Intracellular distribution of haem after uptake by different receptors. Haem-haemopexin and haem-asialo-haemopexin.

How the interaction of haemopexin with two different receptors affects its subsequent metabolism and 'intracellular' haem transport was examined by using mesohaem-haemopexin and mesohaem-asialo-haemopexin. The physical properties of the two haem proteins, including their absorption and c.d. spectra, are similar. Binding studies in vitro showed that haem-asialo-haemopexin interacts with both the haemopexin-specific and galactose-specific receptors on liver plasma membranes, but that haem-haemopexin interacts only with the haemopexin receptor. In vivo haem-asialo-haemopexin rapidly interacts with the liver via the galactose-specific receptor, since the protein is extensively catabolized and uptake is blocked by asialofetuin. Haem iron from haem-asialo-haemopexin is not accumulated in the liver to the same extent as from intact haem-haemopexin, and the native sialylated protein is not proteolysed. Moreover, after fractionation of homogenized liver by using colloidal-silica gradients, liver-associated haem-haemopexin and haem-asialo-haemopexin produced distinctly different patterns for both protein and ligand, consistent with their uptake by two distinct receptors. These results demonstrate that the interaction of haemopexin with different receptors influences its subsequent metabolic fate and that haem iron from haem-haemopexin is efficiently conserved only if it enters the liver cell via the specific haemopexin receptor.     

Emerging strategies in microbial haem capture

Gram-negative pathogenic bacteria have evolved novel strategies to obtain iron from host haem-sequestering proteins. These include the production of specific outer membrane receptors that bind directly to host haem-sequestering proteins, secreted haem-binding proteins (haemophores) that bind haem/haemoglobin/haemopexin and deliver the complex to a bacterial cell surface receptor and bacterial proteases that degrade haem-sequestering proteins. Once removed from haem-sequestering proteins, haem may be transported via the bacterial outer membrane receptor into the cell. Recent studies have begun to define the steps by which haem is removed from bacterial haem proteins and transported into the cell. This review describes recent work on the discovery and characterization of these systems. Reference is also made to the transport of haem in serum (via haemoglobin, haemoglobin/haptoglobin, haemopexin, albumin and lipoproteins) and to mechanisms of iron removal from the haem itself (probably via a haem oxygenase pathway in which the protoporphyrin ring is degraded). Haem protein-receptor interactions are discussed in terms of the criteria that govern protein-protein interactions in general, and connections between haem transport and the emerging field of metal transport via metallochaperones are outlined.     

The effect of polycyclic aromatic hydrocarbons on choline kinase activity in mouse hepatoma cells

Choline kinase catalyzes the first rate-limiting step in the pathway of biosynthesis of phosphatidylcholine. This enzyme was shown previously to be induced in liver by treatment of rats with polycyclic aromatic hydrocarbons (Ishidate et al. (1980) Biochem. Biophys. Res. Commun. 96, 946-952). The present study was undertaken to determine whether choline kinase in the murine hepatoma cell line, Hepa 1c1c7, is inducible by aromatic hydrocarbons and, if so, whether this induction is mediated by the aromatic hydrocarbon receptor. Treatment of Hepa 1c1c7 cells with 10 microM beta-naphthoflavone resulted in a 1.6-fold increase of choline kinase activity, but no response was seen when the cells were exposed to either 5.0 microM benzo[a]pyrene or 1.0 nM 2.3,7,8-tetrachlorodibenzo-p-doxin, both potent inducers of aryl hydrocarbon hydroxylase. Cell line variants with either deficient or elevated aromatic hydrocarbon receptors showed no increase in choline kinase activity following treatment with any of the polycyclic aromatic hydrocarbons. These results are not consistent with a role for the aromatic hydrocarbon receptor in increased choline kinase activity in Hepa 1c1c7 cells.     

