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Telomeres are the nucleoprotein structures found at the ends of linear chromosomes (2, 6, 9). Telomeric DNA consists of tandem arrays of short repeats that have G-rich and C-rich strands. The telomeric DNA is divided into a proximal double-stranded region and a more distal single-stranded 3′ overhang (36). This overhang is comprised solely of the G-rich strand and is elongated through the activity of the reverse transcriptase telomerase (53). Telomerase minimally consists of a template RNA that is reverse transcribed onto telomeric ends by the catalytic protein subunit. Telomeric DNA also associates with a large assemblage of proteins (43, 51). The telomere protein-DNA complex acts as a cap that is dynamic in composition and arrangement. It is the dynamic protein-DNA telomere complex that both controls telomere metabolism and protects telomeres from being recognized as double-stranded breaks. One proposed mechanism for telomere protection is the creation of telomeric secondary structures, such as t-loops, which are stabilized through the action of telomeric DNA binding proteins (19, 49). The maintenance of telomere length is important to the normal functioning of cells. In most human somatic cells, telomerase activity is very low or absent, so that telomeres gradually shorten, eventually reaching a state which triggers a growth arrest called replicative senescence (15, 40). Mutations or other conditions that bypass senescence produce further telomere shortening and take cells into crisis, a state of high genetic instability and cell death caused by widespread telomere dysfunction (41). In order to survive crisis and become immortal, a means of telomere elongation must arise (14, 39). Most cancer cells are immortal due to the presence of telomerase (40). A subset of cells that emerge from crisis maintain their telomeres through alternate lengthening of telomeres (ALT), a process that involves intertelomeric recombination (3, 16, 38). Studies have shown that telomeres of ALT cells are both highly heterogeneous in size and frequently much longer than telomeres in normal human cells and telomerase-positive cancer cells (3, 4). ALT cells also commonly contain intranuclear structures known as ALT-associated PML bodies, or APBs. APBs contain recombination proteins, telomeric DNA, and replication factor A and have therefore been suggested to play a role in the mechanism of ALT telomere elongation (33, 55). While the structure of telomeric DNA associated with APBs is unknown, extrachromosomal telomeric repeats (ECTR) have recently been extracted from ALT cell lines and have been shown to at least partially exist in circular conformations (7, 52). Yeast cells have constitutively active telomerase and normally maintain their telomeres within a fixed size range (17). However, deletion of telomerase in yeast cells leads to telomere shortening and coincident growth senescence (27, 29, 42). Although most cells eventually die from this senescence, some postsenescence survivors emerge. These survivors, which are dependent upon the recombination gene RAD52 (26, 28), elongate their telomeres through recombinational telomere elongation (RTE), a process that appears to be analogous to human ALT. In Saccharomyces cerevisiae, there are two pathways of RTE, each of which produces a distinctly different elongation pattern. Type I survivors have amplification of the long Y′ subtelomeric elements and short extensions of the telomeric TG(1-3) repeats (10, 26, 46). Type II survivors lack subtelomeric amplification and are characterized by long telomeric TG(1-3) extensions which are heterogeneous in length (10, 46). Both type I and type II survivors of Saccharomyces require RAD52 as well as type-specific sets of recombination genes. Type I survivors require RAD51, RAD54, and RAD57, while type II survivors require components of the MRX complex, RAD59, and the helicase SGS1 (10, 12, 21, 24, 45). Only type II survivors arise when the telomerase RNA gene (TER1) is deleted from Kluveromyces lactis cells (28). Inspection of telomeric sequence from K. lactis ter1-Δ survivors derived from cells with telomeres composed of basal wild-type and terminal phenotypically silent mutant repeats revealed repeating patterns containing both types of repeats (35). Most or all telomeres within a clone shared a single pattern, but patterns varied between survivor clones. These data led us to propose the roll-and-spread model, which proposes that the first elongated telomere is made within a cell by rolling circle replication around a circle as small as 100 bp and that the sequence of this long telomere is then spread by gene conversion to most or all other telomeres (35). Consistent with this model, K. lactis cells can utilize transformed telomeric circles of 1.6 kb and 100 nucleotides (nt) to elongate their telomeres (34, 35). Other recent work has confirmed that the sequence of one elongated telomere spreads at very high efficiency to all other telomeric ends during survivor formation (49a). In this work we show that the unusually small telomeric circles proposed by the roll-and-spread model can be made by recombination in a long telomere mutant of K. lactis.

