DNA Damage and Repair in Central Nervous System Injury
National Institute of Neurological Disorders and Stroke Workshop Summary
Background and Purpose DNA damage and repair are areas of research with important implications for stroke and cerebral trauma. DNA damage is present in central nervous system (CNS) injury, and defects in repair mechanisms are associated with neurodegenerative disease.
Methods A workshop, DNA Damage and Repair in CNS Injury, was organized by the National Institute of Neurological Disorders and Stroke in Bethesda, Md, on September 11, 1995. The objective of this workshop was to promote inquiry and to foster application of research in DNA damage and repair after stroke and trauma.
Results The participants discussed the connection between the fields of DNA damage and repair and stroke and trauma and identified gaps in knowledge to be filled to expand research of DNA damage and repair in CNS injury. Specific recommendations were made targeting research opportunities in the areas of DNA repair and damage in stroke and trauma.
Conclusions Research in the science of DNA injury and repair will likely provide new and important information on mechanisms of cell damage and provide opportunities for the development of novel and effective therapies to reduce CNS injury in stroke and trauma.
There is evidence of DNA damage in experimental stroke and trauma, and neurological syndromes have been attributed to impairment in DNA repair mechanisms. Thus, DNA damage is a likely factor in promoting CNS pathophysiology, and DNA repair may be an important mechanism for maintenance of normal physiological function. The purpose of the workshop was to bring scientists and clinicians with expertise in DNA injury and repair together with investigators in stroke and CNS trauma to promote inquiry and foster application of this new area of science to stroke and head injury research. The workshop focused on four topics: overview of DNA injury and repair and CNS injury; PCD and DNA damage; mechanisms of DNA damage and repair; and mechanisms of DNA damage and repair in CNS injury. To promote the growth and application of research in DNA damage and repair in stroke and brain injury, recommendations targeting specific research opportunities were made. The consensus of the participants was that the application of the science of DNA injury and repair may yield new and important information on mechanisms of neuronal damage and provide opportunities for the development of novel and effective therapies to reduce CNS injury in stroke and trauma.
The terms “apoptosis,” “DNA fragmentation,” and “programmed cell death” describe related but not necessarily congruent events. Apoptosis and necrosis are descriptive terms for types of cell death exhibiting distinct sets of morphological features. Morphological features of apoptosis include condensed chromatin, plasma membrane blebs, and shrunken cytoplasm in which the individual organelles remain intact. The morphological consequences of necrosis differ from apoptosis, and necrotic cells typically swell and lyse, release intracellular contents, and provoke an inflammatory response. Nuclear DNA is not systematically degraded in necrotic cells. DNA fragmentation is a biochemical term that describes endonuclease cleavage of DNA into segments of approximately 200 bp in length. DNA fragmentation is closely associated with apoptosis. PCD is a functional term describing an active process of cell death that generally requires the activation of a genetic program. These terms have been used interchangeably because the consequences of PCD are apoptosis and DNA fragmentation. However, there are cellular systems that may exhibit one or more of these characteristics independently.
Overview—DNA Damage and Repair and CNS Injury
An overview of the mechanisms of DNA damage1 2 3 and experimental evidence for DNA-related damage from studies of apoptosis in neuronal cultures and experimental stroke models were presented.4 5 6 There are two types of structural abnormalities that lead to mutations or permanent changes in DNA sequences. The first type involves normal bases that are positioned in a wrong sequence context (eg, mismatched, bulged, or looped), and the second type involves abnormal nucleotides (eg, modified, fragmented, or cross-linked) that are in normal sequence context. The biochemical reactions that attempt to rectify these abnormalities are generally referred to as the DNA repair mechanisms. In principle, the repair mechanisms that are responsible for repair of mismatches are similar: the incorrect or damaged base is removed individually (base excision) or is removed as an oligonucleotide (nucleotide excision). In the latter case, the single-stranded gap is filled in by the action of polymerase (repair synthesis), and the newly synthesized DNA is ligated. Thus, the DNA damage/repair process can be monitored by either an incision/excision assay or a repair synthesis assay. A listing of different DNA lesions repaired by excision repair is presented in the Table⇓. The Figure⇓ illustrates mechanisms of base-excision repair and nucleotide-excision repair.