Haem transport to the liver by haemopexin. Receptor-mediated uptake with recycling of the protein

Rat [(59)Fe]haem-(125)I-labelled haemopexin complexes (700pmol/rat) associate rapidly and exclusively with the liver after intravenous injection into anaesthetized rats. The two isotopes exhibit different patterns of accumulation. Liver (125)I-labelled haemopexin is maximum 10min after injection (20+/-4.9pmol/g of liver) and then declines by 2h to the low values (about 3pmol/g of liver) seen after injection of the apoprotein. In contrast, [(59)Fe]haem accumulates in the liver for at least 2h. Haemopexin undergoes no extensive proteolysis during 2h of haem transport as shown by precipitation with acid (98%) and specific antiserum (92%) and by electrophoresis. Moreover, only 1-2% of the dose is located in extrahepatic tissues, and there is no significant urinary excretion of either (125)I or (59)Fe. Hepatic uptake at 10min is saturable, reaching 200pmol of haemopexin/g of liver and 350pmol of haem/g of liver at a dose of 9nmol/rat, whereas uptake of the apoprotein is 3-5% of the dose. This suggests that the interaction of haem-haemopexin with the liver is a specific receptor-mediated process. The complex probably interacts via the protein moiety, since the haem analogues mesohaem and deuterohaem do not affect association of the protein with the liver but the species of haemopexin does. Increasing amounts of protein are associated with the liver 5min after injection in the order: human>rabbit>rat, and haem uptake is consistently increased. For both rat and rabbit haemopexin saturation is reached at the same concentration of protein, i.e. 180-200pmol/g of liver, indicating that the different protein species bind to a common receptor. We propose that haemopexin transports haem to the liver by a specific receptor-mediated process and then returns to the circulation.     

Haem utilization in Vibrio cholerae involves multiple TonB-dependent haem receptors

Vibrio cholerae has multiple iron transport systems, one of which involves haem uptake through the outer membrane receptor HutA. A hutA mutant had only a slight defect in growth using haemin as the iron source, and we show here that V. cholerae encodes two additional TonB-dependent haem receptors, HutR and HasR. HutR has significant homology to HutA as well as to other outer membrane haem receptors. Membrane fractionation confirmed that HutR is present in the outer membrane. The hutR gene was co-transcribed with the upstream gene ptrB, and expression from the ptrB promoter was negatively regulated by iron. A hutA, hutR mutant was significantly impaired, but not completely defective, in the ability to use haemin as the sole iron source. HasR is most similar to the haemophore-utilizing haem receptors from Pseudomonas aeruginosa and Serratia marcescens. A mutant defective in all three haem receptors was unable to use haemin as an iron source. HutA and HutR functioned with either V. cholerae TonB1 or TonB2, but haemin transport through either receptor was more efficient in strains carrying the tonB1 system genes. In contrast, haemin uptake through HasR was TonB2 dependent. Efficient utilization of haemoglobin as an iron source required HutA and TonB1. The triple haem receptor mutant exhibited no defect in its ability to compete with its Vib- parental strain in an infant mouse model of infection, indicating that additional iron sources are present in vivo. V. cholerae used haem derived from marine invertebrate haemoglobins, suggesting that haem may be available to V. cholerae growing in the marine environment.     

Iron acquisition systems in the pathogenic Neisseria

Pathogenic neisseriae have a repertoire of high-affinity iron uptake systems to facilitate acquisition of this essential element in the human host. They possess surface receptor proteins that directly bind the extracellular host iron-binding proteins transferrin and lactoferrin. Alternatively, they have siderophore receptors capable of scavenging iron when exogenous siderophores are present. Released intracellular haem iron present in the form of haemoglobin, haemoglobin-haptoglobin or free haem can be used directly as a source of iron for growth through direct binding by specific surface receptors. Although these receptors may vary in complexity and composition, the key protein involved in the transport of iron (as iron, haem or iron-siderophore) across the outer membrane is a TonB-dependent receptor with an overall structure presumably similar to that determined recently for Escherichia coli FhuA or FepA. The receptors are potentially ideal vaccine targets in view of their critical role in survival in the host. Preliminary pilot studies indicate that transferrin receptor-based vaccines may be protective in humans.     

The receptor for 2,3,7,8-tetrachlorodibenzo-p-dioxin in the mouse hepatoma cell line Hepa 1c1c7. A comparison with the glucocorticoid receptor and the mouse and rat hepatic dioxin receptors

