Why Do Neurons Enter the Cell Cycle?

[Cell Cycle 3:6, 769-773; June 2004]; ©2004 Landes Bioscience

Why Do Neurons Enter the Cell Cycle? Review

KEY WORDS DNA damage, cell cycle, DNA repair, apoptosis

ABBREVIATIONS

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reactive oxygen species base excision repair transcription-coupled repair global genome repair ataxia telangiectasia mutated ATM and Rad 3-related cyclin-dependent kinase retinoblastoma protein murine double minute-2 checkpoint kinase 2 Bcl-2 associated X protein apoptotic protease-inducing factor 1 Alzheimer’s disease amyotrophic lateral sclerosis 8-hydroxy-2'-deoxyguanosine harlequin mutation

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INTRODUCTION

DNA lesions are constantly produced in living cells by the deleterious actions of both endogenous and environmental DNA-damaging agents. More than 104 DNA damaging events occur in each mammalian cell every day from spontaneous decay, replication errors and cellular metabolism alone.1 The majority of DNA damage has an endogenous origin and is mainly produced by oxidation of DNA by reactive oxygen species (ROS), which are generated in the cells by normal aerobic metabolism. Oxidative stress causes base damage, as well as strand breaks in DNA. Such lesions, if left either unrepaired or misrepaired, interfere with essential DNA-dependent processes (transcription, replication, recombination, and chromosome segregation) leading to mutations, chromosomal instability and cell death, thereby resulting in cancer, and other pathological consequences.2,3 Thus, cells are under the constant threat of DNA damaging agents.

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ROS BER TCR GGR ATM ATR CDK pRb MDM2 Chk2 Bax Apaf-1 AD ALS oxo8dG Hq

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Previously published online as a Cell Cycle E-publication: http://www.landesbioscience.com/journals/cc/abstract.php?id=901

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Received 04/02/04; Accepted 04/02/04

Increasing evidence indicates that postmitotic, terminally differentiated neurons activate the cell cycle before death. The purpose of this cell cycle activation, however, remains elusive. In proliferating cells, cell cycle machinery is a major contributor to the DNA damage response, which is comprised of growth arrest. In quiescent cells such as terminally differentiated neurons, cell cycle-associated events may also be part of the DNA damage response. A link between DNA damage and repair, cell cycle regulation and cell death is becoming increasingly recognized for cycling cells but remains elusive for quiescent cells. Neurons are particularly susceptible to oxidative stress due to the high rate of oxidative metabolism in the brain and the low level of antioxidant enzymes compared to other somatic tissues. This is supported by fact that the intracellular end point of many neurotoxic stimuli is oxidative stress, which also represents a major cause of the neuropathology underlying a variety of neurodegenerative diseases. DNA is perhaps the major target of oxyradicals. Thus, oxidative stress may cause DNA damage, which is countered by a complex defense mechanism, the DNA damage response, which involves not only the elimination of DNA damage, but its coordination with other cellular processes such as cell cycle progression, together directing to preserve genomic integrity. The function of such response is the removal of DNA damage by DNA repair pathways, or the elimination of damaged cells via apoptosis. The present review discusses the idea that the cell cycle machinery is a critical element of the DNA damage response not only in cycling, but also quiescent cells, and may bear the same function: to repair the damage or initiate apoptosis if the damage is too extensive to be repaired.

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Correspondence to: Inna Kruman, Ph.D.; Sun Health Research Institute; 1015 West Santa Fe Drive; Sun City, Arizona 85351 USA; Tel.: 623.876.5607; Fax: 623.876.5695; Email: [email protected]

ABSTRACT

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Inna I. Kruman

DNA DAMAGE RESPONSE IN CYCLING CELLS

To cope with the deleterious consequences of DNA lesions, cells are equipped with efficient defense mechanisms to remove DNA damage by DNA repair pathways, control cell cycle progression and eliminate damaged cells via apoptosis.2,4 The complicated network of DNA repair mechanisms includes base excision repair (BER), transcription-coupled repair (TCR), global genome repair (GGR), mismatch repair (MMR), homologous recombination (HR) and nonhomologous end-joining (NHEJ) damage. Evolution has overlaid the core cell cycle machinery with a series of surveillance pathways termed cell cycle checkpoints. In proliferating cells, checkpoints tightly control progress through the cell cycle. Cells may be arrested at any of the checkpoints, and the DNA will be repaired, or cells may die by apoptosis.5 The latter is a mechanism to ensure that irrepairable DNA modifications are not passed on to the progeny of a damaged cell. Thus, the overall function www.landesbioscience.com