At least nine genes have been identified in human diseases to be associated with excision repair, eg, XP and trichothiodystrophy; many other genes from rodents may indeed have known human homologues. Although much has been discovered on the excision-repair reactions, little is known about repair synthesis. At least two polymerases and the proliferating cell nuclear antigen gene are involved. A debate exists concerning the potential connection between tumor suppressor genes (such as p53) and DNA repair and cell cycle elements such as CDK7 and cyclin H (which constitute the CDK-activating kinase) that may tie up DNA repair and cell replication.
The basis of current knowledge and the concept of DNA damage and repair are largely derived from nonneuronal cells and/or tissues and nonmammalian research. Therefore, a significant gap of knowledge exists concerning the fundamental biochemical processes that may lead to DNA damage in the brain, as well as the physiological relevance of such processes. In addition, the genetic/biochemical repair systems that exist in neurons, glia, and other brain cells must also be studied.
Two forms of neuronal damage in stroke and CNS trauma, necrosis and apoptosis, have gained recent attention. Both forms may be activated independently but may lead into the other mode of the death pathway, independent of the inciting insult. Mechanisms that lead to neuronal death may involve de novo protein synthesis, and genes associated with proapoptotic and antiapoptotic programs may be activated. Both in vitro and in vivo studies support the hypothesis that necrosis and apoptosis are multimodal pathways toward death. In in vitro studies, neuronal death induced by oxygen-glucose deprivation was associated with acute cell-body swelling without internucleosomal DNA fragmentation; it could be attenuated by addition of the glutamate receptor antagonists MK-801 plus CNQX to the exposure medium but not by addition of the protein synthesis inhibitor cycloheximide. However, if the duration of oxygen-glucose deprivation was extended to overcome the protective effect of MK-801 plus CNQX, a neuronal death resulted that was associated with cell-body shrinkage and DNA laddering. This presumptive apoptotic death could be attenuated by cycloheximide. The idea that oxygen-glucose deprivation can induce in cortical neurons both excitotoxic necrosis and apoptosis dependent on new macromolecule synthesis has been further supported by two sets of recent in vivo experiments utilizing a rat model of transient middle cerebral artery occlusion. In the first experiments, mild transient ischemia (insufficient to induce infarction at 24 hours) was followed by the appearance of substantial infarction between 3 and 14 days. This very delayed infarction was preceded by neuronal thymidine/deoxythymidine-mediated deoxyUTP-biotin nick end labeling (TUNEL) staining and tissue DNA laddering, and that was largely blocked by a single pretreatment injection of cycloheximide. In the second experiments with severe transient ischemia, combined pretreatment with a glutamate receptor antagonist, dextrorphan, and cycloheximide reduced heat shock protein HSP70 expression and infarct volume to a greater extent than either drug alone. Thus, apoptosis is viewed to be a major pathway in neuronal response to injury and must be given proper attention in studies of neuronal vulnerability and resistance to injuries.
PCD and DNA Damage
The discussions on this topic focused on mechanisms of PCD and the molecular and genetic cascades that initiate and propagate suicide programs in cells. The importance of ICE proteins, p53, the molecular signaling mechanisms in PCD, and the role of Bcl-2 proteins in PCD were discussed.
The genetic control of cellular suicide in neurons is represented by PCD in the nematode Caenorhabditis elegans.7 8 The CED-3 gene (cell death-3) activates, whereas the CED-9 gene blocks, PCD. Hence, the most favorable situation for cell death involves inhibition of death suppression and activation of death-promoting programs. Expression of a CED-3 homologue occurs in mammalian cells that is now the prototype for a family of mammalian proteins exhibiting ICE activity. The ICE family consists of at least six distinct enzymes that both positively and negatively regulate PCD in vertebrate animals. The ICE family members share cysteine protease activity, exhibit strong sequence homology to CED-3, and show specific substrate cleavage requirements. ICE, the first family member identified, shows 28% overall sequence homology and complete conservation of the CED-3 active site. ICE itself must undergo cleavage for activity. The active enzyme processes pro-interleukin 1β, a proinflammatory cytokine associated with inflammation and cell death in some systems. The substrates for other ICE family members are not known, although there are a number of candidate proteins.