The molecular properties of the receptor for 2,3,7,8-tetrachlorodibenzo-p-dioxin in the mouse hepatoma cell line Hepa 1c1c7 were investigated. The receptor was found to represent a highly asymmetrical molecule with a sedimentation coefficient, s20,w, of approximately 8 S, a Stokes radius of 7-8 nm, and a calculated Mr approximately equal to 260,000-300,000. In comparison, the Hepa 1c1c7 glucocorticoid receptor in analogy to the glucocorticoid receptor in general as well as the C57BL/6 mouse and rat hepatic dioxin receptors are molecules with an s20,w value of 4-5 S, a Stokes radius of approximately 6 nm, and a calculated Mr approximately equal to 100,000. In the presence of 20 mM sodium molybdate, a large Mr approximately equal to 270,000-310,000 form of the Hepa 1c1c7 glucocorticoid receptor is stabilized which is hydrodynamically indistinguishable from the Mr approximately equal to 260,000-300,000 Hepa 1c1c7 dioxin receptor. Sodium molybdate does not have any effect on the molecular properties of the Hepa 1c1c7 dioxin receptor. In conclusion, the large form of dioxin receptor present in Hepa 1c1c7 mouse hepatoma cells in the absence of sodium molybdate is strikingly similar to molybdate-stabilized steroid hormone receptors as well as the molybdate-stabilized form of the dioxin receptor previously demonstrated in rat hepatic cytosol. Therefore, the Hepa 1c1c7 dioxin receptor might offer an interesting model for studies on the structure and function of Mr approximately equal to 300,000 forms of soluble receptors.     

The signaling potential of the receptors for insulin and insulin-like growth factor I (IGF-I) in 3T3-L1 adipocytes: comparison of glucose transport activity, induction of oncogene c-fos, glucose transporter mRNA, and DNA-synthesis.

The receptors for insulin and insulin-like growth factor I (IGF-I) have in common a high sequence homology and diverse overlapping functions, (e.g., the stimulation of acute metabolic events and the induction of cell growth.). In the present study, we have compared the potential of insulin and IGF-I receptors in stimulating glucose transport activity, glucose transporter gene expression, DNA-synthesis, and expression of proto-oncogene c-fos in 3T3-L1 adipocytes which express high levels of both receptors. Binding of both hormones to their own receptors was highly specific as compared with binding to the respective other receptor (insulin receptor: KD = 3.6 nM, KI of IGF-I greater than 500 nM; IGF-I receptor, KD = 1.1 nM, KI of insulin = 191 nM). Induction of proto-oncogene c-fos mRNA by insulin and IGF-I paralleled their respective receptor occupancy and was thus induced by both hormones via their own receptor (EC50 of insulin, 3.7; IGF-I, 3.9 nM). Similarly, both insulin and IGF-I increased DNA synthesis (EC50 of insulin, 5.8 nM; IGF-I, 4.0 nM), glucose transport activity (EC50 of insulin, 1.7 nM; IGF-I, 1.4 nM), and glucose transporter (GLUT4) mRNA levels in concentrations corresponding with their respective receptor occupancy. These data indicate that in 3T3-L1 cells the alpha-subunits of insulin and IGF-I receptors have an equal potential to stimulate a metabolic and a mitogenic response.     

Transferrin iron interactions with cultured hepatocellular carcinoma cells (PLC/PRF/5)

Hepatocellular carcinoma cells of the PLC/PRF/5 cell line had 1.9 x 10(5) transferrin receptors per tumor cell with a Kd of 1.5 x 10(-8) M. At high concentrations of transferrin the binding was not saturable. Transferrin internalization by hepatoma cells was shown by time and temperature-dependent binding studies and by pronase experiments. Transferrin recycling was confirmed by the demonstration of a progressive increase in the cellular molar ratios of iron to transferrin and by chase experiments. Ammonium chloride interfered with iron unloading. The vinca alkaloid vincristine inhibited iron and transferrin uptake. The hepatocarcinoma cells appeared to lack asialoglycoprotein receptors and therefore internalized partially desialated transferrin by the regular route. Iron uptake from transferrin was markedly inhibited by the hydrophobic ferrous chelator 2,2' bipyridine but was relatively unaffected by the hydrophilic ferric chelator desferroxamine. The implication that ferrous iron was involved in postendocytic transvesicular membrane iron transport was supported by a study in which hepatoma cells were shown to take up large amounts of ferrous iron suspended in 270 mM sucrose at pH 5.5. The interaction at this pH between surface labeled hepatoma cell extracts and ferrous iron on a Sephacryl S-300 column suggested that the postendocytic transvesicular transport of iron through the membrane was in part protein mediated. The endocytosed iron in hepatoma cells was found in association with ferritin (33%), transferrin (31%) and a low molecular weight fraction (21%).     

