Characterization, size estimation and solubilization of α-macroglobulin complex receptors in liver membranes

Gliemann, Jørgen; Davidsen, Ole; Moestrup, Søren K.

doi:10.1016/0005-2736(89)90320-9

Cited by 23 publications

(12 citation statements)

References 26 publications

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“…At this pH, in the range of 20-200 g/ml the binding of P-A2M was dose-dependent, contrasting with that of N-A2M. It could be inhibited by an excess of unlabeled protein, a characteristic shared with that described for the A2M receptor (Debanne et al 1976;Van Leuven et al 1979;Gliemann et al 1989;Moestrup and Gliemann 1991). Considering that in vertebrates, A2MR belongs to the LDL family of receptors (reviewed in Moestrup 1994), our results, associated with those showing the binding of LDL to the T. cruzi surface (Soares and De Souza 1991), lead us to speculate that an A2M/LDL receptor occurs in T. cruzi.…”

Section: Discussionmentioning

confidence: 86%

Alpha-2-macroglobulin binds to the surface of Trypanosoma cruzi

Coutinho

Cavalcanti

Leuven

et al. 1997

Parasitology Research

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Trypanosoma cruzi, the causative agent of Chagas disease, infects vertebrate cells after an initial step of parasite/host-cell recognition. Alpha-2-macroglobulin (A2M), an important type of physiological proteinase inhibitor found in tissues and in the plasma of mammals, inhibits cell invasion by T. cruzi and accumulates in sites of the inflamed myocardium associated with parasite antigens. To study whether A2M would bind to T. cruzi, an indirect immunofluorescence reaction was performed using two different anti-mouse A2M sera. Intense labeling was observed in the membrane lining the cell body and the flagellum of bloodstream trypomastigotes obtained from experimentally infected mice in the peak of parasitemia, suggesting that the antisera recognize plasma A2M associated with the parasite surface. Metacyclic trypomastigotes obtained in a serum-free defined medium reacted with anti-A2M only after previous incubation with purified human A2M. Enzyme-linked immunosorbent assay (ELISA) studies were applied to characterize better the binding of native (N-A2M) and of proteinase-complexed (P-A2M) forms of A2M. The "in vitro" binding of N-A2M to trypomastigotes was better at pH 5.0, followed by pH 10.0 and pH 7.4. Cysteinly and serine proteinase inhibitors, E-64 and STI, respectively, inhibited the reaction. P-A2M also bound to T. cruzi in a dose-dependent way. Flow-cytometry studies showed that about 80% of the parasites stained with fluorescein isothiocyanate (FITC)-labeled P-A2M (50 micrograms/ml) with high affinity at pH 7.4 (but also at pH 10.0) in a process that was reverted by the addition of unlabeled P-A2M or the calcium-chelator agent EDTA and also by incubation at an acid pH (4.0). These results suggest that (a) native-A2M binds to T. cruzi proteinase(s) and (b) T. cruzi expresses a receptor(s) that binds proteinase-complexed A2M.

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Section: Discussionmentioning

confidence: 86%

Alpha-2-macroglobulin binds to the surface of Trypanosoma cruzi

Coutinho

Cavalcanti

Leuven

et al. 1997

Parasitology Research

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“…Additionally, expression of CD91 was observed to correlate with the ability of an APC to re-present gp96-chaperoned peptides (9). CD91 has been shown to be a receptor for the serum protein ␣2-macroglobulin (␣ 2 M) (13), and this protein was shown to inhibit re-presentation of gp96-chaperoned peptides. Collectively, these observations led to the suggestion that CD91, a pedigreed receptor for ␣ 2 M and a host of other proteins (see ref.…”

mentioning

confidence: 99%

Essential role of CD91 in re-presentation of gp96-chaperoned peptides

Binder

Srivastava

2004

Proc. Natl. Acad. Sci. U.S.A.