Another recently discovered ICE-like cysteine protease, CPP-322B, may also be important in mammalian systems. Like ICE, CPP-322B activity is critically dependent on proteolytic cleavage of an inactive precursor, is highly homologous to CED-3, and cleaves the nuclear enzyme PARP, an early biochemical event in PCD. CPP-322B cleavage is not autocatalytic, raising the intriguing possibility that one or more ICE family members may promote cleavage and activation of CPP-322B.
There is functional evidence implicating ICE in mammalian PCD. Ectopic expression of ICE induces apoptosis in transfected cells by a mechanism reversed by coexpression of an ICE inhibitor, crm, a pox virus, or coexpression with Bcl-2. In addition, ICE and CPP-322B cleave and thereby inactivate PARP to block DNA repair without diminishing auto-ADP ribosylation. Despite strong evidence for CED-3/ICE participation in PCD, ICE may not be a universal trigger; ICE-knockout mice show normal embryological development. Similarly, PARP is probably not the sole effector for ICE or CPP-322B activity: PARP-knockout mice, rather than exhibiting gross developmental abnormalities, exhibit minor pathological features such as epidermal hyperplasia. The results in knockout mice underscore the need to define the precise chain of molecular events in PCD.
The nuclear phosphoprotein p53 has a role in DNA injury and repair.9 10 Originally identified as a tumor suppressor, p53 is a DNA-binding transcription factor that is involved in the control of cell proliferation, DNA repair, and apoptosis. Normally, p53 is believed to inhibit growth by halting the cell cycle and thereby providing the time needed to repair damaged DNA before division. Mutations in the p53 gene, which occur frequently, have been linked with diverse types of human cancer. The level of p53 is increased after treatment of cells with agents that cause DNA damage. It has been suggested that p53 binds to and alters the activity of several factors that are involved in the repair of damaged DNA; however, there is presently no consensus about the precise function of p53 in DNA repair. When extensive DNA damage occurs, p53 is believed to be involved in the process of apoptotic cell death.
There have been only a few studies of the relationship between p53 and brain injury. In models of focal cerebral ischemia, expression of p53 increases in regions undergoing ischemic injury and is associated with DNA fragmentation. In a model of excitotoxic injury, expression of p53 occurs in neurons that exhibit morphological features of injury and DNA fragmentation. Although these results demonstrate that p53 is a marker of neuronal injury, they do not indicate whether p53 is involved in DNA repair or apoptosis. Indeed, the wide range of cellular functions of p53 only increases the difficulty of interpreting changes in p53 expression.
Wild-type and mutant p53 can bind in vitro human TCR factors involved in strand-specific DNA repair. All these factors belong to a recently identified DNA and RNA helicase superfamily. Investigators have demonstrated differences in the transcriptional activity of wild-type and mutant p53. Cells from patients with Li-Fraumeni syndrome are heterozygous for the p53 mutant allele and repair UV-induced pyrimidine dimers at a slower rate than normal human cells. These and other results indicate that p53 may play both an indirect (G1 checkpoint function) and a direct role in modulating nucleotide-excision repair pathways. Although p53 has the potential to modulate important processes of DNA damage, DNA repair, and PCD, the evidence for such a role in brain injury remains circumstantial.
The triggering mechanisms for PCD involve novel receptor-ligand binding sites both at the cell surface and within cells.11 12 13 Investigators have developed a monoclonal antibody that initiates apoptosis selectively in cultured neurons. This antibody recognizes a glycosphingolipid surface binding site selectively expressed on neurons and on no other tested cells such as glia. Death domains, as these activation sites are known, may also reside on the intracytoplasmic segments of receptor proteins, such as the low-affinity neurotrophin receptor p75NTR. Expression of p75NTR by neurons lowers the apoptosis threshold in the absence of neurotrophins. High sequence homology between p75NTR domain and the death domain of tumor necrosis factor suggests the possibility that common signaling mechanisms may initiate and propagate PCD. Precisely how this is accomplished demands further study, but common PCD mechanisms offer novel therapeutic opportunities in CNS disease.