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WHY DO NEURONS ENTER THE CELL CYCLE?

upstream of p53, phosphorylating checkpoint kinase 2 (Chk2).29 Chk2 phosphorylates p53, thereby stabilizing it.30 E2F-1-induced apoptosis may be p53-dependent.31,32 In addition, p53 might also directly induce cell death by stimulating the expression of pro-apoptotic genes including bax and apaf-1.33,34 p53 has also been shown to induce apoptosis by a transcriptionally independent mechanism involving direct activation of Bax.35

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DNA DAMAGE AND DNA REPAIR IN TERMINALLY DIFFERENTIATED NEURONS

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Functionally mature cells such as neurons are generally deficient in DNA repair. DNA repair is progressively attenuated in the course of cellular differentiation, leading to an increased vulnerability to DNA damaging conditions.36,37 In spite of the inability of postmitotic, terminally differentiated cells to replicate their genomic DNA, DNA repair is critical for the nervous system, as illustrated by the neurological abnormalities in patients with hereditary diseases associated with defects in DNA repair.38 The most devastating characteristic of these human genetic disorders is progressive neurodegeneration.28 Since neurons thought to be particularly susceptible to oxidative stress due to the high rate of oxidative metabolism in the brain and the low levels of antioxidant enzymes compared to other somatic tissues, oxidative stress in neurons may result in the accumulation of numerous oxidative DNA lesions and may thereby compromise transcription.39 The logical suggestion would be the existence of mechanisms maintaining the integrity of those genes needed for viable cell function. Terminally differentiated cells including neurons are indeed able to repair DNA damage, although the process is slower than in proliferative cells, and neurons are more prone to DNA damage-initiated apoptosis.40,41 Given the critical importance of preserving genomic integrity for living organisms, it is not surprising that the DNA damage response consists of machinery overlaid with multiple signal transduction pathways that expend considerable energy.2,5 Since DNA repair requires a large amount of ATP,42 which is overwhelming for such metabolically active cells like neurons, the evolutionary processes “minimized” the energy cost of DNA repair in differentiated cells at the global genome level. Global genome repair (GGR) machinery responsible for the removal of lesions at any location in the genome is deficient in neurons, compared to proliferating cells.43 It likely contributes to the higher sensitivity to DNA damage-initiated apoptosis seen in terminally differentiated cells. Efficient removal of lesions from transcribed strands of active genes is essential for proper cell function. Indeed, repair of active genes has been found to be maintained or even enhanced by differentiation employing a specific repair mode, referred to as transcription-coupled repair (TCR; see ref. 44). TCR specifically removes lesions from the transcribed strand.45,46 It is probably triggered by an RNA polymerase stalled on a lesion.47 TCR is a discrete pathway for initiating the rapid removal of lesions that block transcription.48 Defective TCR may lead to the arrest of RNA polymerase II translocation, p53 activation and apoptosis, especially if transcription blocking DNA damage occurs in genes encoding essential proteins.49 Together, these response pathways ensure efficient repair of the lesion, thereby maintaining the integrity of the transcribed strands in active genes of terminally differentiated cells or, if necessary, elimination of damaged cells. The efficiency of DNA repair by TCR and GGR in mammalian cells depends on a number of factors, including species, cell type and type of DNA lesion. TCR and GGR are thought to be activated via the common DNA damage response pathways but the exact molecular mechanism of TCR still remains obscure.48