Hemopexin joins transferrin as representative members of a distinct class of receptor-mediated endocytic transport systems

Receptor-mediated transport of heme by hemopexin in vivo and in vitro results in catabolism of heme but not the protein, suggesting that intact apohemopexin recycles from cells. However, until now, the intracellular transport of hemopexin by receptor-mediated endocytosis remained to be established. Biochemical studies on cultured human HepG2 and mouse Hepa hepatoma cells demonstrate that hemopexin is transported to an intracellular location and, after endocytosis, is subsequently returned intact to the medium. During incubation at 37 degrees C, hemopexin accumulated intracellularly for ca. 15 min before reaching a plateau while surface binding was saturated by 5 min. No internalization of ligand took place during incubation at 4 degrees C. These and other data suggest that hemopexin receptors recycle, and furthermore, incubation with monensin significantly inhibits the amount of cell associated of heme-[125I]hemopexin during short-term incubation at 37 degrees C, consistent with a block in receptor recycling. Ammonium chloride and methylamine were less inhibitory. Electron microscopic autoradiography of heme-[125I]hemopexin showed the presence of hemopexin in vesicles of the classical pathway of endocytosis in human HepG2 hepatoma cells, confirming the internalization of hemopexin. Colloidal gold-conjugated hemopexin and electron microscopy showed that hemopexin bound to receptors at 4 degrees C is distributed initially over the entire cell surface, including microvilli and coated pits. After incubation at 37 degrees C, hemopexin-gold is located intracellularly in coated vesicles and then in small endosomes and multivesicular bodies. Colocalization of hemopexin and transferrin intracellularly was shown in two ways. Radioiodinated hemopexin was observed in the same subcellular compartment as horseradish peroxidase conjugates of transferrin using the diaminobenzidine-induced density shift assay. In addition, colloidal gold derivatives of heme-hemopexin and diferric transferrin were found together in coated pits, coated vesicles, endosomes and multivesicular bodies. Therefore, hemopexin and transferrin act by a similar receptor-mediated mechanism in which the transport protein recycles after endocytosis from the cell to undergo further rounds of intracellular transport.     

Elemental iron does repress transferrin, haemopexin and haemoglobin receptor expression in Haemophilusinfluenzae

The iron repressible nature of Haemophilus influenzae transferrin binding proteins suggests a regulatory role for elemental iron in their expression. The existence of a Haemophilus ferric uptake repressor (Fur) binding motif identified in the promoter region of both tbpA and tbpB further supports this hypothesis. However, a recent study using brain heart infusion growth medium suggested that transferrin binding protein synthesis in H. influenzae was haem- rather than iron-regulated. The present study re-investigates this observation and using a chemically defined medium, we demonstrate that elemental iron haem or protoporphyrin IX can each regulate Haemophilus influenzae transferrin, haemopexin and haemoglobin receptor expression.     

Quelling the red menace: haem capture by bacteria

Haem is an important bacterial nutrient. As a prosthetic group of several proteins, haem functions as a cofactor mediating oxygen transport, energy generation, and mixed-function oxidation. In addition, the iron chelated in the porphyrin ring may serve as an iron substrate for growth. However, because of its propensity for oxidizing cellular constituents, haem is always associated with proteins. Therefore, the uptake and transit of haem across bacterial membranes requires the participation of protein escorts. Bacteria have evolved a diverse array of surface-exposed receptors dedicated to binding haem and haem-proteins. Following this selective recognition at the bacterial cell surface, haem is transported across the outer membrane via a TonB-dependent process. The control of receptor expression appears to be multifactorial, probably involving a number of global regulators. A model integrating this information is presented.     

The dioxin receptor: a comparison with the glucocorticoid receptor

The physico-chemical properties of the dioxin and glucocorticoid receptors from rat liver and wild-type and mutant cell lines were investigated and compared. In rat liver, the receptors are virtually indistinguishable. Both are highly asymmetrical proteins with axial ratios of 12-15, have Stokes radii of 6 nm and sedimentation coefficients of approximately 4 S. This results in a calculated apparent mol. wt of approximately 100,000. The dioxin receptor from the mouse hepatoma cell line Hepa 1c1c7 represents an atypical form of the dioxin receptor with a pronounced tendency to aggregate to form Mr approximately equal to 300,000 complexes in high ionic strength and in the absence of sodium molybdate. In the presence of sulphydryl reducing agents, however, the Hepa 1c1c7 dioxin receptor dissociates to an Mr approximately 100,000 species. In analogy to the nt- mutant glucocorticoid receptor in mouse lymphoma cells, there is no gross change in the structure of the nt- dioxin mutant in mouse hepatoma cells compared with the wild-type receptor. The nt- dioxin receptor does, however, have a reduced affinity for DNA.     