There appears to be a relationship between cell susceptibility and oxidative stress in PCD. Overexpression of copper/zinc superoxide dismutase inhibits neuronal apoptosis induced by growth factor withdrawal. The production of reactive oxygen species may stimulate an early proapoptotic cell signal rather than activate more traditional pathways that promote toxicity to lipids, proteins, and DNA. Transient ischemia, exposure to β-amyloid, and high oxygen tension are conditions that both promote apoptosis and stimulate the production of reactive oxygen species.
Bcl-2 decreases the net generation of free radicals, prevents lipid peroxidation, and through unknown mechanisms augments the more reduced state of NADH and GSH over NAD and GS to render cells relatively more resistant to oxidative stress. Perhaps these findings underlie recent observations that resistance to ischemic brain injury develops when Bcl-2 is overexpressed in transgenic mice and after brain cells of normal mice are stimulated to express Bcl-2.
Consideration of injury repair mechanisms in the brain, however, raises issues concerning the antiapoptotic Bcl-2 gene family and protein-protein interactions. Overexpressed in some human cancers, Bcl-2 confers resistance to chemotherapy. p53 gene expression transcriptionally represses Bcl-2 gene expression, whereas loss of p53 augments Bcl-2 production. At least three peptide domains are required for the death suppressor function. Targeting these peptide domains could become an important focus for drug development, although protein-protein interactions are notoriously difficult to target therapeutically. Regulation of PCD by the Bcl-2 family members appears to be dependent on a complex balance between cell death–enhancing and cell death–suppressing homologues. Other members of the Bcl-2 family include BAX, BIP, and BAK. BAX and BIP are proapoptotic, whereas BAK and Bcl-2 enhance cell survival. Homodimer and heterodimer formation are important to this process. For example, BAX/Bcl-2 binding confers antiapoptotic activity, whereas Bcl-2/Bcl-2 homodimers are inactive.
Mechanisms of DNA Damage and Repair
There is evidence that neuronal damage and neurological deficits in neurodegenerative diseases are caused by inherited deficits in DNA repair mechanisms. Other studies have indicated that oxidative DNA damage and repair can importantly modulate pathological responses of CNS neurons, including apoptotic cell death. Finally, since the introduction of alkaline sucrose gradient sedimentation methods, there have been considerable advances in techniques used to detect single-strand breaks that may be applicable to in vivo studies.
The neuronal cell death seen in XP most likely results from unrepaired DNA damage producing neuronal cell death.14 15 16 All neurologically affected XP patients appear unable to repair a type of free radical–induced DNA damage requiring DNA nucleotide-excision repair in expressed genes. These patients eventually develop similar neurological complications, and the more severe the nucleotide-excision repair defect the sooner complications arise. These important studies demonstrate that DNA repair of neurons is required to maintain the human CNS. It is possible that neuronal death due to inability to repair DNA damage by free radicals could also contribute to other hereditary neurodegenerative diseases, including Cockayne’s syndrome and Alzheimer’s disease. These observations raise important questions of how investigators will establish causal relationships between DNA damage and specific pathological features of stroke and trauma. Although the mechanisms underlying DNA damage and repair, as well as the methods used to study these mechanisms, differ from those in earlier studies of pathological processes underlying stroke and trauma, the general research strategies should be similar. For example, investigators have enjoyed considerable success in providing evidence that excitotoxic mechanisms contribute to pathology after stroke and trauma. These studies have used techniques including blockade of specific receptors that gate calcium entry after CNS injury. Thus, it would be important to explore how we can manipulate mechanisms of DNA damage and repair and how these changes affect specific end points in stroke and trauma model systems.