The human DNA damage sensors, Rad17-replication factor C (Rad17-RFC) and the Rad9-Rad1-Hus1 (9-1-1) checkpoint complex, are thought to be involved in the early steps of the DNA damage checkpoint response. Rad17-RFC and the 9-1-1 complex have been shown to be structurally similar to the replication factors, RFC clamp loader and proliferating cell nuclear antigen polymerase clamp, respectively. Here, we demonstrate functional similarities between the replication and checkpoint clamp loader/DNA clamp pairs. When all eight subunits of the two checkpoint complexes are coexpressed in insect cells, a stable Rad17-RFC/9-1-1 checkpoint supercomplex forms in vivo and is readily purified. The two individually purified checkpoint complexes also form a supercomplex in vitro , which depends on ATP and is mediated by interactions between Rad17 and Rad9. Rad17-RFC binds to nicked circular, gapped, and primed DNA and recruits the 9-1-1 complex in an ATP-dependent manner. Electron microscopic analyses of the reaction products indicate that the 9-1-1 ring is clamped around the DNA.

In human somatic cells, telomeres shorten every cell division due to the end replication problem. Once reaching a critically short length, the cells will undergo permanent cell cycle arrest and become senescent. Cancer cells acquire unlimited proliferation ability by activation of a telomere maintenance mechanism, either the enzyme telomerase or the homologous recombination-based mechanism Alternative Lengthening of Telomeres (ALT). Cancers that utilize the ALT mechanism commonly are deficient for ATRX protein expression, are difficult to treat, and have a poor prognosis. We discovered that ICP0-null herpes simplex virus type 1 (HSV-1) was ten to one thousand-fold more effective in killing cancer cell lines that are ATRX-deficient. Sensitivity to mutant HSV-1 infection resulted from ATRX-dependent regulation of PML expression at both the transcriptional and post-transcriptional levels. Reduction of PML protein resulted in a concomitant reduction in PML nuclear bodies, which weakened the innate cellular immunity to viral infection. Infection of co-cultured primary and ATRX-null cancer cells revealed that mutant ICP0-null HSV-1 treatment preferentially killed ATRX-null cells. Our results suggest that mutant ICP0-null HSV-1 may have therapeutic benefit against ATRX-null cells. Moreover, these data provide an applicable approach for predicting, based on tumor ATRX or PML protein status, which tumors will respond to an oncolytic herpes virus. Citation Format: Mingqi Han, Christine E. Napier, Sonja Frölich, Roger D. Everett, Anthony J. Cesare, Roger R. Reddel. Synthetic lethality of cytolytic HSV-1 in cancer cells with ATRX and PML deficiency [abstract]. In: Proceedings of the AACR Precision Medicine Series: Opportunities and Challenges of Exploiting Synthetic Lethality in Cancer; Jan 4-7, 2017; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2017;16(10 Suppl):Abstract nr A22.

Double strand break (DSB) repair is suppressed during mitosis because RNF8 and downstream DNA damage response (DDR) factors, including 53BP1, do not localize to mitotic chromatin. Discovery of the mitotic kinase-dependent mechanism that inhibits DSB repair during cell division was recently reported. It was shown that restoring mitotic DSB repair was detrimental, resulting in repair dependent genome instability and covalent telomere fusions. The telomere DDR that occurs naturally during cellular aging and in cancer is known to be refractory to G2/M checkpoint activation. Such DDR-positive telomeres, and those that occur as part of the telomere-dependent prolonged mitotic arrest checkpoint, normally pass through mitosis without covalent ligation, but result in cell growth arrest in G1 phase. The discovery that suppressing DSB repair during mitosis may function primarily to protect DDR-positive telomeres from fusing during cell division reinforces the unique cooperation between telomeres and the DDR to mediate tumor suppression.

Loss of chromosome end protection through telomere erosion is a hallmark of aging and senescence. Here we developed an experimental system that mimics physiological telomere deprotection in human cells and discovered that the telomere deprotection response is functionally distinct from the genomic DNA damage response. We found that, unlike genomic breaks, deprotected telomeres that are recognized as DNA damage but remain in the fusion-resistant intermediate state activate differential ataxia telangiectasia mutated (ATM) signaling where CHK2 is not phosphorylated. Also unlike genomic breaks, we found that deprotected telomeres do not contribute to the G2/M checkpoint and are instead passed through cell division to induce p53-dependent G1 arrest in the daughter cells. Telomere deprotection is therefore an epigenetic signal passed between cell generations to ensure that replication-associated telomere-dependent growth arrest occurs in stable diploid G1 phase cells before genome instability can occur.