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of cell cycle checkpoints is to detect damaged DNA, and to coordinate cell cycle progression with DNA repair.3,4 The process of DNA replication itself, in addition to standard DNA-damaging insults, adds intrinsic risks, such as base misincorporation errors and stalled replication forks, which demand an immediate response from the checkpoint machinery to preserve genomic integrity. If not corrected, the errors may lead to nucleotide sequence alterations and chromosomal rearrangements, usually associated with cellular transformation.6 The integrity of S-phase is monitored mainly via two phosphoinositide 3-kinase (PI3) related kinases—ATM (ataxia telangiectasia mutated) and ATR (ATM and Rad 3-related)—which play critical roles in early signal transmission.4,7 These two serine/threonine kinases play a central role in DNA damage signaling.4 G1- to S-phase transition requires the coordinated action of many proteins including members of the E2F family of transcription factors.8 E2F activity is required for the transcriptional induction of many genes required for cell cycle progression, including cyclins, cyclin-dependent kinases (CDK) and enzymes involved in DNA replication. In addition, E2F functions in apoptosis initiated by DNA damage regulating the expression of pro-apoptotic genes including apaf-1 and some caspases.9 ATM and ATR are thought to be involved in the transmission of signals from stalled replication forks through a multitude of signaling cascades. E2F-1 protein levels, DNA binding activity and transcriptional activity have been found to increase following exposure to UV light10-12 and other DNA-damaging agents.13 Thus DNA-damage may induce apoptosis through E2F-1. In turn, the association of this apoptosis with the S-phase may result from the fact that E2F-1 is in its free form (i.e., not complexed with retinoblastoma protein Rb (pRb)) and thus is most active during this phase of the cell cycle.14 pRb and its related proteins (p107, p130) are thought to act as a checkpoint for G1/S transition in proliferating cells.15 In its inactive state, pRb is unphosphorylated and is believed to inhibit cell cycle progression by the repression of E2F transcription factors. Upon mitogenic stimulation, pRb is phosphorylated by active cyclin D/ CDK4/6 complexes and released from E2F, resulting in the derepression and transactivation of E2F-target genes required for S phase entry.14,16 However, there is evidence of the involvement of pRb, possibly activated and recruited to E2F-binding sites, in response to DNA damage.17 In addition, several genes involved in DNA repair were shown to be regulated by E2F. E2F deregulation up-regulates the expression of genes involved in the DNA-damage response, including ATM, BRCA1 and RAD51.18-21 The E2F-1 protein has been shown to be stabilized and its levels to elevate in response to DNA damage.22 These findings support a role for E2F transcription factors in the DNA damage response. E2F regulation of DNA repair-associated genes at the G1/S transition is consistent with the view that DNA replication is an error-prone process and that DNA repair is tightly linked to DNA synthesis.20,23 Thus, DNA damage response, apoptosis and the cell cycle share some common participants such as pRb, E2F, and ATM/ATR.24,25 Another factor important for the cell cycle arrest, DNA repair, and apoptosis is the p53 tumor suppressor protein, which is also one of the most important sensors of DNA damage. In unstressed cells, p53 is maintained at low levels by targeted degradation due to binding by murine double minute-2 (MDM2) protein.26 Genotoxic stress initiates signaling pathways, leading to its stabilization and activation as a transcription factor.27 The G1/S cell cycle checkpoint is mediated primarily by activation and accumulation of p53, which in turn activates the gene encoding p21, a cyclin-dependent kinase inhibitor.28 In response to double-strand breaks or replication blocks, ATM acts

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distinct neuronal populations, but rather characterizes a common pathway involved in neuronal apoptosis. Since oxidative stress and the associated DNA damage, as well as cell cycle-related mechanisms, likely contribute to neuronal cell death52,85 and therefore to the pathophysiology of neurodegenerative diseases, the cell cycle activation may be requisite for DNA damage. While a number of laboratories have reported a correlation between neuronal apoptosis in neurodegenerative diseases and the appearance of cell cycle-related proteins, a role for cell cycle activation in terminally differentiated cells remains obscure. Does the cell cycle activation represent an obligatory part of DNA damage response in general, like in proliferative cells, or does it take place when a cell ”is making a choice” between life and death, DNA repair and apoptosis? Is cell cycle activation obligatory only for the execution of DNA damage-initiated apoptosis? In other words, do neurons need cell cycle activation only for death or also for DNA repair? There is evidence of greater activity of various DNA repair enzymes in proliferating, as compared to resting cells.86,87 DNA repair pathways might be acting as a function of the cell cycle stage because of greater DNA repair activity of the involved enzymes. However, this aspect of cell cycle activation in neurons remains obscure. The in vivo evidence that the cell cycle re-entry of postmitotic terminally differentiated neurons is associated with DNA damage came from the analysis of the X-linked harlequin (Hq) mutation in the gene encoding apoptosis-inducing factor (AIF) that causes progressive ataxia.88 In Hq mice, many cerebellar granule cells had newly synthesized nuclear DNA, as demonstrated by BrdU incorporation, and were positive for caspase-3 indicating the association with apoptosis. In addition, all cycling neurons were positive for oxo8dG, although not all of oxo8dG-positive neurons were in S-phase. In cycling cells, whether cells exposed to DNA-damaging conditions undergo growth arrest or apoptosis may depend in part on where the cell resides in the cell cycle when insulted. It has been found, for instance, that the majority of the apoptotic fibroblasts were in the S-phase of the cell cycle following their exposure to H2O2.89 Similar results have been shown for HL-60 cells exposed to topoisomerase I and II inhibitors.90 The fact that not all of oxo8dG-positive neurons in brains of Hq mice were in S-phase, indicate that these neurons may enter S-phase later and then die by apoptosis or that these oxo8dG-positive neurons may repair their DNA and do not enter S-phase followed by apoptosis, but continue normal function. Using flow cytometry and BrdU incorporation analyses, we have demonstrated that cell cycle activation followed by apoptosis was induced by DNA damage and can be blocked together with DNA damage response.64 These observations and the detection of markers for oxidative DNA damage (long before neurodegeneration) demonstrate that the induction of unscheduled cell cycle re-entry is highly correlated with, and is likely induced by DNA damage. Furthermore, the amount of oxo8dG, a major oxidative DNA lesion has been found to be elevated in DNA of neurons in brains of patients with neurodegenerative disorders.91 Increases in oxo8dG are also correlated with increased incidence of cancer66 demonstrating the importance of the same factors in the pathogenesis of both types of disorders.

