Nutrition modulates both production and composition of milk. Milk composition was studied in rats chronically fed a diet without additional lipids, and therefore eating only traces of the recommended supply of essential polyunsaturated fatty acid. Despite a large decrease in milk-protein synthesis, only protein composition, but not protein concentration, was found to change in the milk of rats following a lipid-deprived diet. Correlatively, we observed a substantial increase in the lactose concentration of milk. Analysis of milk proteins by two-dimensional electrophoresis demonstrated that the relative proportion of the various molecular forms of k-casein, an O-glycosylated protein, was modified in the milk of rats receiving the lipiddeprived diet. In tissues, differences in the two-dimensional pattern of k-casein between control and lipid-deprived rats were similar, if not identical. In contrast to k-casein, the molecular forms of a-lactalbumin, an N-glycosylated protein, were not affected by the diet. These data provide evidence that O-glycosylation of milk proteins in the secretory pathway of mammary epithelial cells is modulated by the lipid content of experimental diets.