Intrinsic limits on cellular proliferation in human somatic tissue serves as a tumor suppressor mechanism by restricting cell growth in aged cells with accrued pre-cancerous mutations. This is accompanied by the potential cost of restricting regenerative capacity and contributing to cellular and organismal aging. Emerging data support a model where telomere erosion controls proliferative boundaries through the progressive change of telomere structure from a protected state, through two distinct states of telomere deprotection. In this model telomeres facilitate a controlled permanent cell cycle arrest with a stable diploid genome during differentiation and may serve as an epigenetic sensor of general stress in DNA metabolism processes.

Summary Telomere shortening and disruption of telomeric components are pathways that induce telomere deprotection. Here we describe another pathway, where prolonged mitotic arrest induces damage signals at telomeres in human cells. Exposure to microtubule drugs, kinesin inhibitors, proteasome inhibitors or the disruption of proper chromosome cohesion resulted in the formation of damage-foci at telomeres. Induction of mitotic telomere deprotection coincided with dissociation of TRF2 (Telomere Repeat binding Factor 2) from telomeres, telomeric 3’-overhang degradation and ATM (Ataxia Telangiectasia Mutated) activation, and could be suppressed by TRF2 overexpression or inhibition of Aurora B kinase. Normal cells that escape from prolonged mitotic arrest halted in the following G1 phase, whereas cells lacking p53 continued to cycle and became aneuploid. We propose a telomere dependent mitotic duration monitoring system that reacts to improper progression through mitosis.

Unlimited cellular proliferation depends on counteracting the telomere attrition that accompanies DNA replication. In human cancers this usually occurs through upregulation of telomerase activity, but in 10-15% of cancers - including some with particularly poor outcome - it is achieved through a mechanism known as alternative lengthening of telomeres (ALT). ALT, which is dependent on homologous recombination, is therefore an important target for cancer therapy. Although dissection of the mechanism or mechanisms of ALT has been challenging, recent advances have led to the identification of several genes that are required for ALT and the elucidation of the biological significance of some phenotypic markers of ALT. This has enabled development of a rapid assay of ALT activity levels and the construction of molecular models of ALT.

Telomere lengths are maintained in many cancer cells by the ribonucleoprotein enzyme telomerase but can be further elongated by increasing telomerase activity through the overexpression of telomerase components. We report here that increased telomerase activity results in increased telomere length that eventually reaches a plateau, accompanied by the generation of telomere length heterogeneity and the accumulation of extrachromosomal telomeric repeat DNA, principally in the form of telomeric circles (t-circles). Telomeric DNA was observed in promyelocytic leukemia bodies, but no intertelomeric copying or telomere exchange events were identified, and there was no increase in telomere dysfunction-induced foci. These data indicate that human cells possess a mechanism to negatively regulate telomere length by trimming telomeric DNA from the chromosome ends, most likely by t-loop resolution to form t-circles. Additionally, these results indicate that some phenotypic characteristics attributed to alternative lengthening of telomeres (ALT) result from increased mean telomere length, rather than from the ALT mechanism itself.