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Neuronal apoptosis is a prominent feature in a number of acute and chronic neurological diseases including stroke, Alzheimer’s disease (AD), and amyotrophic lateral sclerosis (ALS).50-52 At the same time, markers of oxidative stress are also found to be associated with many neurodegenerative disorders.53 Oxidative damage has been reported in Alzheimer’s and Parkinson’s diseases as well as other age-related neurodegenerative diseases, including progressive supranuclear palsy and prion disorders.54-57 DNA damage is an important initiator of neuronal death and has been implicated in a wide variety of neuropathological conditions.50,58,59 Damage to nucleic acids caused by reactive oxygen species occurs in the form of base modifications, single-strand breaks, double-strand breaks and one of the most common adducts formed from the reaction of reactive oxygen species with DNA 8-hydroxy-2’-deoxyguanosine (oxo8dG).60,61 Evidence for widespread single- and double-strand breaks in AD brains has been provided by in situ labeling methods.62,63 Thus, neuronal DNA damage often occurs as a result of oxidative stress and the increased production of reactive oxygen species. According to our data, elevated levels of oxo8dG in brains of APP mutant transgenic mice, a mouse model of AD, correlated with increased levels of soluble Aβ1-42/Aβ1-40.64 The amount of oxo8dG was found to be elevated in nuclear and mitochondrial DNA in neurons in diseased brain regions of patients with AD.65 Furthermore, studies have shown increases in this modified base to correlate with increased incidence of cancer.66 Thus, DNA damage is an important inducer of both carcinogenesis and neurodegeneration. In support, neurological abnormalities accompany a defective DNA damage response in different human syndromes such as ataxia telangiectasia and Cocayne syndrome,38 demonstrate connections between abnormalities of the DNA damage response and neurodegeneration.

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CELL CYCLE ACTIVATION IN POSTMITOTIC NEURONS

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Accumulating evidence suggests that neurodegeneration is linked to paradoxical re-entry of neurons into the cell cycle.16 However, while terminally differentiated neurons retain the ability to reactivate the cell cycle, it rarely leads to neuronal proliferation and typically induces apoptosis instead.16,67,68 Consistent with this idea are the rare incidence of brain tumors of neuronal origin and the resistance of neurons to oncogenic transformation.69 On the other hand, aneuploidy was demonstrated to be quite common in the brain of adult animals,70 meaning that DNA replication in terminally differentiated cells may lead to aneuploidy. Nevertheless, re-expression and activation of cell cycle proteins have been observed in dying neurons in brains of patients with neurodegenerative disorders.68,71-73 Recently, a direct interaction between cell cycle machinery and the apoptotic program was demonstrated in postmitotic neurons with the observation that cdc2, a cell cycle regulator, initiates apoptosis via direct activation of Bad, a trigger of apoptosis.74 The suppression of cyclin-dependent kinase (CDK) activity has been shown to produce a neuroprotective effect both in vivo and in vitro,75-77 supporting the idea that cell cycle re-entry underlies neuronal apoptosis. Both cell cycle and apoptosis are controlled by a highly conserved machinery, exhibit morphological similarities of cell rounding and chromatin condensation, and are controlled by several common factors such as ATM, E2F and p53.24,78 The cell cycle activation has been shown in different paradigms of neurodegeneration such as Alzheimer’s73 and Parkinson’s79 diseases, ischemia,80 ALS,81 Down’s syndrome,82 excitotoxicity,83 Niemann-Pick’s disease.84 Thus, re-entry into the cell cycle is not specific to particular neurodegenerative diseases or