Erythropoietin. Receptor characteristics during the ontogeny of hamster yolk sac erythroid cells

Erythropoietin (Epo) binds specifically to receptors on the surface membrane of responsive erythroid cells. In search of ontogenic changes in Epo receptor behavior, we studied characteristics of specific binding to hamster yolk sac erythroid cells during hamster ontogeny. We detected receptors specific for Epo on these cells throughout the duration of their intravascular existence (hamster gestational days 8 through 13). These receptors are saturable at an Epo concentration of 1.2 nM in the incubation medium. Attainment of equilibrium of binding prior to hormone internalization, a requirement for receptor binding assays, was possible at 10 degrees C but not at 37 degrees C. Hence, all incubations of cells with Epo were carried out at 10 degrees C. Data on specific binding analyzed by the method of Scatchard demonstrated that yolk sac erythroid cells possess a single class of Epo receptors at each stage of gestation examined. Binding affinity and numbers of receptors per cell change as ontogeny progresses: Kd (the dissociation constant) increases, a phenomenon observed in other differentiating cell systems, whereas the number of receptors per cell peaks on gestational day 10. The variability in number of receptors per cell is consonant with up and down regulation controlled by Epo availability. We propose that the progressive increase in Kd might be best explained by ontogenic changes in cell membrane structure contiguous to the receptors themselves.     

Sodium transport by the acetylcholine receptor of cultured muscle cells.

Activation of the acetylcholine receptors of cultured muscle cells by carbamylcholine increases the rate of passive 22-Na+ uptake into the muscle cells up to 20-fold. The Na+ transport activity of the receptor desensitizes during exposure to carbamylcholine. The rate and extent of desensitization is reduced by lowering the assay temperature from 36 degrees to 2 degrees, allowing accurate measurements of initial rates of Na+ transport by the receptor. Activation of the receptor by carbamylcholine and acetylcholine is significantly cooperative (Hill coefficients of 1.4 to 2.0). Inhibition by D-tubocurarine is not cooperative. The carbamylcholine-induced Na+ transport activity of the receptor is inhibited 50% by 4 muM D-tubocurarine, 100 muM atropine, or 1.6 nM diiodo-alpha-bungarotoxin but is not affected by tetrodotoxin. The initial rate of Na+ transport by the receptor is temperature-independent between 2 degrees and 36 degrees. Receptor Na+ transport is saturable by Na+ at 2 degrees with an apparent Km of 150 plus and minus 20 mM. Saturation by Na+ not observed at 36 degrees at the concentrations tested. Saturation by Na+ is observed at 2 degrees both under conditions of net Na+ influx and under conditions of isotopic exchange at equilibrium. The receptor does not catalyze obligatory exchange diffusion at a detectable rate. Comparison of binding of [125-I]diiodo-alpha-bungarotoxin with rates of Na+ transport indicates a turnover number of 2 times 10-7 ions per min per receptor. These results are discussed in terms of the mechanism of Na+ transport by the receptor.     

Expression of haptoglobin receptors in human hepatoma cells.

The uptake of radio-labeled hemoglobin-haptoglobin complex (Hb-Hp) by human hepatoma PLC/PRF/5 and HepG2 cells was investigated in an attempt to characterize the uptake process and intracellular transport. Human hepatoma cells took up Hb-Hp in a receptor-mediated manner. Scatchard analysis of binding revealed that PLC/PRF/5 and HepG2 cells exhibited about 21,000 and 63,000 haptoglobin receptors/cell, with a dissociation constant (Kd) of 8.0 and 17 nM, respectively. Human hepatocytes in primary culture also expressed about 84,000 receptors/cells, with a Kd of 7.4 nM. The hemoglobin-haptoglobin complex was internalized and subsequently the internalized Hb-Hp was slowly degraded in the cells. Preincubation of the cells with Hb-Hp resulted in a decrease in binding of the radioactive Hb-Hp to the cell surface, and was accompanied with an accumulation of intracellular receptors. The uptake of Hb-Hp by the cells was not inhibited by 100 microM chloroquine or by 10 mM methylamine, but was inhibited by 50 microM monodansylcadaverine. Hemoglobin-heme taken up by the cells induced microsomal heme oxygenase. Thus, human hepatoma PLC/PRF/5 and HepG2 cells can take up Hb-Hp by haptoglobin receptor-mediated endocytosis and Hb-Hp probably causes translocation of the haptoglobin receptors from the cell surface to the cell interior where they can be degraded. The internalized heme-moiety of hemoglobin can regulate the expression of heme oxygenase.     