Telomeres are the nucleoprotein structures at eukaryotic chromosome termini that cap the chromosome ends. They consist of tandem G-rich repeats that terminate in a single-stranded 3′ overhang and proteins that interact with both the single-stranded and double-stranded regions of telomeric DNA. Disruption of telomere capping can lead to one or more outcomes, including telomere fusions, increased telomeric recombination, and replicative senescence (12). In higher eukaryotes, telomeres are commonly arranged into a structure termed a telomere loop (t-loop) that involves strand invasion of the 3′ overhang into internal telomeric repeats of the same chromosome end (14). Such loops have been seen in human, murine, chicken, plant, and ciliate telomeres (5, 14, 24, 27). T-loop formation in vitro is facilitated by human TRF2 protein (1, 14, 29), and inhibition of TRF2 in vivo results in telomere dysfunction and chromosome fusions (3, 35), suggesting that the ability to stabilize t-loops may be an important part of telomeric capping. Telomeres in the budding yeast Saccharomyces cerevisiae have been proposed to assume a fold-back structure mediated by protein-protein interactions, although the variable telomeric-repeat sequence may or may not allow strand invasion of the 3′ overhang to generate a t-loop (10, 11). The Taz1 protein, from the fission yeast Schizosaccharomyces pombe, is an ortholog TRF2, and in vitro studies have shown that, like TRF2, it has the ability to arrange model telomere substrates into looped structures (32). However, direct examination of native yeast telomere architecture by electron microscopy (EM) to confirm the presence or absence of t-loops in yeast has proven problematic, due to the difficulty of isolating the short telomeres in these species. This is because long telomere restriction fragments (TRFs), which can be separated from the short genomic restriction fragments by gel filtration chromatography, are a technical requirement for observation of native telomeric DNA by EM. While telomere attrition is most often counterbalanced by telomerase activity, homologous recombination-dependent telomere maintenance mechanisms also exist. A subset of human tumor and immortalized cells use a telomerase-independent mechanism termed alternative lengthening of telomeres (ALT) that is believed to be dependent on recombination (16), and recombinational telomere elongation (RTE) is well characterized in yeast mutants with telomerase deleted (23). Such mutants display gradual telomere shortening, leading to senescence. However, survivors can emerge from senescence if chromosome ends have become elongated by recombination (20, 22). Two types of survivors are characterized in S. cerevisiae: type I has short telomeres and exhibits amplification of the subtelomeric Y′ elements, while type II is similar to human ALT cells, exhibiting elongated, heterogeneous telomeric DNA (7, 20, 30). Survivors in the related budding yeast, Kluyveromyces lactis, are exclusively type II (22). Current data support a model of type II RTE termed “roll-and-spread” in which extrachromosomal telomeric repeat (ECTR) DNA circles (t-circles), created by telomeric recombination, act as a template for rolling-circle replication, elongating one (or more) telomere (26). The elongated telomere(s) then spreads to other chromosome ends via nonreciprocal recombination events. Support for this model comes mainly from K. lactis, in which exogenous t-circles were shown to promote RTE and a DNA sequence from one elongated telomere was shown to spread to other telomeres following RTE onset (25, 26, 33). Evidence of a roll-and-spread mechanism was also observed in the mitochondria of the yeast species Candida parapsilosis, which contains linear mitochondrial DNA capped by telomeres maintained in a telomerase-independent fashion (28). Consistent with a role in RTE, t-circles are a conserved feature in human ALT cells (4, 36), S. cerevisiae survivors (18, 19), and C. parapsilosis mitochondria (31), and as described below, they likely reflect a t-loop origin. Recent data also indicate that Ku70 deletion in Arabidopsis thaliana results in t-circle formation and the growth of callus cultures with ALT-like telomeric phenotypes (39). K. lactis provides an excellent opportunity for investigating telomere structure in a yeast species. Unlike S. cerevisiae, the 25-bp K. lactis telomeric repeat contains TA dinucleotides that allow interstrand cross-linking by psoralen (Fig. ​(Fig.1D),1D), a necessary step for maintaining t-loop structure during isolation (14). K. lactis mutants with telomeres long enough to permit isolation from the bulk genomic DNA following restriction enzyme fragmentation are available, either with or without obvious telomere-capping defects (34). The ter1-16T mutant, in particular, is well suited for such studies. Mutation of the telomerase RNA gene in ter1-16T generates telomeric repeats with a base change that alters the binding site for Rap1, a protein crucial to telomere length regulation and capping (Fig. ​(Fig.1D)1D) (34). The ter1-16T mutation leads to long, uncapped telomeres with abnormally long 3′ overhangs, both of which can be suppressed by RAP1 overexpression (34). ter1-16T cells have also been shown to produce small t-circles using homologous recombination, and this can be inhibited by RAD52 deletion (15). FIG. 1. Phenotypic characteristics and enrichment of telomeric DNA from K. lactis strains. (A) Detection of single-stranded G-rich telomeric DNA by in-gel hybridization without denaturation. Uncut genomic DNA (1 μg) was separated on a 0.8% agarose ... In this study, we have isolated telomeric DNA from the K. lactis ter1-16T strain for direct examination by EM and 2D gel analysis. We show that telomeric DNA from ter1-16T cells contains looped molecules of telomeric DNA with the physical characteristics of t-loops previously isolated from mammalian and plant cells, as well as abundant t-circles ranging up to the size of an entire telomere. In contrast, telomeric DNA from ter1-16T RAP1OE (overexpressing RAP1) and ter1-16T rad52Δ cells contained significantly fewer t-loops and t-circles. Cumulatively, these data suggest that increased recombination at K. lactis telomeres in the absence of Rap1 results in increased strand invasion at t-loop junctions and resolution of t-loops by homologous recombination into free t-circles.

