We describe a cell surface protein that is abundant in liver and has close structural and biochemical similarities to the low density lipoprotein (LDL) receptor. The complete sequence of the protein containing 4544 amino acids is presented. From the sequence a remarkable resemblance to the LDL‐receptor and epidermal growth factor (EGF) precursor is apparent. Three types of repeating sequence motifs entirely account for the extracellular domain of the molecule. These are arranged in a manner resembling four copies of the ligand binding and the EGF‐precursor homologous region of the LDL‐receptor. Following a proline‐rich segment of 17 amino acids are found six consecutive repeats with close homology to EGF. A single membrane‐spanning segment precedes a carboxy‐terminal ‘tail’ of 100 amino acids. This contains two seven‐amino acid sequences with striking homology to the cytoplasmic tail of the LDL‐receptor in the region that contains the signal for clustering into coated pits. The mRNA for this protein is most abundant in liver, brain and lung. By using an antibody raised against a 13‐amino acid peptide corresponding to the deduced amino acid sequence of the carboxy‐terminus of the protein we have demonstrated its existence on the cell surface and its abundance in liver. Like the LDL‐receptor this protein also strongly binds calcium, a cation absolutely required for binding of apolipoproteins B and E to their receptors. We propose that this LDL‐receptor related protein (LRP) is a recycling lipoprotein receptor with possible growth‐modulating effects.

Construction of a family of bacterial expression vectors, pEX1‐3, is described. These vectors are derived from a cro‐lacZ gene fusion plasmid which expresses large quantities of fusion protein under the control of the PR promoter of bacteriophage lambda. A polylinker has been engineered into the 3′ end of the lacZ gene in all three translational reading frames, and stop signals for transcription and translation inserted, so that any open reading frame DNA may be expressed as a hybrid beta‐galactosidase protein. cDNA fragments cloned in these vectors can be detected with an efficiency of greater than 1 in 3, thus enabling the detection of rare cDNA molecules. In addition, the low solubility of hybrid proteins leads to a rapid isolation procedure allowing antibodies of pre‐determined specificity to be made against expressed regions of cloned DNA. We describe the cloning of albumin and complement C9 genes from a human cDNA library using polyclonal and monoclonal antibodies.

Organelle-specific integral membrane proteins were identified by a novel strategy which gives rise to monospecific antibodies to these proteins as well as to the cDNA clones encoding them. A cDNA expression library was screened with a polyclonal antiserum raised against Triton X-114-extracted organelle proteins and clones were then grouped using antibodies affinity-purified on individual fusion proteins. The identification, molecular cloning and sequencing are described of a type 1 membrane protein (TGN38) which is located specifically in the trans-Golgi network.

High density lipoprotein is the major carrier of lipid hydroperoxides in human blood plasma from fasting donors (atherosclerosis/low density lipoprotein/oildation/ubiquinol/metabolism)

The low-density-lipoprotein (LDL) receptor is a cell-surface protein that plays an important part in the metabolism of cholesterol by mediating the uptake of LDL from plasma into cells. Although LDL particles bind to the LDL receptor through their apolipoprotein B (apo B) and apolipoprotein E (apo E) moieties, other apo E-containing particles, like chylomicron remnants, are not dependent on the LDL receptor for uptake into cells. Chylomicrons formed in the intestinal mucosa during the absorption of the products of digestion, are processed by the peripheral circulation by lipoprotein lipase, which catalyses the breakdown of triglycerides in chylomicrons to free fatty acids and glycerol. The resulting chylomicron remnants, which are cholesterol-rich lipoproteins, are subsequently taken up in the liver. A second distinct protein that binds to apo E-containing lipoproteins, but not to LDL, has been proposed to be the receptor mediating the clearance of chylomicron remnants from the plasma. This protein has a relative molecular mass (Mr) of 56,000 (56K). More recent studies have failed, however, to establish whether this protein is a cell-surface receptor. Here we describe crosslinking experiments in which apo E liposomes were found to bind specifically to the cell surface of hepG2 cells and to human liver membranes. The size and immunological cross-reactivity of the protein to which the liposomes bound was indistinguishable from that of the recently cloned and sequenced LDL-receptor-related protein, LRP. We therefore conclude that the LRP might function as an apo E receptor.

