Structures of Class A Macrophage Scavenger Receptors

Section: Discussionmentioning

confidence: 93%

mentioning

confidence: 99%

Ligand Binding to Macrophage Scavenger Receptor-A Induces Urokinase-type Plasminogen Activator Expression by a Protein Kinase-dependent Signaling Pathway

Hsu¹,

Hajjar²,

Khan³

et al. 1998

“…This notion is based on the fact that neither this segment of MARCO (VHNEDAGVECS) nor the corresponding one of Tesr (GHAEDAGVTCTVP) contains any positively charged residues, which are considered to be crucial for the interaction with AcLDL, a polyanionic macromolecule. The differences in AcLDL binding cannot be because of any major structural alterations either, because both segments contain a cysteine residue, which participates in the formation of an intradomain disulfide bond (3,27,28). This bond fixes the position of the very C-terminal end.…”

Section: Discussionmentioning

confidence: 99%

A Phage Display Screen and Binding Studies with Acetylated Low Density Lipoprotein Provide Evidence for the Importance of the Scavenger Receptor Cysteine-rich (SRCR) Domain in the Ligand-binding Function of MARCO

Chen

Sankala

Ojala

et al. 2006

MARCO is a class A scavenger receptor capable of binding both Gram-negative and -positive bacteria. Using the surface plasmon resonance technique, we show here that a recombinant, soluble form of MARCO, sMARCO, binds the major Gram-negative and -positive bacterial surface components, lipopolysaccharide and lipoteichoic acid. Yet, the interaction of these two polyanions with sMARCO is of much lower affinity than that of polyinosinic acid, a polyanionic inhibitor of bacterial binding to MARCO. To further elucidate the ligand-binding functions of MARCO, we performed a phage display screen with sMARCO. The screening resulted in the enrichment of only a handful of phage clones. Contrary to expectations, no polyanionic peptides, but only those with a predominantly hydrophobic nature, were enriched. One peptide, VRWGSFAAWL, was displayed on two-thirds of the phages recovered after four rounds of screening. The VRWGSFAAWL phage-sMARCO interaction had significantly slower dissociation kinetics than that between sMARCO and lipopolysaccharide or lipoteichoic acid. Further work with this phage, and the second most enriched phage, displaying the peptide RLNWAWWLSY, demonstrated that both peptides bind to the SRCR domain of MARCO, and that they probably bind to the same site. Data base searches suggested that the VRWGSFAAWL peptide represents complement component C4, but we could not convincingly confirm this suggestion. A study with chimeric scavenger receptors indicated that even minor sequence changes in the MARCO scavenger receptor cysteine-rich (SRCR) domain can have profound effects on the binding of the prototypic scavenger receptor ligand, acetylated low density lipoprotein. As shown by differential binding of glutathione S-transferase-VR-WGSFAAWL, these differences were very likely due to conformational changes. These findings led to experiments that demonstrated a crucial role of the SRCR domain for acetylated low density lipoprotein binding in MARCO. Thus, our results strengthen the notion that the SRCR domain is the major ligand-binding domain in MARCO. Furthermore, they suggest that the domain may contain multiple ligand-binding sites.MARCO, a close relative of scavenger receptor A (see Ref. 1), is a trimeric type II transmembrane protein with an N-terminal intracellular domain, a transmembrane domain, and an extracellular portion composed of a short spacer domain, a long triple-helical collagenous domain, and a C-terminal scavenger receptor cysteine-rich (SRCR) 3 domain (2). MARCO has a very restricted expression pattern in adult mice living under pathogen-free conditions. It is expressed at significant levels only in the marginal zone macrophages of the spleen, in macrophages of the medullary cord of lymph nodes, and in the peritoneal macrophages (2). 4 In bacterial infections MARCO expression is up-regulated in macrophages of most tissues (3-6). Cells transfected with a plasmid encoding MARCO avidly bind both Gram-negative and -positive bacteria, but not yeast (2, 3). MARCO also has other than microbial ligand...

“…These results, however, cannot distinguish whether the multimerization was produced by inter-molecular or by intra-molecular disulfide bonds. Recent studies on the structure of SRCR domains demonstrated that intra-molecular disulfide bonds are critical for formation of the native structure of the domain (17).…”

Section: Soluble Hensin Exists In Different Forms In the Twomentioning

confidence: 99%

Only Multimeric Hensin Located in the Extracellular Matrix Can Induce Apical Endocytosis and Reverse the Polarity of Intercalated Cells

Hikita

Takito

Vijayakumar

et al. 1999

When an intercalated epithelial cell line was seeded at low density and allowed to reach confluence, it located the anion exchanger band 3 in the apical membrane and an H ؉ -ATPase in the basolateral membrane. The same clonal cells seeded at high density targeted these proteins to the reverse location. Furthermore, high density cells had vigorous apical endocytosis, and low density cells had none. The extracellular matrix of high density cells was capable of inducing apical endocytosis and relocation of band 3 to the basolateral membrane in low density cells. A 230-kDa extracellular matrix (ECM) protein termed hensin, when purified to near-homogeneity, was able to reverse the phenotype of the low density cells. Antibodies to hensin prevented this effect, indicating that hensin is necessary for conversion of polarity. We show here that hensin was synthesized by both low density and high density cells. Whereas both phenotypes secreted soluble hensin into their media, only high density cells localized it in their ECM. Analysis of soluble hensin by sucrose density gradients showed that low density cells secreted monomeric hensin, and high density cells secreted higher order multimers. When 35 Slabeled monomeric hensin was added to high density cells, they induced its aggregation suggesting that the multimerization was catalyzed by surface events in the high density cells. Soluble monomeric or multimeric hensin did not induce apical endocytosis in low density cells, whereas the more polymerized hensin isolated from insoluble ECM readily induced it. These multimers could be disaggregated by sulfhydryl reagents and by dimethylmaleic anhydride, and treatment of high density ECM by these reagents prevented the induction of endocytosis. These results demonstrate that hensin, like several ECM proteins, needs to be precipitated in the ECM to be functional.The plasma membrane of epithelial cells is polarized into two domains; apical and basolateral, each of which has characteristic protein and lipid composition (1-3). Although targeting sequences were found in the structure of several polarized proteins, recent studies have shown that some proteins are polarized in a cell type-specific manner; for instance the Na,KATPase is targeted to the apical membrane of retinal pigment epithelium but to the basolateral membrane of most other epithelia (4). Several other proteins exhibit this flexibility in targeting; the most dramatic example is the targeting of the proton-translocating ATPase and the Cl:HCO 3 exchanger of the intercalated cell of the renal tubule (reviewed in Ref. 5). These cells exist in a spectrum of forms. One extreme, the ␣ type, has an apical H ϩ -ATPase and a basolateral anion exchanger that is an alternately spliced form of the erythroid band 3 (kAE1) 1 and hence is capable of trans-epithelial secretion of H ϩ . In contrast, the ␤ form secretes HCO 3 by a basolateral H ϩ -ATPase and an apical kAE1 (6). (However, one study did not find kAE1 by immunoblot analysis but confirmed the presence of its mRNA (7).) In an...