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CONCLUSION

In summary, evidence is mounting to show that oxidative stress and the DNA damage produced by this stress, may activate cell cycle machinery in postmitotic neurons as an essential part of DNA damage response. As in cycling cells, DNA damage may result in

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17. Knudsen KE, Booth D, Naderi S, Sever-Chroneos Z, Fribourg AF, Hunton IC, et al. RBdependent S-phase response to DNA damage. Mol Cell Biol 2000; 20:7751-63. 18. Ishida S, Huang E, Zuzan H, Spang R, Leone G, West M, et al. Role for E2F in control of both DNA replication and mitotic functions as revealed from DNA microarray analysis Mol Cell Biol 2001; 21:4684-99. 19. Polager S, Kalma Y, Berkovich E, Ginsberg D. E2Fs up-regulate expression of genes Involved in DNA replication, DNA repair and mitosis. Oncogene 2002; 21:437-46. 20. Ren B, Cam H, Takahashi Y, Volkert T, Terragni J, Young RA, et al. E2F integrates cell cycle progression with DNA repair, replication, and G2/M checkpoints. Genes Dev 2002; 16:245-56. 21. Berkovich E, Ginsberg D. ATM is a target for positive regulation by E2F-1. Oncogene 2003; 22:161-7. 22. Stevens C, La Thangue NB. A new role for E2F-1 in checkpoint control. Cell Cycle 2003; 2:435-7. 23. Weinmann AS, Yan PS, Oberley MJ, Huang TH, Farnham PJ. Isolating human transcription factor targets by coupling chromatin immunoprecipitation and CpG island microarray analysis. Genes Dev 2002; 16:235-44. 24. King KL, Cidlowski JA. Cell cycle and apoptosis:Common pathways to life and death. J Cell Biochem 1995; 58:175-80. 25. Stevaux O, Dyson NJ. A revised picture of the E2F transcriptional network and RB function.Curr Opin Cell Biol 2002; 14:684-91. 26. Sionov RV, Moallem E, Berger M, Kazaz A, Gerlitz O, Ben-Neriah Y, et al. c-Abl neutralizes the inhibitory effect of Mdm2 on p53. J Biol Chem 1999; 274:8371-4. 27. Appella E, Anderson CW. Post-translational modifications and activation of p53 by genotoxic stresses. Eur J Biochem 2001; 268:2764-72. 28. Shiloh Y. ATM and related protein kinases:Safeguarding genome integrity. Nat Rev Cancer 2003; 3:155-68. 29. Matsuoka S, Rotman G, Ogawa A, Shiloh Y, Tamai K, Elledge SJ. Ataxia telangiectasia-mutated phosphorylates Chk2 in vivo and in vitro. Proc Natl Acad Sci USA 2000; 97:10389-94. 30. Hirao A, Kong YY, Matsuoka S, Wakeham A, Ruland J, Yoshida H, et al. DNA damageinduced activation of p53 by the checkpoint kinase Chk2. Science 2000; 287:1824-7. 31. Kowalik TF, DeGregori J, Schwarz JK, Nevins JR. E2F1 overexpression in quiescent fibroblasts leads to induction of cellular DNA synthesis and apoptosis. J Virol 1995; 69:2491500. 32. Matsumura I, Tanaka H, Kanakura Y. E2F1 and c-Myc in cell growth and death.Cell Cycle 2003; 2:333-8. 33. Miyashita T, Kitada S, Krajewski S, Horne WA, Delia D, Reed JC. Overexpression of the Bcl-2 protein increases the half-life of p21Bax. J Biol Chem 1995; 270:26049-52. 34. Fortin A, Cregan SP, MacLaurin JG, Kushwaha N, Hickman ES, Thompson CS, et al APAF1 is a key transcriptional target for p53 in the regulation of neuronal cell death. J Cell Biol 2001; 155:207-16. 35. Chipuk JE, Kuwana T, Bouchier-Hayes L, Droin NM, Newmeyer DD, Schuler M, et al. Direct activation of Bax by p53 mediates mitochondrial membrane permeabilization and apoptosis. Science 2004; 303:1010-4. 36. McCombe P, Lavin M, Kidson C. Control of DNA repair linked to neuroblastoma differentiation. Int J Radiat Biol Relat Stud Phys Chem Med 1976; 29:523-1. 37. Hanawalt PC, Gee P, Ho L, Hsu RK, Kane CJ. Genomic heterogeneity of DNA repair. Role in aging? Ann N Y Acad Sci 1992; 663:17-25. 38. Rolig RL, McKinnon PJ. Linking DNA damage and neurodegeneration. Trends Neuroscie 2000; 23:417-24. 39. Brooks PJ. Brain atrophy and neuronal loss in alcoholism: A role for DNA damage? Neurochem Int 2000; 37:403-12. 40. Gobbel GT, Bellinzona M, Vogt AR, Gupta N, Fike JR, Chan PH. Response of postmitotic neurons to X-irradiation:Implications for the role of DNA damage in neuronal apoptosis. J Neurosci1998; 18:147-55. 41. Morris EJ, Geller HM. Induction of neuronal apoptosis by camptothecin, an inhibitor of DNA topoisomerase-I:Evidence for cell cycle-independent toxicity. J Cell Biol1996; 134, 757-70. 42. Roca AI, Cox MM. RecA protein:Structure, function, and role in recombinational DNA repair. Prog Nucleic Acid Res Mol Biol 1997; 56:129-223. 43. Nouspikel T, Hanawalt PC. Terminally differentiated human neurons repair transcribed genes but display attenuated global DNA repair and modulation of repair gene expression. Mol Cell Biol 2000; 20:1562-70. 44. Ho L, Hanawalt PC. Gene-specific DNA repair in terminally differentiating rat myoblasts Mutat Res 1991; 255:123-41. 45. Bohr VA, Smith CA, Okumoto DS, Hanawalt PC. DNA repair in an active gene:Removal of pyrimidine dimers from the DHFR gene of CHO cells is much more efficient than in the genome overall. Cell 1985; 40:359-69. 46. Mellon I, Spivak G, Hanawalt PC. Selective removal of transcription-blocking DNA damage from the transcribed strand of the mammalian DHFR gene.Cell 1987; 51:241-9. 47. van den Boom V, Jaspers NG, Vermeulen W. When machines get stuck—obstructed RNA polymerase II:Displacement, degradation or suicide. Bioessays. 2002; 24:780-4. 48. Nouspikel T, Hanawalt PC. DNA repair in terminally differentiated cells. DNA Repair (Amst) 2002; 1:59-75. 49. Ljungman M, Zhang F, Chen F, Rainbow AJ, McKay BC. Inhibition of RNA polymerase II as a trigger for the p53 response. Oncogene 1999; 18:583-92. 50. Mattson MP. Apoptosis in neurodegenerative disorders. Nat Rev Mol Cell Biol 2000; 1:120-9.