185

152

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Heat shock proteins (HSPs) such as gp96 are released from cells as a result of necrotic cell death. The ability of endogenous HSPpeptide complexes to elicit antigen-specific T cells requires representation of the chaperoned peptides by antigen-presenting cells. Re-presentation requires the uptake of HSP-peptide complexes through a receptor, suggested to be the low-density lipoprotein receptor-related protein or CD91. We have used short interfering RNA for CD91 to show that, as antigen-presenting cells lose expression of CD91, their re-presenting ability undergoes a corresponding and dramatic decline. Furthermore, as the cells recover from extinction of CD91 expression, they regain the ability to re-present peptides. The ability of cells to bind ␣2 macroglobulin, a previously known CD91 ligand, or HSP gp96, and their ability to process peptides chaperoned by ␣2 macroglobulin, undergo identical variations. These results have been obtained from studies in vitro and from an assay that measures re-presentation in vivo. In additional studies in vivo, protective tumor immunity elicited by tumor-derived gp96 -peptide complexes is shown to be abrogated by anti-CD91 antisera. These studies show that CD91 is essential for re-presentation of gp96-chaperoned peptides by MHC molecules and have an important bearing on the mechanism of immunogenicity of necrotic cells.heat shock proteins ͉ tumor immunity ͉ ␣2-macroglobulin ͉ receptor-associated protein ͉ LRP H eat shock proteins (HSPs) purified from antigen-expressing cells chaperone antigenic peptides generated in that cell (1-4). Immunization with such HSP-peptide complexes purified from tumors or pathogen-infected cells elicits specific immunity directed against the tumor or pathogen, respectively (5-7). The mechanism of immunogenicity of HSP-peptide complexes is increasingly clear. The HSP-peptide complex is targeted to the antigen-presenting cells (APCs) through interaction of HSP with a receptor identified as CD91 (8,9). In case of the HSP gp96, it appears that the HSP-peptide complex is endocytosed into a compartment positive for Fc receptor and MHC I and negative for Rab5a, CD1, and transferrin (10). The subsequent fate of the HSP molecule itself remains unclear, but the peptide is transported to the cytosol through a mechanism yet to be determined. The peptide is further transported into the endoplasmic reticulum through a transporterassociated with antigen processing-dependent (9, 11) or -independent (see refs. 11 and 12) mechanism. Following a classical pathway within the endoplasmic reticulum, the peptide is charged onto a cognate MHC I molecule, and the MHC I-peptide complex present at the cell surface now stimulates cognate CD8 ϩ T cells. Because the endocytosed HSP-peptide complex is taken up into a compartment that is MHC I ϩ (10), the possibility that the gp96-chaperoned peptide is charged onto the MHC I in that compartment, independently of the endoplasmic reticulum, merits consideration.Among the several HSPs that follow this path, the endoplasmic reticulum-r...

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“…The liver membrane preparations from rat and human livers were prepared by Ultra-Turrax homogenization followed by differential centrifugation as described [27]. The protein content was determined with the dye binding method [28] using bovine serum albumin as a standard and a commercial reagent (Bio-Rad, Munich, FRG).…”

Section: Samples For Light Microscopic Autoradiographymentioning

confidence: 99%

Cellular targets and receptors for interleukin‐6 I. In vivo and in vitro uptake of IL‐6 in liver and hepatocytes

Sonne

Davidsen

Møller

et al. 1990

Eur J Clin Investigation

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Interleukin-6 (IL-6) is a potent stimulator of the hepatic synthesis of acute-phase proteins. 125I-labelled IL-6 disappeared from the blood of rats with an overall half-time of about 1.5 min; 41% of the injected tracer dose was recovered in the liver by 15 min. The clearance was biphasic. The simultaneous injection of tracer and an excess of unlabelled IL-6 eliminated the initial rapid phase, and reduced the hepatic uptake to 14%. Light microscopic autoradiography showed 5% of the grains over non-hepatocytes, and 80% over hepatocytes, accumulating in areas around the bile canaliculi. Thereafter, degradation products accumulated in the bile. At 4 degrees C, isolated rat hepatocytes bound IL-6 with an apparent Kd of 39 pmol l-1 to a uniform class of 4500 receptors per cell with an apparent molar mass of 115-120 kg mol-1. The HepG2 human hepatocellular cell line bound IL-6 with an apparent Kd of 21 pmol l-1 to a uniform class of 1200 receptors per cell with an apparent molar mass of 155-160 kg mol-1. At 37 degrees C, both cell types endocytosed the bound ligand slowly, and degradation products appeared in the medium after a relatively long lag period (40 min in hepatocytes and 1 h in HepG2 cells).

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Characterization, size estimation and solubilization of α-macroglobulin complex receptors in liver membranes

Cited by 23 publications

References 26 publications

Alpha-2-macroglobulin binds to the surface of Trypanosoma cruzi

Alpha-2-macroglobulin binds to the surface of Trypanosoma cruzi

Essential role of CD91 in re-presentation of gp96-chaperoned peptides

Cellular targets and receptors for interleukin‐6 I. In vivo and in vitro uptake of IL‐6 in liver and hepatocytes

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