PerspectiveCell Biology

A Cellular Striptease Act

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Science  13 Nov 1998:
Vol. 282, Issue 5392, pp. 1279-1280
DOI: 10.1126/science.282.5392.1279

The cell surface is a dynamic place [HN1], [HN2]. During its life history the cell alters the repertoire of proteins displayed on its surface many times. Membrane-anchored adhesion molecules, receptors, ligands, and enzymes are removed and replaced as the cell proceeds through development and as its activation state changes [HN3], [HN4]. How is this wholesale refurnishing of the cell membrane orchestrated? One key mechanism is proteolytic processing of the ectodomain (extracellular domain) of such membrane proteins. Cleavage or shedding of the ectodomains of plasma membrane proteins—widely observed in cells in culture—is blocked by inhibitors of metalloproteinases (1, 2). This result suggests that transmembrane and soluble metalloproteinases, such as matrix metalloproteinases (MMPs) and their relatives, are rate-limiting for cleavage and shedding. Other evidence also implicates serine proteinases in these processing events (3, 4).

The first such “sheddase” characterized was the tumor necrosis factor-α (TNF-α) converting enzyme (TACE) (5) [HN5]. The study by Peschon and colleagues (6) on page 1281 of this issue now points to TACE's essential role in the shedding of ectodomains during mouse development. The surprise comes from the observation that mice lacking TACE do not show a phenotype indicative of a lack of TNF-α availability. Rather, they show the same phenotype as mice engineered to be without the epidermal growth factor (EGF) receptor—because TACE-mediated proteolysis makes available ligands for the EGF receptor, particularly transforming growth factor-α (TGF-α).

TACE turns out to be a membrane-anchored proteinase that is a member of the ADAM (a disintegrin and metalloproteinase) domain family of proteins that combines features of both cell surface adhesion molecules and proteinases (8) [HN6]. ADAMs all have a common domain organization, which endows these proteins with several potential functions—proteolysis, adhesion, signaling, and fusion (see figure below). The proteolytically competent ADAMs, such as TACE (ADAM17), are zinc-dependent metalloproteinases, closely related to the MMP family with which they share small molecule inhibitors and even one tissue inhibitor, TIMP-3 (9, 10). Several newly discovered MMPs appear to be hybrids of both MMP and ADAM domains (11), indicating that these two types of enzymes are part of one, larger family.

The ADAM proteinases are themselves targets of proteolytic events that ultimately strip off the catalytic domains (5, 8). This action could be a mechanism of functionally blunting the effects of the proteinases (see the figure on the next page). These soluble ADAMs may have proteolytic activity, as is the case for snake venom enzymes (8), but soluble TACE is much less active than membrane-bound enzyme (5, 6). The residual adhesive domains of ADAMs left after cleavage may have regulatory or adhesive functions. In support of this idea, a catalytic domain-deleted mutant of Kuz (ADAM10/SUP17), first identified as being required for cleavage of Notch during neural development in Drosophila, exerts a dominant negative effect (8, 12) [HN7]. During sperm maturation fertilin, a heterodimeric ADAM essential for sperm-egg interaction (13), also loses its catalytic domains by proteolytic processing. The remaining adhesive disintegrin domain is then competent to bind integrins.

How does TACE act? TACE is widely expressed in the animal. Mutation of the catalytic domain of TACE (6) reveals several distinct functions for this ADAM in development. Ligands for the EGF receptor, which is essential for epithelial development (7), are usually made and used locally (14). Although the growth factor precursors may have some biological activity (15), the new results imply that the membrane-anchored forms are essentially inactive precursors (6). TACE also cleaves ectodomains of other receptors and ligands, such as TNF-α, the p75 TNF receptor, and L-selectin, and thus participates in inflammatory and pathological reactions (6).

Processing membrane proteins by the ADAMs and other sheddases requires both the membrane-anchored enzyme and its substrate to be present in cis on the same cell (6, 8, 12). This presents several interesting problems. How are the active cell surface proteinases kept separate from their cell surface substrates until shedding is triggered? How do you exert selectivity for only certain ectodomain targets, out of many transmembrane proteins displayed on the cell surface? And how are the proteinase and substrate brought together in a coordinated manner so that all the cell surface substrate molecules can be removed within seconds, as occurs for the adhesion molecules L-selectin [HN8] and syndecans (4, 16)?

Despite nonconserved cleavage sites that may be adjacent to the membrane or further out on the molecule, there are clues that a common strategy may operate in most cases. First, all ectodomain shedding is inhibited in a single mutant cell line (1). Second, the proteolysis is regulated in different cell types by activation of protein kinase C (PKC) [HN9], calcium/calmodulin kinases, or receptor tyrosine kinases (1, 17). A model that accounts for these observations requires the processing proteinases and their transmembrane substrates both to be anchored in distinct domains of the plasma membrane, probably through cytoskeletal interactions (see figure below). Upon cell activation, the attachments change and the proteinases and substrates become coclustered and can interact. Alternatively, the signaling cascade could modify the cytoplasmic domains of the proteinases or substrate, producing a conformational change that either activates the enzyme or makes the cleavage site available.

Activation of sheddases.

The ADAM proteases (as dimers) and substrates are anchored apart in the plane of the membrane. Upon activation (via protein kinases and other pathways) they are brought together and proteolysis takes place, leading to free ectodomains.