(Thr-59)-insulin-like growth factor I stimulates 2-deoxyglucose transport in BC3H1 myocytes through the insulin-like growth factor receptor, not the insulin receptor

The murine non-fusing muscle cell line contains distinct receptors for insulin and insulin-like growth factors. Pretreatment of myocytes with insulin for 20 h at 37 degrees C inhibits the binding of [125I]iodoinsulin by 60% without affecting the binding of [125I]iodoinsulin-like growth factor I. The ED50 values for down-regulation of the insulin and insulin-like growth factor receptor by their respective ligands are 1 nM and 3 nM, respectively. Insulin, (Thr-59)-insulin-like growth factor I and multiplication-stimulating activity stimulate 2-[3H]deoxyglucose transport in myocytes with ED50 values of 5 nM, 5.6 nM and 33 nM, respectively. In order to determine whether (Thr-59)-insulin-like growth factor I stimulates 2-[3H]deoxyglucose transport in myocytes via its own receptor or the insulin receptor, we determined the activity of these peptides after down-regulation of the insulin receptor. The rate of 2-[3H]deoxyglucose transport in myocytes pretreated with insulin (5 nM) is elevated but returns to control levels by 1 h after the washout of insulin. The dose-response curve for insulin-stimulated 2-[3H]deoxyglucose transport is shifted to the right (ED50 greater than 100 nM) immediately after insulin washout but is normal by 1 h after insulin washout. In contrast, the dose-response curve for (Thr-59)-insulin-like growth factor I is unchanged in insulin-pretreated cells immediately after insulin washout. These data show that (Thr-59)-insulin-like growth factor I stimulates 2-[3H]deoxyglucose transport in myocytes by acting through an insulin-like growth factor receptor and not through the insulin receptor. Since multiplication-stimulating activity is 6-fold less active than (Thr-59)-insulin-like growth factor, they both may be acting through a type 1 insulin-like growth factor receptor.     

Stimulation of facilitated [3H]uridine transport by thyroid hormone in GH1 cells. Evidence for regulation by the thyroid hormone nuclear receptor

We have previously shown that 3,5,3'-triiodo-L-thyronine (L-T3) stimulates cell growth and a 4- to 8-fold increase in growth hormone mRNA in GH1 cells. These effects appear to be mediated by a thyroid hormone nuclear receptor with an equilibrium dissociation constant for L-T3 of 0.2 nM and an abundance of about 10,000 receptors per cell nucleus. In this report, we show that L-T3 exerts a pleiotypic effect on GH1 cells to rapidly (within 2 h) stimulate [3H]uridine uptake to a maximal value of 2.5- to 3-fold after 24 h. This results from an increase in the number of functional uridine "transport sites" as shown by studies documenting an increase in the apparent Vmax with no change in the Km, 17 microM. Although the labeling of the cellular uridine pool and pools of all phosphorylated uridine derivatives was increased by L-T3, there was no change in the relative amounts of the individual pools in cells incubated with or without hormone. The intracellular concentration of [3H]uridine was estimated to be similar to that of the medium, suggesting that facilitated transport mediates [3H]uridine uptake. That this increase in [3H]uridine transport was nuclear receptor-mediated is supported by the excellent correspondence of the L-T3 dose-response curve for [3H]uridine uptake and that for L-T3 binding to receptor. Finally, inhibition of protein synthesis by cycloheximide and RNA synthesis by actinomycin D demonstrated that the L-T3 effect required continuing protein and RNA synthesis. These results are consistent with an effect of the L-T3-nuclear receptor complex to increase uridine uptake in GH1 cells by altering the expression of gene(s) essential for the transport process.