Abstract-CD36 is 1 of the class B scavenger receptor expressed on monocytes, monocyte-derived macrophages (M), platelets, and adipocytes. In our previous studies, we reported that the uptake of oxidized low density lipoproteins (OxLDLs) is reduced by Ϸ50% in M from CD36-deficient patients compared with that in control subjects. Recently, we have shown that CD36 is highly expressed in human atherosclerotic aorta. Possibilities have been raised that besides the wide distribution and multifunctional characteristics of CD36, this molecule may also be involved in the mediation of intracellular signaling. The aim of the present study was to elucidate the role of CD36 in cytokine secretion and to investigate the CD36-mediated intracellular signaling stimulated by OxLDL. On addition of OxLDL or thrombospondin-1, the M from CD36-deficient patients secreted significantly less amounts of tumor necrosis factor-␣ (TNF-␣) and interleukin-1 (IL-1) compared with those from controls. RNase protection assay with multiprobe template sets demonstrated that after incubation with OxLDL, the mRNAs of a variety of cytokines, including genes encoding IL-1Ra, IL-1, IL-6, TNF-␣ and -, and interferon (IFN)-␥ and -, were significantly lower in the M of patients. The addition of antibody against CD36 attenuated this OxLDL-induced response in controls. We also observed a reduced response in nuclear factor-B (NF-B) activity in OxLDL-stimulated M from CD36-deficient patients. Unlike OxLDL, stimulation by lipopolysaccharide induced an increase in NF-B activity in M from CD36-deficient patients, suggesting that lipopolysaccharide-mediated signaling was conserved. These results demonstrate that in addition to the reduced OxLDL uptake that we reported previously, CD36-deficient patients may also have an impaired response of OxLDL-induced NF-B activation and subsequent cytokine expression. (Arterioscler Thromb Vasc Biol.
Abstract-The scavenger receptor class B type I (SR-BI) and its human homologue CLA-1 (CD36 and LIMPII Analogous-1) have recently been identified to bind HDL and mediate the selective uptake of HDL lipids. Tissue distribution of both murine and human receptors is quite similar, in that they are expressed abundantly in liver and steroidogenic tissues. However, expression and function of the human SR-BI (hSR-BI), in the periphery of reverse cholesterol transport such as macrophages, are still unclear. In the present study, we have raised two different kinds of anti-hSR-BI polypeptide antibodies (Abs): one against the extracellular domain and the other against the intracellular domain. We have investigated the expression of hSR-BI mRNA and immunoreactive mass in freshly isolated cultured human monocyte-derived macrophages (hM) and in atherosclerotic lesions. Contrary to the earlier report, hSR-BI mRNA was expressed in cultured hM and markedly upregulated with differentiation, determined by Northern blot and reverse transcriptase-based polymerase chain reaction analyses. The mRNA expression pattern during differentiation of hM was very similar to those of SR class A and another member of SR class B, CD36. Protein expression was confirmed by Western blot analyses with the above Abs to show a major 83-kDa band. Modified lipoproteins such as oxidized LDL and acetylated LDL induced a 5-fold increase in mRNA and protein expression of hSR-BI. Confocal immunofluorescence microscopy demonstrated that hSR-BI immunoreactive mass was detectable as a heterogeneous, punctate staining pattern. Furthermore, immunohistochemical analysis showed that immunoreactive mass of hSR-BI was detected in foam cells in human aortic atherosclerotic lesions and that there was no significant difference of staining patterns between the two Abs. This study clearly demonstrates that hSR-BI is expressed in the lipid-laden macrophages in human atherosclerotic lesions, suggesting that it is very important to know its function and regulation in hM to understand the biological utility of this molecule. (Circ Res. 1999;85:108-116.)