Although activation of the shedding reaction appears to control the rapid and complete removal of cell surface molecules such as L-selectin (an adhesion molecule involved in leukocyte rolling and extravasation into inflammatory sites) for most processing reactions there appears to be a constitutive level of ectodomain shedding. Processing is necessary to make available paracrine growth and survival factors such as TGF-α, EGF, HB-EGF, the kit ligand, and amphiregulin (18). This makes sense to allow for the consistent supply of growth factors (see figure below).

Versatile shedding.

Sheddases can supply or down-regulate ligands for receptors. Cleavage of adhesion molecules on cell surface or exposure of the disintegrin domain of ADAM regulate cell-cell and cell-extracellular matrix interactions.

Endogenous inhibitors allow even finer control of the action of the shedding enzymes. Recently TACE was shown to be inhibited by TIMP-3, but not by the three other TIMPs that also inhibit MMPs (10). If TACE liberates a survival factor, then the presence of TIMP-3 could lead to cell death. This may explain why TIMP-3, but not other TIMPs, induce apoptosis (19) [HN10].

Proteolysis of the ectodomains of growth factor coreceptors such as syndecan provide a second mechanism for regulating growth factor availability. Shedding the ectodomain of syndecan converts it to a potent inhibitor of FGF-2 (20). Just as shedding can make growth factor ligands available and control proliferation and survival, cleavage can also control cell death. Membrane-bound Fas ligand induces apoptosis by binding to the Fas receptor. Proteolysis functionally down-regulates the ligand and short-circuits apoptosis in lymphoid cell (21).

Cell surface adhesive molecules can also be regulated by proteolysis. An emerging paradigm is that cleavage of adhesive molecules not only alters adhesion, but completely revamps cell signaling. In the case of Notch, cleavage by Kuz is required to make it functional as a receptor, promoting adhesion, signaling, and cell lineage choices (12). Shedding of L-selectin by TACE or related enzymes inhibits leukocyte rolling and blunts their extravasation to inflammatory sites (16). The shedding of the ectodomains of E-cadherin (22) and transmembrane protein tyrosine phosphatases such as LAR have profound effects on cell-cell adhesions and also on important signaling pathways (17). These changing adhesion receptors and ligands may also be part of the apparatus for pathfinding in the nervous system.

Cells use a limited number of strategies to remodel their microenvironments. It is clear that the shedding process is an ancient, conserved, and fundamental pathway present from worms to humans. Thus, proteolysis by cell surface shedding enzymes provides a mechanism by which the wardrobe of externally displayed molecules can be changed or discarded. Spatial restriction of the enzymes and their substrates allows for either instant action or sustained activity.

HyperNotes Related Resources on the World Wide Web

General Hypernotes

The MIT Biology Hypertextbook, developed by the Experimental Study Group at the Massachusetts Institute of Technology, provides background information on the biology of cells.

Pedro's BioMolecular Research Tools is a collection of WWW links to information and services useful to molecular biologists. It provides links to molecular biology search and analysis tools; bibliographic, text, and Web search services; guides and tutorials; and biological and biochemical journals and newsletters.

The World Wide Web Virtual Library: Biosciences points to virtual library pages for Biological Molecules and for Biochemistry and Molecular Biology. Each of these pages presents a long list of Web resources. The World Wide Web Virtual Library Biological Molecules covers molecular sequence and structure databases, metabolic pathway databases, and other lists of Web resources. The World Wide Web Virtual Library: Biochemistry and Molecular Biology is a list of resources listed by provider.

Cell & Molecular Biology Online is a well-organized list of Web resources for cell and molecular biologists. For each resource, a brief description is provided.

Cells Alive!, an educational service of Quill Graphics, provides timely and visually exciting material about cells of the immune system, bacteria, and parasites. The section on apoptosis provides a description of programmed cell death, illustrations, and a well-annotated list of links to other resources. The section on the cytoskeleton includes a video which illustrates the redistribution of the cytoskeleton during phagocytosis.

Numbered Hypernotes

1. Cells I and Introduction to Membranes by Anthony Moss describes cells and the cell surface. This page was written to accompany a course in biology at Auburn University.

2. An On-Line Biology Book by M. J. Farabee includes a chapter on Cells II: Cellular Organization. This chapter describes the plasma membrane and provides a diagram. Links to related Web resources are available.

3. Membranes and their proteins describes biological membranes and the categories of proteins associated with them. This page is a section of The Principles of Protein Structure, an Internet course developed at Birkbeck College, the University of London.

4. Membranes are described in NetBiochem, a multimedia support center for the health sciences. This page covers the chemical components of membranes, membrane structure, compartmentalization, and membrane receptors.

5. Tumor Necrosis Factor outlines the biological activity of TNF-alpha, describes diseases associated with TNF, and presents the structure of TNF.

6. The ADAM family of proteins is briefly described.

7. Kuzbanian Evolutionary Homologs describes the functions of ADAMs in Drosophila. This page is a component of the Interactive Fly, a guide to Drosophila genes and their roles in development.

8. Neutrophil in Action outlines the role of neutrophils and L-selectin in the acute inflammatory process. This animated page is a chapter of WebPath, an electronic course in pathology.

9. The Protein Kinase Resource (PKR) is a Web-accessible compendium of information on the protein kinase family of enzymes. This resource includes tools for structural and computational analyses as well as links to related information maintained by others.

10. The Cell Death Society provides definitions, illustrations, and other resources for studying apoptosis.

11. Department of Anatomy, University of California, San Francisco.

References and Notes

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