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DNA repair or apoptosis if DNA damage is left unrepaired, and the consequent loss of neurons by apoptosis could result in the neurodegeneration seen in patients with neurodegenerative diseases or diseases associated with impairment of DNA repair. It is quite possible that the involvement of cell cycle machinery in both DNA repair and apoptosis is as critical for neurons, as it is for proliferating cells. However, the mechanisms by which a cell is forced into apoptosis versus DNA repair are elusive for cycling cells and even more so for postmitotic cells. It is likely that both outcomes are managed by the DNA damage response. In neurons, suppression of the ATM, which is a proximal component of DNA damage response, attenuates both apoptosis and cell cycle re-entry,64 suggesting that both cell cycle activation and apoptosis constitute the DNA damage response. ATM deficiency in humans is characterized by predisposition to cancer and progressive neurodegeneration,92 suggesting the impairment of DNA repair when main player in the DNA damage response does not act. Cells derived from patients deficient in ATM are hypersensitive to inducers of DNA double-strand breaks.93 In addition, neurons from ATM-deficient mice are more resistant to apoptosis initiated by DNA damage,64,94-96 suggesting that ATM may function to eliminate neural cells when they have incurred sufficient DNA damage. However, inhibition of ATM not only protected neurons against DNA damage-triggered apoptosis, but also prevented S-phase re-entry of the cells.64 Complementary to these results, cycling cells deficient in ATM exhibit defective cell cycle checkpoints at the G1/S transition, during S-phase, and at the G2/M boundary28 in support the idea that neurons may be influenced by the same cell cycle checkpoints that govern apoptosis in cycling cells. While the role and mechanisms of cell cycle regulation of neuronal apoptosis are just beginning to be characterized, the fundamental question of why postmitotic neurons engage cell cycle machinery to control apoptosis and maybe DNA repair, remains the subject of considerable speculation.

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