Oxidative mechanisms can mediate DNA damage to cultured cortical neurons, and this DNA damage is associated with nuclear fragmentation and DNA laddering typical of apoptosis in response to irradiation.17 18 Studies carried out to investigate the response of neurons to radiation-induced DNA double-strand breaks in comparison with other normal brain cells capable of division (such as astrocytes) indicate that neurons do repair double-strand breaks, but they do so significantly more slowly than astrocytes. Many neurons underwent nuclear fragmentation and DNA laddering typical of apoptosis in response to irradiation. In contrast, no DNA laddering was seen in astrocytes. The protein synthesis inhibitor cycloheximide and the RNA transcription inhibitor actinomycin D prevented the radiation-induced apoptosis of neurons. To test whether the decreased DNA repair capabilities of the neurons might be related to a greater level of differentiation, undifferentiated astrocytes were treated with dibutyryl cAMP to induce differentiation. Treatment with cAMP enhanced rejoining of DNA double-strand breaks. Furthermore, treatment of neurons with cAMP significantly decreased the level of radiation-induced DNA apoptosis. These results are consistent with those of previous studies indicating that increased oxidative stress can lead to neuronal apoptosis and that certain neuronal populations may be particularly susceptible to apoptosis due to the slow rate of DNA repair in these cells. In addition, modifiers of DNA repair may be useful for blunting apoptosis, an important issue for investigators of acute CNS injury. Although oxidative damage is implicated in stroke and traumatic brain injury, there is a need to link oxidative stress in stroke and trauma models to DNA damage and apoptosis.
Recent technical advances include methods for measuring single-strand breaks and increasing sensitivity, speed, and sample-handling capacity.19 20 Many of these methods used to detect single-strand breaks have been adapted for measurement of double-strand breaks, which are often considered more relevant for chromosome aberrations and cell death. A large variety of factors may contribute to the response of a cell to a DNA-damaging agent. In addition to investigating the complexity of factors mediating responses to DNA damage, it will be important to define points in pathological cascades resulting from DNA damage, including (1) trigger points initiating cascades, (2) points in the cascade at which reversible damage occurs, and (3) points in the cascade at which irreversible damage occurs.
DNA Damage and Repair Mechanisms in CNS Injury
In this session, the two major pathways for DNA excision and repair, a global pathway and a TCR pathway (which only affects transcribed DNA) were discussed. Preliminary results showing an association between neurological disorders and genomic instability were provided.
Recent studies have identified a subpathway of nucleotide-excision repair in which transcriptionally active genes are more rapidly cleared of certain types of DNA lesions than are silent domains of the genome.21 22 23 This TCR operated on transcribed DNA strands within transcription units. Only RNA polymerase II–transcribed genes are subject to TCR, and the process requires that the polymerase is actively translocating on the DNA template.
Several genes implicated in Cockayne’s syndrome are required for TCR but not for global genomic repair. A defect in TCR in early development might account for the neurological problems in Cockayne’s syndrome patients. The demonstration of TCR but not global repair of UV-induced cyclobutane pyrimidine dimer in the XP129 partial revertant of XP group A cells indicates that UV resistance correlates with repair of cyclobutane pyrimidine dimer in active genes. Repair measured as an average over the genome can be misleading, and the results underscore the necessity of considering the genomic locations of DNA lesions and their repair for an accurate assessment of the biological importance of these lesions.
p53 mutants appear to be a double-edged sword. Li-Fraumeni syndrome fibroblasts expressing only the mutant p53-tumor suppressor gene are more UV resistant and exhibit less UV-induced apoptosis than normal human cells or heterozygotes with mutations in only one allele of p53. The p53-defective cells exhibit a marked reduction in global-excision repair capacity, but they retain normal TCR. The loss of p53 function may enhance genomic instability by reducing the efficiency of global DNA repair, while cellular resistance may be assured through the operation of TCR and the elimination of apoptosis. This system constitutes yet another example of a situation in which cell survival does not correlate with the efficiency of global DNA repair.
Rat PC12 cells have been used as a model system to study TCR in differentiating neuronlike cells. Global repair of cyclobutane pyrimidine dimer dropped from 15% to an undetectable level in the terminally differentiated cultures. Repair in the induced GAP43 gene was upregulated, while repair in the constitutively expressed synapsin I gene was unaffected by differentiation. These results provide evidence that postmitotic neuronal cells remove cyclobutane pyrimidine dimer from their DNA, and they confirm the general expectation that repair is more efficient in expressed genes than in silent genomic domains. TCR may be a key mechanism for assuring that transcriptionally active genes injured by oxidative stress are rapidly repaired.