Seven dominant mutations showing greatly enhanced resistance to the glucose repression of galactokinase synthesis have been isolated from GAL81 mutants, which have the constitutive phenotype but are still strongly repressible by glucose for the synthesis of the Leloir enzymes. These glucose-resistant mutants were due to semidominant mutations at either of two loci, GAL82 and GAL83. Both loci are unlinked to the GAL81-gal4, gal80, or gal7-gallO.-gall locus or to each other. The GAL83 locus was mapped on chromosome V at a site between arg9 and chol. The GAL82 and GAL83 mutations produced partial resistance of galactokinase to glucose repression only when one or both of these mutations were combined with a GAL81 or a gal80 mutation. The GAL82 and GAL83 mutations are probably specific for expression of the Leloir pathway and related enzymes, because they do not affect the synthesis of a-D-glucosidase, invertase, or isocitrate lyase.In the on-off control of the lac operon in Escherichia coli, there is dual control of enzyme synthesis (9): negative control by a repressor coded by the lacI gene, and positive control by a complex molecule consisting of a specific catabolite gene activator protein (CAP protein) and cyclic adenosine 3',5'-monophosphate. The repressor detects the presence or absence of,-galactoside in the cytoplasm, and the CAP protein detects glucose through the alteration of the intracellular level of cyclic adenosine 3',5'-monophosphate. These signals are conveyed to the appropriate sites in the promoter region of the lac operon.Glucose repression or carbon catabolite repression is also commonly observed in yeasts.Recent biochemical studies clearly demonstrate that glucose repression of cytochrome c synthesis in Saccharomyces cerevisiae is at the level of gene transcription (32, 33), whereas the contribution of cyclic adenosine 3',5'-monophosphate is, in general, ambiguous in yeasts. To elucidate the genetic mechanism of carbon catabolite repression in yeast, mutants with altered regulatory properties have been isolated and characterized by several workers. These include mutations producing resistance to carbon catabolite repression in invertase synthesis (15, 24), pleiotropic mutations conferring resistance to carbon catabolite repression (5, 6), the hex mutation (13) with reduced activity of glucose phosphorylation, which contributes to carbon catabolite repression, and the ADR mutations (4, 7) in a regulatory system for glucose repression of alcohol dehydrogenase synthesis. The genetic data, however, are still insufficient to construct a genetic model for carbon catabolite repression.In previous communications (22, 23), we proposed a genetic model for the role of the inducer in the synthesis of the galactose pathway enzymes (the Leloir enzymes) in S. cerevisiae. The structural genes for the Leloir enzymes, the gall locus (encodes galactokinase [EC 2.7.1.6]), gal7 (encodes a-D-galactose-1-phosphate uridylyltransferase [EC 2.7.7.12]), and gallO (encodes uridine diphosphoglucose 4-epimerase [EC 5.1.3....
Abstract-CD36 has been reported to be a receptor for oxidized LDL (Ox-LDL). In our previous study, the uptake of Ox-LDL in CD36-deficient macrophages was reduced by approximately 50% compared with that in control macrophages, suggesting an important role of CD36 as a receptor for Ox-LDL in humans. In the current study, we examined the immunohistochemical localization of CD36 in human aorta in comparison with that of scavenger receptor class A type I and type II (SRA). Cryostat sections were made from aortic tissues. For immunohistochemical staining, the following antibodies were used: (1) FA6-152, anti-CD36 antibody, and (2) SRI-2, which recognizes both type I and type II SRAs. Immunohistochemical staining for CD36 and SRA was performed using labeled streptavidin method. In macrophages scattered in aortic walls without atherosclerotic lesions, the expression of CD36 was hardly observed, whereas that of SRA was detected weakly but consistently. In contrast, in atherosclerotic lesions, macrophages around the core region showed a weak immunoreactivity to CD36 and a strong immunoreactivity to SRA. Furthermore, lipid-laden macrophages, which mainly existed in the core region, had a strongly positive immunoreactivity to CD36, but a low or moderate level of immunoreactivity to SRA. The distributions of CD36 and SRA were different from each other, and especially foamed, large-sized macrophages in atherosclerotic plaques tended to more abundantly express CD36 protein. These data demonstrate, for the first time, that the expression of both CD36 and SRA might be differentially regulated in aortic walls, and might play different roles in the formation of foam cells in atherosclerotic lesions.
Two dominant uninducible mutant alleles in the gal80 locus were identified. The GAL80s-1 and GAL80s-2 mutants showed novel phenotypes in response to the newly isolated GAL81-1 mutant allele, a dominant constitutive mutation linked to the gal4 locus; the GAL80s-1 GAL81-1 strain was inducible and the GAL80s-2 GAL81-1 strain was uninducible. Many galactose positive revertants from the GAL80s-2 GAL81-1 strain were isolated. It was proved that each revertant was due to a secondary mutation either in the gal80 or GAL81 locus, whereas revertants due to mutation at the supposed controlling site for the structural gene cluster of the galactose-pathway enzymes have not been isolated.
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