Nick Platt

Recognizing death: the phagocytosis of apoptotic cells.

Although apoptotic cell death is widespread, dying cells are rarely seen in situ because of their rapid clearance by neighbouring phagocytes. Phagocytic recognition of apoptotic cells is less well understood than the death programme itself, but an increasing number of recent studies are highlighting its importance. This review discusses the nature of the receptors that have been implicated in apoptotic cell phagocytosis, the mechanisms of uptake and the immunological consequences of apoptotic cell ingestion.

Is the class A macrophage scavenger receptor (SR-A) multifunctional? - The mouse's tale.

The class A macrophage scavenger receptor (SR-A) is the prototypic member of an expanding family of membrane receptors collectively termed scavenger receptors (SRs) (1–3). Receptors of this group recognize a number of ligands, including chemically modified or altered molecules and, in particular, the modified lipoproteins that are pertinent to the development of vascular disease. As shown in Table ​Table1,1, all characterized SR-A ligands are polyanionic, although many polyanions fail to bind SR-A. Here, we wish to summarize and comment upon the evidence for SR-A being both a multiligand and multifunctional receptor, considering both the available in vitro data and the results of several studies of SR-A–deficient mice. Two other SRs, namely SR-B1 (Krieger, ref. 4) and LRP (Herz and Strickland, ref. 5), are the subject of separate reviews in this Perspective series. Table 1 SR-A ligands. Receptor recognition or nonrecognition has been shown for these molecules, either through direct binding studies or the inhibition of modified lipoprotein endocytosis. Two themes that can be drawn from studies of SRs as a whole are particularly relevant to this discussion. First, because these receptors display broad and seemingly overlapping ligand-binding properties, biological specificity is likely to be determined not only by ligand structure and the signal transduced following ligand binding, but also by other considerations. These include the distribution and availability of the various SRs, their ability to interact with other receptors, and their relative affinities for the various ligands. Second, the apparent redundancy of ligand binding is achieved despite the absence of conserved protein sequences among the distinct classes of SRs. A mechanistic understanding of individual SRs’ broad yet specific ligand recognition and of the features shared by these unrelated molecules will require the resolution of tertiary structures of multiple SRs.

Implications for invariant natural killer T cell ligands due to the restricted presence of isoglobotrihexosylceramide in mammals

Development of invariant natural killer T (iNKT) cells requires the presentation of lipid ligand(s) by CD1d molecules in the thymus. The glycosphingolipid (GSL) isoglobotrihexosylceramide (iGb3) has been proposed as the natural iNKT cell-selecting ligand in the thymus and to be involved in peripheral activation of iNKT cells by dendritic cells (DCs). However, there is no direct biochemical evidence for the presence of iGb3 in mouse or human thymus or DCs. Using a highly sensitive HPLC assay, the only tissue where iGb3 could be detected in mouse was the dorsal root ganglion (DRG). iGb3 was not detected in other mouse or any human tissues analyzed, including thymus and DCs. Even in mutant mice that store isoglobo-series GSLs in the DRG, we were still unable to detect these GSLs in the thymus. iGb3 is therefore unlikely to be a physiologically relevant iNKT cell-selecting ligand in mouse and humans. A detailed study is now warranted to better understand the nature of iNKT cell-selecting ligand(s) in vivo.

Identification and characterization of murine SCARA5, a novel class A scavenger receptor that is expressed by populations of epithelial cells.

Epithelia are positioned at a critical interface to prevent invasion by microorganisms from the environment. Pattern recognition receptors are important components of innate immunity because of their ability to interact with specific microbe-associated structures and initiate immune responses. Several distinct groups of receptors have been recognized. One of these, the scavenger receptors, has been classified into at least eight separate classes. The class A scavenger receptors are characterized by the presence of a collagen-like domain and include macrophage scavenger receptor type A (SR-A1 I/II, SCARA1) and MARCO (SCARA2). These receptors are known to make important contributions to host defense. Here, we identify a novel murine scavenger receptor, SCARA5, which has a structure typical of this class. The cDNA encodes 491 amino acids, which predict a type II protein that contains C-terminal intracellular, transmembrane, extracellular spacer, collagenous, and N-terminal scavenger receptor cysteine rich domains. Expression in Chinese hamster ovary cells confirmed that the receptor assembles as a homotrimer and is expressed at the plasma membrane. SCARA5-transfected cells bound Escherichia coli and Staphylococcus aureus, but not zymosan, in a polyanionic-inhibitable manner. Unlike other class A scavenger receptors, the receptor was unable to endocytose acetylated or oxidized low density lipoprotein. Quantitative RT-PCR and in situ hybridization demonstrate SCARA5 has a tissue and cellular distribution unique among class A scavenger receptors. Because of the restriction of SCARA5 transcripts to populations of epithelial cells, we propose that this receptor may play important roles in the innate immune activities of these cells.

Scavenger receptors: diverse activities and promiscuous binding of polyanionic ligands

Scavenger receptors are a diverse family of proteins that share a common property--the binding of modified lipoprotein--but they have recently been shown to recognise a diverse range of ligands. Understanding the molecular interaction of receptor-ligand binding should provide insight into how scavenger receptors contribute to important biological processes.

