Article

Loss of Wild-Type ATRX Expression in Somatic Cell Hybrids Segregates with Activation of Alternative Lengthening of Telomeres

Cancer Research Unit, Children's Medical Research Institute, Westmead, New South Wales, Australia.
PLoS ONE (Impact Factor: 3.23). 11/2012; 7(11):e50062. DOI: 10.1371/journal.pone.0050062
Source: PubMed

ABSTRACT

Alternative Lengthening of Telomeres (ALT) is a non-telomerase mechanism of telomere lengthening that occurs in about 10% of cancers overall and is particularly common in astrocytic brain tumors and specific types of sarcomas. Somatic cell hybridization analyses have previously shown that normal telomerase-negative fibroblasts and telomerase-positive immortalized cell lines contain repressors of ALT activity, indicating that activation of ALT results from loss of one or more unidentified repressors. More recently, ATRX or DAXX was shown to be mutated both in tumors with telomere lengths suggestive of ALT activity and in ALT cell lines. Here, an ALT cell line was separately fused to each of four telomerase-positive cell lines, and four or five independent hybrid lines from each fusion were examined for expression of ATRX and DAXX and for telomere lengthening mechanism. The hybrid lines expressed either telomerase or ALT, with the other mechanism being repressed. DAXX was expressed normally in all parental cell lines and in all of the hybrids. ATRX was expressed normally in each of the four telomerase-positive parental cell lines and in every telomerase-positive hybrid line, and was abnormal in the ALT parental cells and in all but one of the ALT hybrids. This correlation between ALT activity and loss of ATRX expression is consistent with ATRX being a repressor of ALT.

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    • "ATRX, together with DAXX, act together to deposit H3.3 at specific heterochromatic loci, such as telomeres, in a replication-independent manner143144145146147. Loss of ATRX or DAXX would impair H3.3 loading at telomeres, and mutations in the genes encoding ATRX, DAXX, and H3.3 are associated with ALT cancers[138,148149150. ASF1, originally identified in S. cerevisiae[151], is a histone chaperone involved in both the replication-coupled and replication-independent incorporation of H3.1-H4 and H3.3-H4 histone dimers into nucleosomes[152,153]. "

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    • "However, whether and how ATRX loss may drive telomeric chromatin remodeling during the process of ALT activation is still unknown, as ATRX depletion alone is not sufficient to promote telomeric recombinations (18). It was, however, reported that ATRX loss may facilitate ALT establishment during in vitro immortalization of human fibroblasts with SV40T antigen (33), suggesting that ATRX loss may impact on telomeric HC when this happens during the telomere shortening phase that precedes activation of a TMM. Hence, it is possible that ATRX loss may facilitate subtelomeric DNA invasion by shortened telomeres. "
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    ABSTRACT: Proper telomeric chromatin configuration is thought to be essential for telomere homeostasis and stability. Previous studies in mouse suggested that loss of heterochromatin marks at telomeres might favor onset of Alternative Lengthening of Telomeres (ALT) pathway, by promoting homologous recombination. However, analysis of chromatin status at human ALT telomeres has never been reported. Here, using isogenic human cell lines and cellular hybrids, which rely either on telomerase or ALT to maintain telomeres, we show that chromatin compaction is reduced at ALT telomeres and this is associated with a global decrease in telomeric H3K9me3. This, subsequently, leads to upregulation of telomere transcription. Accordingly, restoration of a more condensed telomeric chromatin through telomerase-dependent elongation of short ALT telomeres reduces telomere transcription. We further show that loss of ATRX chromatin remodeler function, a frequent characteristic of ALT cells, is not sufficient to decrease chromatin condensation at telomeres nor to increase the expression of telomeric RNA species. These results offer new insight on telomeric chromatin properties in ALT cells and support the hypothesis that telomeric chromatin decondensation is important for ALT pathway.
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    • "How would G34R/V mutations function? Notably, H3.3 G34 mutations almost invariably coexist with ATRX mutations (Henikoff, 2008; Schwartzentruber et al., 2012) and are associated with the alternative lengthening of telomeres (ALT) mechanism (Heaphy et al., 2011; Bower et al., 2012; Liu et al., 2012; Lovejoy et al., 2012), a recombinogenic mechanism for telomere elongation . ALT cells contain a modified PML-NB called ALT-associated PML-NB (APB), which we and others have implicated in telomeric damage response and potentially telomere recombination (Stagno D'Alcontres et al., 2007; Lovejoy et al., 2012). "
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