The maintenance of telomere length is essential for the indefinite proliferation of cancer cells. This is most often achieved by the activation of telomerase; however, a substantial number of cancers lack detectable telomerase activity and are classified as using an alternative lengthening of telomeres (ALT) pathway. We showed recently that ALT cells have a high level of extrachromosomal telomeric circles (t circles) that may be a specific marker of the ALT phenotype. The mechanism underlying t circle production and the requirement of t circles in ALT remain unclear. Understanding the specific requirements of ALT is key to developing diagnostic tools and therapies that target this pathway and is critical for the treatment of cancers in which ALT is prevalent, including cancers of neuroepithelial and mesenchymal origin. In this study, we used short hairpin RNAs directed at either Xrcc3 or Nbs1, two proteins involved in the homologous recombination pathway, to determine the role of these proteins in t circle production and the requirement of t circles in maintaining the ALT pathway. We show that Xrcc3 and Nbs1 are indeed required for the production of t circles in human ALT. However, these cells continue to proliferate in the absence of t circles, suggesting that they are not required for the survival of ALT cells. [Cancer Res 2007;67(4):1513–9]

A prerequisite for cellular immortalization in human cells is the elongation of telomeres through the upregulation of telomerase or by the alternative lengthening of telomeres (ALT) pathway. In this study, telomere structure in multiple ALT cell lines was examined by electron microscopy. Nuclei were isolated from GM847, GM847-Tert, and WI-38 VA13 ALT cells, psoralen photo-cross-linked in situ, and the telomere restriction fragments were purified by gel filtration chromatography. Examination of telomere-enriched fractions revealed frequent extrachromosomal circles, ranging from 0.7 to 56.8 kb. t-loops were also observed, with the loop portion ranging from 0.5 to 70.2 kb. The total length of the loop plus tail of the t-loops corresponded to the telomere restriction fragment length from the ALT cell lines as determined by pulsed-field gel electrophoresis. The presence of extrachromosomal circles containing telomeric DNA was confirmed by two-dimensional pulsed-field gel electrophoresis. These results show that extrachromosomal telomeric DNA circles are present in ALT nuclei and suggest a roll-and-spread mechanism of telomere elongation similar to that seen in previous observations of multiple yeast species. Results presented here also indicate that expression of telomerase in GM847 cells does not affect t-loop or extrachromosomal circle formation.

Summary Telomeres vary greatly in size among plants and, in most higher plants, consist of a long array of 5′-TTTAGGG-3′/3′-AAATCCC-5′ (TTTAGGG) repeats. Recently, telomeric DNA in human, mouse, oxytricha, and trypanosome chromosomes have been found arranged into loops (t-loops), proposed to sequester the telomere from unwanted repair events and prevent activation of DNA damage checkpoints. We have asked whether t-loops exist in the higher order plant Pisum sativum (garden pea). DNA was isolated from the shoots and root tips of germinating seeds. Analysis of the telomeric restriction fragments showed that DNA hybridizing to a (TTTAGGG)n probe migrated as a smear centering around 25 kb, and direct sequencing verified the repeat to be (TTTAGGG)n. Total DNA in isolated nuclei was photo-cross-linked, and the telomeric restriction fragments were purified by gel filtration. Electron microscopic (EM) analysis revealed DNA molecules arranged as t-loops with a size distribution consistent with that seen by gel electrophoresis. Some molecules had loops as large as 75 kb. These results show that the arrangement of telomeric DNA into loops occurs in higher plants.

Prior evidence has supported the existence of multiple susceptibility genes for schizophrenia. Multipoint linkage analysis of the 270 Irish high-density pedigrees that we have studied, as well as results from several other samples, suggest that at least one such gene is located in region 6p24-21. In the present study, family-based association analysis of 36 simple sequence-length-polymorphism markers and of 17 SNP markers implicated two regions, separated by approximately 7 Mb. The first region, and the focus of this report, is 6p22.3. In this region, single-nucleotide polymorphisms within the 140-kb gene DTNBP1 (dystrobrevin-binding protein 1, or dysbindin) are strongly associated with schizophrenia. Uncorrected, empirical P values produced by the program TRANSMIT were significant (P.01) for a number of individual SNP markers, and most remained significant when the data were restricted to include only one affected offspring per nuclear family per extended pedigree; multiple three-marker haplotypes were highly significant (P=.008-.0001) under the restricted conditions. The pattern of linkage disequilibrium is consistent with the presence of more than one susceptibility allele, but this important issue is unresolved. The number of markers tested in the adjacent genes, all of which are negative, is not sufficient to rule out the possibility that the dysbindin gene is not the actual susceptibility gene, but this possibility appears to be very unlikely. We conclude that further investigation of dysbindin is warranted.

