ml) of confluent RPTC cultures for 6 consecutive days increased monolayer DNA content 3.3-fold. EGF-stimulated proliferation of RPTC was preceded by a rapid (within 4 h) induction of glycolysis and a decrease in basal and ouabain-sensitive oxygen consumption (20 and 30%, respectively). EGF stimulated the pentose cycle by 58% and decreased gluconeogenesis by 48%. Supplementation of the culture medium with ribose-5-phosphate or ribose abolished the stimulation of glycolysis and the pentose cycle by EGF but had no effect on proliferation. These results show that EGF rapidly stimulates the pentose cycle, shifts glucose metabolism from gluconeogenesis to glycolysis, and decreases oxygen consumption before any increase in proliferation. The lack of an EGF effect on the pentose cycle and glycolysis in the presence of exogenous precursors for DNA synthesis suggests that the stimulation of these pathways before proliferation is due to increased demands for ribose for subsequent nucleic acid synthesis.