Preliminary data supports an association between neurological disorders and genomic instability.24 25 Interactions between hydroxyl radicals and DNA increased the content of several DNA adducts. Mutagenic 8-hydroxyguanine is one of them. A positive correlation exists between the increase in 8-hydroxyguanine content and base-pair deletion in mitochondrial DNA from the cortex and putamen of human brain. An accumulation of DNA damage and somatic mutation in the brain is part of the normal aging process. Unique point mutations have been found in mitochondrial DNA from some patients with certain neurodegenerative diseases such as Alzheimer’s and Parkinson’s diseases. In addition, metal-catalyzed oxidation is thought to contribute to the amyloidogenicity of β-amyloid. Associations of clinical neurological disorders and defective DNA repair have been documented in certain familial forms of human disorders. The association between premature neurodegeneration with traumatic head injuries in early life raises the possibility that reactive oxygen species generated after head injury may damage DNA and accelerate the aging process in humans. Gene instability may be associated with accelerated neurodegeneration and aging.
Preliminary studies to examine the pathogenetic role of reactive oxygen species in DNA damage and repair after experimental cerebral ischemia were described. A significant increase in 8-hydroxyguanine content was observed in C57BL/6 mice subjected to transient forebrain cerebral ischemia. Ischemia and reperfusion induced approximately six G:C damages per gene per cell for every G:C damage in the normal brain. The brain cells responded to ischemia/reperfusion with DNA repair, as shown by a reduction in some but not all DNA damages and by an increase in the expression of DNA polymerase-B gene. Furthermore, the incidence of chromosomal gene mutation increased significantly, suggesting that DNA repair was not without error. Finally, DNA fragmentation appeared in certain nuclei and gradually increased with time 3 to 8 days after cerebral ischemia. These data suggest that the brain can remove DNA damage after ischemia. On the other hand, a delay in the removal of some mutagenic DNA damage during reperfusion and an error-prone repair is likely to be the culprit in causing permanent damage. These findings offer preliminary observations that support the contention that DNA damage and defective repair may initiate genome instability, which causes cell death through apoptotic mechanisms. The results offer new insights into our understanding of stroke mechanisms and into potential new treatment strategies.
Conclusions and Recommendations
On the basis of the presentations and discussions at the workshop, the following conclusions were derived and recommendations were made concerning the application of DNA damage and repair in stroke and traumatic brain injury. (1) DNA damage and repair modulate injury to the CNS. However, the causal relationships between DNA damage and pathological responses to stroke and trauma have not been clarified. (2) Techniques to measure DNA damage are available that are applicable to in vitro and in vivo models of stroke and traumatic brain injury. (3) Two major pathways exist for DNA excision and repair: a global pathway and TCR. (4) ICE, Bcl-2, p53, and DNA polymerase-B genes are implicated in apoptosis and in DNA damage and repair. (5) Knowledge of DNA damage and repair mechanism has been largely derived from nonneuronal tissue, and there is a need to investigate these mechanisms in neurons.
The following proposed research directions were recommended: (1) Define points in pathological cascades resulting from DNA damage ie, trigger points and points at which reversible and irreversible damage occurs; determine the precise chain of molecular and genetic events in neuronal PCD. (2) Identify genomic locations of DNA damage and repair in neurons for an accurate assessment of the biological importance of these lesions; determine whether DNA damage occurs at random locations or more frequently at predictable sequences or locations; and assess the differences between potentially deficient TCR and global repair mechanisms in selective regions after CNS injury. (3) Identify genes, enzymes, and receptors associated with DNA repair in neurons. (4) Identify the contribution of extracellular (eg, N-methyl-d-aspartate, nitric oxide, pH, Ca2+) and intracellular (eg, mitochondrial dysfunction, deacylation of phospholipids) mechanisms in DNA damage and apoptosis; link oxidative stress to pathological end points related to DNA damage and apoptosis. (5) Investigate the role of DNA injury and repair as mechanisms of selective vulnerability, penumbra, preconditioning, delayed cell death, and reperfusion injury. (6) Examine the role of the DNA repair enzymes in cerebral injury from stroke and trauma using transgenic animals with various defects of nucleotide excision repair. (7) Investigate the role of DNA repair mechanisms and responses of the cerebral blood vessel wall cells after ischemia and trauma. Vascular responses are virtually unexplored and could provide insight into the role of angiogenesis in cell and tissue survival. (8) Identify methods to measure DNA damage noninvasively (eg, single-photon emission CT, MRI).
