Cholesterol homeostasis is maintained by coordinate regulation of endogenous synthesis and exogenous uptake of lipoprotein cholesterol by low density lipoprotein (LDL) receptors. In the lymphocyte, limiting the availability of exogenous cholesterol is known to increase the rate of endogenous sterol biosynthesis. However, the effect of cholesterol deprivation on the expression and regulation of the LDL receptor gene has not been delineated in lymphocytes. Here, LDL receptor mRNA was detected in freshly isolated human peripheral mononuclear cells. LDL receptor mRNA levels increased by 3-fold during a one-h in vitro culture in lipoprotein-deficient medium and by 6-fold during a 2-h incubation. Actinomycin D blocked the synthesis of LDL receptor mRNA in these cultures. However, neither cycloheximide nor LDL or oxygenated sterols suppressed the increase in LDL receptor mRNA levels observed after a 2-h incubation. The increase in LDL receptor mRNA was maintained for 24 h of culture in the absence of LDL. Ongoing gene transcription and not mRNA stabilization accounted for this expression. Inhibition of protein synthesis with cycloheximide completely prevented the sustained increase in LDL receptor mRNA levels measured after 24 h. Low concentrations of LDL (5 micrograms of cholesterol/ml) and oxygenated sterols also suppressed the level of LDL receptor mRNA measured after a 24-h incubation. These data show that the initial upregulation of LDL receptor gene expression is independent of protein synthesis and not suppressed by either LDL or oxygenated sterols. In contrast, the continued transcription necessary for the maintenance of steady-state levels of LDL receptor mRNA requires synthesis of new protein and is regulated by LDL and oxygenated sterols.