Telomere shortening and disruption of telomeric components are pathways that induce telomere deprotection. Here we describe another pathway, where prolonged mitotic arrest induces damage signals at telomeres. Exposure to microtubule drugs, kinesin inhibitors, proteasome inhibitors or the disruption of proper chromosome alignment resulted in the formation of damage-foci at telomeres. Induction of mitotic telomere deprotection coincided with dissociation of TRF2 from telomeres, telomeric 3′-overhang degradation and ATM activation, and could be suppressed by TRF2 overexpression or inhibition of Aurora B kinase. Normal cells that escape from prolonged mitotic arrest halted in the following G1 phase, whereas cells lacking p53 continued to cycle and became aneuploid. We propose a telomere dependent mitotic duration checkpoint that ensures elimination of cells, which fail to progress through mitosis. To avoid unwanted checkpoint activation by natural chromosome ends, cells have evolved telomeres. Human telomeres are composed of double stranded TTAGGG repeats and a single stranded G rich 3′ overhang, which are covered and protected by shelterin. Among the six shelterin components TRF2 and POT1 have predominantly been implicated in chromosome end protection by preventing ATM- and ATR-dependent checkpoint activation. Upon disruption of TRF2 or POT1 telomeres are recognized as sites of DNA damage, resulting in phosphorylation of histone H2AX (γ-H2AX) within the telomeric and sub-telomeric chromatin and association of 53BP1 with the chromosome ends. The co-localization of DNA-damage response factors and chromosome ends can be visualized as telomere dysfunction-induced foci (TIF). TIF have also been intimately linked to replicative senescence and shown to occur spontaneously in cancer cell lines. Cells arrested in mitosis are known to either die during mitotic arrest, or skip cytokinesis and “slip” into the subsequent G1 phase of the cell cycle. Mitotic slippage occurs through the degradation of Cyclin B1 in the presence of the active spindle assembly checkpoint (SAC). Cells that exit from prolonged mitotic arrest or progress through mitotic slippage exhibit various fates, including apoptosis or p53-dependent cell cycle arrest. In both normal and cancer cells, cell death during mitotic arrest, or apoptosis or senescence after escape from prolonged mitotic arrest are crucial for preventing chromosome instability. A failure to remove cells from the cycling population following prolonged mitotic arrest may allow cells to continue propagating with an abnormal number of chromosomes. However, despite intense research, the molecular mechanisms that trigger growth arrest or death in mitotically arrested cultures have not yet been identified. In the course of experimentation to explore putative telomeric functions for cohesin, we found that mitotic arrest per se induces telomere deprotection in primary and transformed human cells. Telomere deprotection during mitotic arrest associated with loss of the telomeric 3′-overhangs, led to ATM activation and was ATM dependent. TRF2 was dissociated from telomeres during prolonged mitotic arrest, providing the molecular basis for overhang loss and ATM activation, which was emphasized by the finding that TRF2 overexpression protected telomeres from the damage machinery during mitotic arrest. Inhibition of Aurora B kinase suppressed the telomere deprotection phenotype, suggesting the involvment of the SAC. Cells suffering from mitotic telomere deprotection underwent p53 dependent cell cycle arrest in the following G1 phase after mitotic release, while cells lacking p53 function continued to cycle and became aneuploid. Our findings provide a molecular mechanism explaining the induction of DNA damage signaling, cell cycle arrest or apoptosis following prolonged mitotic arrest, and explain the mechanism of action of therapeutic drugs, such as Taxol, Vinblastine and Velcade, which all inhibit mitotic progression. We propose that telomeric destabilization during mitotic arrest induces DNA damage signaling and serves as a mitotic duration checkpoint, responsible for eliminating cells that fail to progress through mitosis properly. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY23-02. doi:1538-7445.AM2012-SY23-02