Apoptotic Thymocyte Clearance in Scavenger Receptor Class A-Deficient Mice Is Apparently Normal

Studies of apoptotic cell uptake by phagocytes in vitro have implicated a number of different receptors capable of mediating ingestion. However, there is currently little evidence for involvement of any of these candidate receptors in vivo. Previously, we have shown by the use of a blocking mAb against the class A scavenger receptor (SR-A) and thymic macrophages prepared from SR-A null mice, that this receptor is responsible for ∼50% of the uptake of apoptotic thymocytes in vitro. In this study we have investigated the frequency of dying cells in the thymus of mice lacking SR-A. Our inability to demonstrate increased frequencies of nonphagocytosed Annexin V+, TUNEL+, or propidium iodide+ apoptotic thymocytes suggests there is no deficiency in apoptotic thymocyte clearance in these mice. Even when the rate of thymocyte apoptosis was increased by exposure of receptor-deficient mice to gamma irradiation, we did not detect a difference in the numbers of dying cells compared with similarly treated wild-type animals. This provides the first direct evidence of redundancy in apoptotic cell clearance mechanisms in vivo.

The many roles of the class A macrophage scavenger receptor.

The class A macrophage scavenger receptor (SR-A) is the prototypic example of a group of plasma membrane receptors collectively known as scavenger receptors. SR-A displays the ability to bind and endocytose large quantities of modified lipoprotein. Hence, it is thought to be one of the main receptors involved in mediating lipid influx into macrophages (Mphi), which promotes their conversion into foam cells that are abundant in the atherosclerotic lesion. However, as a result of increased interest and research effort and through the development of specific reagents and animal models, it is now appreciated to be multifunctional. These roles include Mphi growth and maintenance, adhesion to the substratum, cell-cell interactions, phagocytosis, and host defense. In this review, we attempt to summarize the evidence and argue that these kinds of activities underlie the biological versatility of Mphi.

Cloning and expression of the beta-N-acetylglucosaminidase gene from Streptococcus pneumoniae. Generation of truncated enzymes with modified aglycon specificity.

The gene encoding a β- N-acetylglucosaminidase from Streptococcus pneumoniae has been obtained by screening an expression library for β- N-acetylglucosaminidase activity. Clones of different nucleotide sizes each having arylglycoside activity were obtained, and DNA sequencing revealed a gene of 3933 base pairs possessing typical bacterial transcription initiation and termination sequences and terminating in an ochre stop codon. Computer analysis of the translated protein of 1311 amino acids (144,210 Da) identified a tandem repeat within which lies a sequence homologous with six other hexosaminidase gene products from a wide variety of species ranging from bacteria to humans. Also found were an amino-terminal putative secretion signal peptide and a carboxyl-terminal cell sorting/anchorage motif typically found in over 20 other Gram-positive surface proteins. The expression of an almost complete DNA clone in Escherichia coli produced a functional and authentic β- N-acetylglucosaminidase with aglycon specificity identical to the wild-type enzyme. However, enzymes produced from truncated DNA clones show more restricted aglycon specificity and are unable to hydrolyze terminal β1-2GlcNAc residues from N-glycans containing a bisecting N-acetylglucosamine. The availability of these clones allows structural analyses to be made of catalytic and oligosaccharide recognition protein domains that enhance functional activity.

Macrophage Membrane Molecules: Markers of Tissue Differentiation and Heterogeneity

Monoclonal antibodies directed against murine macrophage differentiation antigens provide tools to characterize novel glycoproteins, their expression by different macrophage subpopulations in situ and regulation by cytokines and other agents in vitro. Studies are in progress to determine ligands for these molecules, and to establish possible functions. We summarize current knowledge of the following molecules: F4/80, a glycoprotein with homology to the G-protein linked transmembrane 7 hormone receptor family; macrosialin, a member of the lysosomal-associated membrane protein [lamp] family with a macrophage-specific mucin-like extracellular domain also present in its human homologue, CD68; sialoadhesin, a sialic acid binding lectin with multiple Ig superfamily domains; mannosyl receptor, a lectin-like molecule with multiple C-type lectin domains; type 3 complement receptor, a beta 2 integrin involved in cell migration and adhesion; scavenger receptor, a transmembrane homotrimer with a collagenous domain.

Expression of the class A macrophage scavenger receptor on specific subpopulations of murine dendritic cells limits their endotoxin response

Dendritic cells (DC) function at the interface of innate and acquired immunity and are uniquely sensitive to specific stimuli. Pattern recognition receptors (PRR) on these cells are critically important because of their ability to recognise and initiate responses to conserved microbial‐associated molecular signatures. With the exception of Toll‐like receptors (TLR), we know relatively little about the specific distribution of other PRR amongst populations of DC. Here, we describe the expression of the murine class A macrophage scavenger receptor (SR‐A) and show that it is restricted to specific subpopulations of bone marrow‐derived and splenic DC. Importantly, we demonstrate that the receptor significantly alters the response of DC to endotoxin. In contrast to the activities of other PRR that have so far been examined, uniquely SR‐A limits the maturation response; SR‐A–/– cells display enhanced CD40 expression and TNF‐α production. We discuss the potential contributions of SR‐A to DC biology in the context of the known multiple activities of this receptor.