Selected Abbreviations and Acronyms
|CNS||=||central nervous system|
|PCD||=||programmed cell death|
Apoptosis A descriptive term for a type of cell death exhibiting a distinct set of morphological features consistent with condensed chromatin, plasma membrane blebs, and shrunken cytoplasm in which the individual organelles remain intact. Apoptotic cells frequently cleave their DNA between nucleosomes.
Bc1-2 gene family A family of genes that promote cell survival (BAK, Bcl-2) or promote apoptosis (BAX, BIP).
CED genes/cell death genes A family of genes that regulate (promote or inhibit) programmed cell death in the nematode C elegans.
Cyclin Proteins whose concentrations rise and fall during the course of eukaryotic cell cycle. Cyclins form complexes with cylin-dependent protein kinases and thereby activate and determine substrate specificity of these enzymes, which control passage through the cell cycle.
DNA fragmentation Cleavage of DNA at nucleosomes, associated with apoptosis and programmed cell death.
DNA and RNA helicase superfamily A class of enzymes that move along a DNA duplex utilizing the energy of ATP hydrolysis to separate strands.
Endonuclease Bacterial enzymes that recognize specific base-pair sequences and then cleave both DNA strands at this site.
Exonuclease Bacterial enzymes that digest nucleic acids from an end.
Excision repair A DNA repair mechanism that involves the following steps: recognition of damage, incision near the damage, removal of damaged stretch, polymerase filling of the gap, and ligation of the two ends.
Interleukin-converting enzyme A family of proteins that regulates programmed cell death.
p53 A DNA-binding transcription factor involved in control of cell proliferation, DNA repair, and apoptosis.
p75NTR A neurotrophin receptor that lowers the apoptosis threshold in the absence of neurotrophins.
Point mutations Substitution of one base pair for another (eg, base substitution, mutations), change of one purine or pyrimidine for another (transition mutations).
Poly(ADP-ribose) polymerase An enzyme that influences the efficiency of DNA ligation during base-excision repair but does not have any effect of nucleotide-excision repair.
Programmed cell death A directed form of cell death, mediated by genes or proteins. These activated cells often exhibit apoptotic morphology.
Proliferating cell nuclear antigen A necessary component of the machinery that copies DNA so that cell division can occur. It is also needed for resynthesis of DNA after damaged portions are removed.
Transcription-coupled repair pathway Transcriptionally active genes are more rapidly cleared of certain types of DNA lesions than are silent domains of the genome.
TUNEL staining Thymidine/deoxythymidine-mediated deoxyUTP-biotin nick end labeling; histochemical method used to identify apoptotic cells.
We gratefully acknowledge the assistance of the participants and discussants who contributed to this article: Dale Bredesen (La Jolla, Calif), Pak H. Chan (San Francisco, Calif), Mary Ellen Cheung (Bethesda, Md), Dennis W. Choi (St Louis, Mo), Michael Chopp (Detroit, Mich), Giora Feuerstein (King of Prussia, Pa), Myron Ginsberg (Miami, Fla), Glenn T. Gobbel (San Francisco, Calif), Zach W. Hall (Bethesda, Md), John M. Hallenbeck (Bethesda, Md), Philip C. Hanawalt (Stanford, Calif), Curtis Harris (Bethesda, Md), Ronald L. Hayes (Houston, Tex), Chung Y. Hsu (St Louis, Mo), Paul L. Huang (Charlestown, Mass), Thomas P. Jacobs (Bethesda, Md), Matthew Linnik (Cincinnati, Ohio), Philip Liu (Houston, Tex), Tracy K. McIntosh (Philadelphia, Pa), Michael A. Moskowitz (Boston, Mass), Peggy Olive (Vancouver, BC, Canada), Jay H. Robbins (Bethesda, Md), Aziz Sancar (Chapel Hill, NC), Frank Sharp (San Francisco, Calif), Michael D. Walker (Bethesda, Md), Frank A. Welsh (Philadelphia, Pa), and Junying Yuan (Charlestown, Mass).
- Received December 15, 1995.
- Revision received January 15, 1996.
- Accepted January 15, 1996.
- Copyright © 1996 by American Heart Association
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