Tetraspanins: molecular organisers of the leukocyte surface Jacqueline M. Tarrant 1 , Lorraine Robb 1 , Annemiek B. van Spriel 2 and Mark D. Wright 2 1 The Walter and Eliza Hall Institute, 1G Royal Parade, Parkville, Victoria, 3050 Australia 2 The Austin Research Institute, A&RMC, Studley Road, Heidelberg, Victoria, 3084, Australia Tetraspanins are a large superfamily of cell surface membrane proteins characterised by their four trans- membrane domains. They are expressed in a wide var- iety of cell types and have functional roles in processes, such as cellular adhesion, motility, activation and tumour invasion. Leukocytes express # 20 tetraspanin proteins on their surface. Tetraspanins have an excep- tional ability to interact with a diverse array of import- ant proteins, such as integrins, immunoreceptors and signalling molecules. They are thought to organise a novel type of cell surface membrane microdomain, which in leukocytes functions to regulate activation, motility and antigen presentation. The importance of tetraspanin microdomains in immunity is highlighted by the perturbations in the immune system observed in tetraspanin-deficient mice. The tetraspanins (also known as the transmembrane four superfamily) are a large superfamily of cell surface membrane proteins. They are best characterised by conserved sequences in their four transmembrane domains and by the CCG and PXSC motifs in their major extracellular domain (EC2) [1–3]. There are several other families of proteins that also have four transmem- brane domains but these can be distinguished from genuine tetraspanins by the absence of these conserved amino acid residues [2,4]. The crystal structure of the EC2 domain of human CD81 has been recently solved [5] and molecular modelling of the EC2 of other tetraspanins indicates they have comparable features [6]. No structural information is available on the highly conserved trans- membrane domains. Interest in the role of tetraspanins in the immune system began with functional screens for novel lymphocyte surface antigens that uncovered several tetraspanin molecules (reviewed in Refs [1,7]). Now, with the avail- ability of the sequence of the human genome, 28 human genes encoding tetraspanins have been documented [8]. Of these, we estimate 20 tetraspanins can be expressed at the surface of leukocytes (Table 1) [9–27]. This multiplicity of tetraspanin proteins makes it difficult to identify the specific role of individual tetraspanin molecules in the immune response. Although tetraspanins mediate import- ant functions in many different cell types, this Review concentrates on the role of tetraspanins in immunity. How new insights into tetraspanin biochemistry have refined hypotheses concerning their function in immune regu- lation will be discussed. The results of recent gene targeting experiments that show individual tetraspanins to have significant, specific functions in the immune response will then be described. Tetraspanin microdomains The major role of tetraspanins is to organise other proteins into signal transducing complexes at the cell surface [3,8,28,29]. The ability of tetraspanins to interact with each other, and the observation that signals transduced through various tetraspanin family members often lead to similar functional outcomes, has led to the hypothesis that tetraspanins predominantly exist in a supramolecular complex or microdomain, often referred to as the tetraspanin web [30]. The characterisation of microdomains with different physical properties and specialised functions is a current major topic in cell biology. Several investigators have sought to determine whether tetraspanins and associated proteins cluster in unique membrane microdomains or form a component of other recognised membrane domains. Of specific interest are recent reports evaluating tetraspanin complexes in relation to lipid microdomains (‘rafts’). Tetraspanin com- plexes are autonomous units, existing both within lipid rafts and as non-raft domains [31–33]. The existence of tetraspanin complexes has also been reported in type III glycosynapses, which are microdomains of glycosphingo- lipids that co-exist in, or with, lipid rafts [34]. Thus, biochemical data strongly support the hypothesis that the primary role of tetraspanins is to organise other proteins into supramolecular complexes at the cell surface. A major issue, not yet fully resolved, is the degree of functional overlap in tetraspanin biology. Precisely which functions are commonly mediated by tetraspanins and which are specific to individual family members? Recent advances in the biochemistry of interactions that occur within the tetraspanin web have started to answer this question. Tetraspanin–protein interactions have been classified into three categories based on their stringency in different detergents [28,31] (Box 1). The weakest and perhaps most controversial are the level 3 interactions. However, several of these have been independently confirmed using a variety of techniques, such as density centrifugation and gel filtration analyses [31], fluor- escence resonance energy transfer [35], site-directed Corresponding author: Mark D. Wright (m.wright@ari.unimelb.edu.au). Review TRENDS in Immunology Vol.24 No.11 November 2003 610 http://treimm.trends.com 1471-4906/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. doi:10.1016/j.it.2003.09.011