Sorting of proteins destined for different plasma membrane domains, lysosomes and secretory pathways takes place in the trans‐Golgi network (TGN). TGN38 is an integral membrane protein found in this intracellular compartment. We show that TGN38 contains an autonomous targeting signal within its cytoplasmic domain which determines its intracellular location. Deletion analysis and site‐directed mutagenesis of this domain demonstrate that a tyrosine motif homologous to the internalization signal of surface receptors is necessary and sufficient for correct localization. These findings suggest that TGN38 is maintained in the TGN by retrieval from the plasma membrane and employs a different mechanism for retention from that of the transferase enzymes of the trans‐Golgi.

Perturbations in the IL-7/IL-7R system were clearly associated with disease progression but did not reliably predict immune reconstitution.

Background-Male sex is an independent risk factor for the extent and severity of atherosclerosis. The influence of androgens on foam cell formation, a key event in atherogenesis, has not yet been investigated. Methods and Results-Primary human monocytes were allowed to differentiate into macrophages. RNA was then extracted from healthy male-donor (nϭ8) and premenopausal female-donor (nϭ8) macrophages, and message for the androgen receptor (AR) was examined by RT-PCR. There was a significantly higher level of AR mRNA in macrophages isolated from men than in those from women (0.64Ϯ0.06 versus 0.15Ϯ0.02 amol/g total RNA; PϽ0.001). AR mRNA levels were similar in macrophages from postmenopausal and premenopausal women (Pϭ0.16). The functional consequence of this sex difference was then explored. Lipid-loading studies were performed on male (nϭ9) macrophages treated with the androgen dihydrotestosterone (DHT) and/or the AR antagonist hydroxyflutamide. These showed that DHT caused a dose-dependent and receptor-mediated increase in macrophage cholesteryl ester content (109Ϯ10%, 117Ϯ3%, and 120Ϯ4% for 4, 40, and 400 nmol/L DHT, respectively, as a percentage of control, Pϭ0.002; 95Ϯ8% for DHT with hydroxyflutamide, Pϭ0.58 versus controls). By contrast, there was no significant effect of androgen on lipid loading in female-donor macrophages (PϾ0.2 versus controls). Conclusions-Sex differences in androgen-mediated macrophage lipid loading may contribute to the greater prevalence and severity of atherosclerosis in men. en have an earlier onset and higher incidence of atherosclerosis than women. 1 The potential protective effects of estrogens on vascular structure and function have been studied extensively. Much less work, however, has addressed the possible effects of androgens on atherogenesis. In animal models, androgen treatment may increase plaque formation in the aorta and coronary vessels. 2,3 Furthermore, in humans, we have recently demonstrated that androgens enhance monocyte adhesion to endothelial cells 4 and are associated with impaired vascular reactivity. 5 Foam cell formation is a key early event in atherosclerosis and is largely due to the uptake of modified lipoproteins, principally LDL, by peripheral blood monocyte-derived macrophages (MDMs) in the arterial wall. 6,7 The possibility that androgen receptors (ARs) and androgen-related intracellular pathways may be involved in the process of lipid loading and foam cell formation in atherosclerosis has not been explored previously. We therefore investigated whether the AR is expressed in primary human MDMs and whether there is a sex difference in its expression and have examined the biological effects of androgen stimulation on cholesteryl ester (CE) formation/lipid loading in these cells. Methods Isolation of Human MonocytesWhite cell concentrates (Red Cross Blood Bank) were obtained from the peripheral blood of individual healthy men (20 to 58 years old) and premenopausal (18 to 45 years old) and postmenopausal (45, 55, and 72 years old) women. The postmenopau...