Cell death induction by receptors of the TNF family: towards a molecular understanding (1997)

It has been known for a long time that programmed cell death (PCD) plays an essential role in the formation of a multicellular, differentiated animal [1]. A striking example is the development of the nematode Caenorhabditis elegans. The fully formed animal consists of 1089 cells, of which 131 are programmed to die. This system has provided a breakthrough in understanding of the molecular processes underlying PCD. Indeed, genetic studies first revealed the presence of genes needed for PCD, such as ced-3 and ced-4, as well as other genes, such as ced-9, which counteract PCD [2]. In addition to its role in development, PCD occurs prominently in various immune processes, such as negative selection of T-cells in the thymus, elimination of overactivated lymphocytes in the periphery, etc. But PCD—or lack of it—has especially become a focus of considerable interest because of its relevance to many diseases [3]. For example, cancer cells have often become resistant to PCD-inducing stimuli, various lympho-proliferative and/or autoimmune diseases evade ‘death’ signals, etc. On the other hand, in some neuro-degenerative disorders such as Parkinson, Alzheimer or amyotrophic lateral sclerosis, one may wish to interfere with PCD. Cells dying in the course of development are in fact fairly difficult to study, because the cell corpses are rapidly phagocytosed by neighbouring cells. Therefore, morphological and biochemical processes associated with PCD can best be studied in cell culture systems, and the typical events leading to cell death are usually referred to as ‘apoptosis’. ‘PCD’ and ‘apoptosis’ are often used interchangeably, although the former has more a connotation of genetically predetermined, while the latter is more linked to morphological and biochemical changes [4]. The principal characteristics of apoptosis are blebbing of the plasma membrane, phosphatidylserine externalization, cytoskeletal disruption, accumulation and/or activation of transglutaminase, condensation of nuclear chromatin, fragmentation of nuclear DNA to approximately 50 kb segments, and subsequently to internucleosomal fragments; at later stages, cytoplasm and nucleus become compartmentalized and form membrane-bound apoptotic bodies, which are engulfed by neighbouring cells or infiltrated tissue macrophages [4]. Cell death by apoptosis is a very neat way to eliminate unwanted cells: no traces are left and the cell contents are never released or accessible to the immune system. Hence, there is no inflammation. This is in contrast to death by necrosis. Under these conditions, normally the cell swells and then, when membrane integrity comes under attack, the cell collapses like a balloon and the contents spill out into the extracellular milieu [5]. This may result in an inflammatory response. Although PCD is sometimes referred to as ‘cell suicide’, in most systems studied the process of apoptosis is initiated by external inducers or treatments. Some examples are UV- or γ-irradiation, heat shock, oxidative stress, chemotherapeutic drugs (directed to topoisomerases like etoposide, to protein kinases like staurosporine, to DNA or its building blocks like nitrogen mustard and methotrexate, respectively, etc.), viral infection, loss of matrix attachment, glucocorticoids, growth factor withdrawal, soluble or membrane-bound cytokines like TNF or Fas/Apo-1 (CD95) ligand, and many others. In general, each cell contains precursors to apoptogenic mediators, as well as counteracting, protecting proteins. It is the balance between these two which determines the outcome after a given stimulus. It is remarkable how morphologically and biochemically similar apoptotically dying cells are, almost irrespective not only of the cell type, but also of the species. This suggests already that the molecular mechanisms of apoptosis are ubiquitous in nature and highly conserved in evolution. Indeed, various lines of evidence point to a universal, biochemical pathway. For example, the human anti-apoptotic protein Bcl-2 (to be discussed in a later section) can partially restore a defective ced-9 function in C. elegans [6, 7]. Equally important, in most systems of apoptosis experimentally studied, clear evidence for involvement of caspases was found. Caspases are a recently discovered, extended family of Cysteine-type proteases, which cleave after an aspartic acid residue; the C. elegans CED-3 protein and interleukin-1β-converting enzyme (ICE) are typical examples. Strong evidence for the involvement of caspases is the fact that usually the process of apoptosis can be interfered with by specific caspase inhibitors, such as zVAD-fmk or viral antagonistic pseudo-substrates (see below). TNF and Fas/Apo-1 ligand play a prominent role in a variety of immunological, inflammatory and pathological conditions. Therefore, many studies on PCD have been done with TNF-R55 (CD120a) and Fas/Apo-1-induced death. Both receptors contain near their intracellular C-terminus a homologous region of about 90 amino acids, referred to as the ‘death domain’ (DD). Upon clustering, this DD is sufficient for signalling cell death. TNF-R55-DD in addition leads to activation of the transcription factor NF-κB and to gene induction. Many of the induced genes are important for inflammation (IL-6, IL-8, ICAM-1, E-selectin), while activation of NF-κB is also involved in synthesis of resistance proteins, such as A20 [8]. Recently, other receptors of the same family have been cloned and shown to contain a DD-related sequence, such as DR3/WSL-1/TRAMP [9-11], chicken CAR-1 [12]and DR4 [13]. The TNF receptor and ligand families are currently the object of intense research, covering a variety of different aspects of cellular signalling (reviewed in [14-20]). These ligand and receptor families, although large and continuing to increase in size, display rather homogeneous patterns of the structures involved in recognition. 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