A Brief Introduction to ApoptosisAnimal cells can self-destruct via an intrinsic program of cell death (Steller, 1995). Apoptosis is a form of programmed cell death that is characterized by specific morphologic and biochemical properties (Wyllie et al., 1980). Morphologically, apoptosis is characterized by a series of structural changes in dying cells: blebbing of the plasma membrane, condensation of the cytoplasm and nucleus, and cellular fragmentation into membrane apoptotic bodies (Steller, 1995; Wyllie et al., 1980).
Biochemically, apoptosis is characterized by the degradation of chromatin, initially into large fragments of 50-300 kilobases and subsequently into smaller fragments that are monomers and multimers of 200 bases (Oberhammer et al., 1993; Wyllie, 1980). Other biochemical indicators of apoptosis are induced or increased levels of the protein clusterin (Pearse et al., 1992), also known as TRPM-2 or SGP-2, and activation of the enzyme typeII transglutaminase, which crosslinks proteins to the envelope of apoptotic bodies (Fesus et al., 1991). Apoptosis is a complex phenomenon of related morphological and biochemical processes that can vary with tissue and cell type (Zakeri et al., 1995).
The execution of apoptosis minimizes the leakage of cellular constituents from dying cells. For example, proteases could damage adjacent cells or stimulate an inflammatory response. This cardinal feature of apoptosis distinguishes it from necrosis, which usually results from trauma that causes injured cells to swell and lyse, releasing the cytoplasmic material that stimulates an inflammatory response (Steller, 1995; Wyllie et al., 1980).
Apoptosis research has been spurred by the observation that normal metazoan development and health require the precise regulation of cell death. Apoptosis plays a critical role in important biological processes such as morphogenesis, tissue homeostasis, the elimination of damaged or virally infected cells, and elimination of self-reactive clones from the immune system (Steller, 1995). Although apoptosis is important for the normal development and health of an animal, its aberrant activation may contribute to a number of diseases, for example, AIDS, neurogenerative disorders, and ischemic injury (Thompson, 1995). In contrast, impaired apoptosis may be a significant factor in the etiology of such diseases as cancer, autoimmune disorders, and viral infections (Thompson, 1995).
Some of the molecular components of the apoptotic program have been conserved through evolution (Steller, 1995). Genetic studies in C. elegans have identified mutations in 14 genes that affect programmed cell death in this organism (Steller, 1995). Two of these genes, ced-9 and ced-3 (ced stands for cell death defective), were homologous to mammalian genes: the proto-oncogene bcl-2 and ice (interleukin-1-ß-converting enzyme), respectively. Recent work has demonstrated the importance of these genes in the regulation and execution of apoptosis.
Ecotopic expression of ced-9/bcl-2 blocks apoptosis in many experimental systems (Reed, 1994). The protein Bcl-2 appears localized or associated with intracellular membranes of mitochondria, endoplasmic reticulum, and nuclei (Reed, 1994). Although the mechanism of Bcl-2 action is unknown, biochemical studies have implicated this protein in the regulation of the redox potential of the cell, as ecotopic expression of bcl-2 suppresses cell death induced by oxidizing agents and appears to affect glutathione levels (Korsmeyer et al., 1993; Zhong et al., 1993). Genetic evidence indicates that ced-9/bcl-2 belongs to an emerging family. Some of the members of this family can suppress apoptosis like bcl-2, for example, bcl-xL (Boise et al., 1993), while other members make cells more susceptible to apoptotic stimuli, for example bax and bcl-xS (Boise et al., 1993; Oltvai et al., 1993). In addition, bcl-2 family members can form hetero- and homo-dimers, suggesting a model of regulation where the cellular susceptibility or threshold for apoptosis is partly influenced by the repertoire and level of expression (Oltvai et al., 1993).
ced-3/ice encodes a cysteine protease that cleaves peptide bonds after Asp residues (Martin and Green, 1995). Mutations that inactivate ced-3 block apoptosis in C. elegans, while ectopic/over expression of ice causes apoptosis in rat fibroblasts (Yuan et al., 1993; Miura et al., 1993). Studies from several laboratories have identified ced-3/ice gene family proteases (Ich-1/Nedd-2, cpp32ß/yama, Tx/Ich-2, and Mch-2), all of which appear to cause apoptosis when overexpressed in various cell types (Martin and Green, 1995). Additional evidence supporting a role for these proteases in apoptosis comes from studies showing that Ice inhibitors such as the cowpox viral protein CrmA and the peptide YVAD block apoptosis induced by the Fas ligand or Fas receptor cross-linking (Martin and Green, 1995). Among the potential substrates for these proteases are other members of this gene family, poly(ADP-ribose) polymerase (PARP), and lamins (Martin and Green, 1995).
Recently, considerable research has focused on identifying the molecular sensors or triggers of apoptosis. Perhaps the best studied mammalian proteins that fit the classification of sensors or triggers are the tumor suppressor p53 and the Fas ligand and its cognate receptor Fas, respectively (Donehower and Bradley, 1993; Nagata and Golstein, 1995). p53 is a DNA binding protein and transcriptional activator that may have a role in DNA repair, because it appears to accumulate following DNA damage (Donehower and Bradley, 1993). Studies using p53 minus cell lines transfected with a temperature sensitive mutant of p53 have clearly shown that wildtype p53 function lowers the threshold for inducing apoptosis following genotoxic damage (Lowe et al., 1993). Thus, p53 may function as a sensor of DNA damage and has been called the "guardian" of the genome (Lane, 1992).
In contrast to p53, the Fas ligand/receptor system is an example of a trigger of apoptosis. The Fas ligand, is a cell surface protein that is found predominantly on activated T cells and is a member of the tumor necrosis factor gene family (Nagata and Golstein, 1995). Fas ligand triggers apoptosis in a variety of cells bearing Fas on their cell surface (Nagata and Golstein, 1995). Antibodies against Fas also can trigger apoptosis, presumably by cross-linking Fas into larger complexes (Nagata and Golstein, 1995). Mutational studies of Fas have revealed a cytoplasmic domain of about 70 amino acids that is required for transduction of the apoptotic signal and has been dubbed the death domain (Itoh and Nagata, 1993; Tartaglia et al., 1993). Recent work on the mechanism of Fas signalling suggests that production of ceramide via activation of an acid sphingomyelinase may be important. The addition of synthetic C2-ceramide alone to culture medium can mimic Fas activation and trigger apoptosis (Cifone et al., 1994).
In summary, a major goal of apoptosis research is to identify its molecular components and mechanisms of regulation. This information may lead to therapeutic agents that can modulate this process in the treatment of degenerative diseases such as neurodegenerative disorders or proliferative diseases such as cancer.
Return to Apoptosis in Development
This review is based upon a brochure prepared and distributed by Upstate Biotechnology, Inc. Reproduced by permission.
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