Functional telomeres are required for the replicability of cancer cells. The G-rich strand of telomeric DNA can fold into a 4-stranded structure known as the G-quadruplex (G4), whose stabilization alters telomere function limiting cancer cell growth. Therefore, the G4 ligand RHPS4 may possess antitumor activity. Here, we show that RHPS4 triggers a rapid and potent DNA damage response at telomeres in human transformed fibroblasts and melanoma cells, characterized by the formation of several telomeric foci containing phosphorylated DNA damage response factors γ-H2AX, RAD17, and 53BP1. This was dependent on DNA repair enzyme ATR, correlated with delocalization of the protective telomeric DNA-binding protein POT1, and was antagonized by overexpression of POT1 or TRF2. In mice, RHPS4 exerted its antitumor effect on xenografts of human tumor cells of different histotype by telomere injury and tumor cell apoptosis. Tumor inhibition was accompanied by a strong DNA damage response, and tumors overexpressing POT1 or TRF2 were resistant to RHPS4 treatment. These data provide evidence that RHPS4 is a telomere damage inducer and that telomere disruption selectively triggered in malignant cells results in a high therapeutic index in mice. They also define a functional link between telomere damage and antitumor activity and reveal the key role of telomere-protective factors TRF2 and POT1 in response to this anti-telomere strategy.
Polycomb response elements (PREs) are regulatory sites that mediate the silencing of homeotic and other genes. The bxd PRE region from the Drosophila Ultrabithorax gene can be subdivided into subfragments of 100 to 200 bp that retain different degrees of PRE activity in vivo. In vitro, embryonic nuclear extracts form complexes containing Polycomb group (PcG) proteins with these fragments. PcG binding to some fragments is dependent on consensus sequences for the GAGA factor. Other fragments lack GAGA binding sites but can still bind PcG complexes in vitro. We show that the GAGA factor is a component of at least some types of PcG complexes and may participate in the assembly of PcG complexes at PREs.Polycomb group (PcG) proteins from complexes in vivo at regulatory sites, the Polycomb response elements (PREs), which mediate the silencing of neighboring genes. Transposons containing PREs generate new binding sites for PcG proteins on polytene chromosomes, indicating that PREs are the physical targets for PcG complex formation. Chromatin cross-linking experiments have also shown that PcG proteins are bound to and in the vicinity of known PRE sites (28,35,36). In these experiments, PcG proteins are found cross-linked over a few kilobases centered over fragments with known PRE activity, suggesting the possibility that a silencing complex initiated at a PRE involves at least a few kilobases either because of a cooperative spreading of the complex or because the PRE is in fact not a site but a region containing multiple sequences that interact with PcG proteins. None of the well-characterized PcG proteins can be shown to bind to DNA in vitro. One simple explanation for the fact that PcG complex formation appears to be specific for the PRE might be that it depends on other hitherto-unknown PcG proteins. One such candidate, the product of the pleiohomeotic (pho) gene, has recently been identified (3). However, it has not been shown yet that PHO interacts with other PcG proteins, and the presumptive consensus sequence for the binding of PHO is not always present in DNA fragments that have PRE activity, suggesting that other DNA-binding proteins might be involved. Other possibilities are suggested by the properties of PcG proteins and of PREs. Several of the PcG proteins can interact with one another, and experiments with Pc protein targeted to LexA or GAL4 binding sites indicate that a single PcG protein can recruit a silencing complex (26; S. Poux, D. McCabe, and V. Pirrotta, submitted for publication). Different genomic PcG sites show different degrees of dependence on different members of the PcG genes, both in the strength of the signal generated by immunostaining and the effect of different PcG mutations on the silencing of the accompanying genes. The silencing ability of a given PRE is strongly dependent on the genomic context in which it is introduced and on homologous pairing or the physical proximity to other PRE sequences in the nucleus (5, 10, 17, 34). All these observations suggest that the formation of a PcG c...
The study of the proteins that bind to telomeric DNA in mammals has provided a deep understanding of the mechanisms involved in chromosome-end protection. However, very little is known on the binding of these proteins to nontelomeric DNA sequences. The TTAGGG DNA repeat proteins 1 and 2 (TRF1 and TRF2) bind to mammalian telomeres as part of the shelterin complex and are essential for maintaining chromosome end stability. In this study, we combined chromatin immunoprecipitation with high-throughput sequencing to map at high sensitivity and resolution the human chromosomal sites to which TRF1 and TRF2 bind. While most of the identified sequences correspond to telomeric regions, we showed that these two proteins also bind to extratelomeric sites. The vast majority of these extratelomeric sites contains interstitial telomeric sequences (or ITSs). However, we also identified non-ITS sites, which correspond to centromeric and pericentromeric satellite DNA. Interestingly, the TRF-binding sites are often located in the proximity of genes or within introns. We propose that TRF1 and TRF2 couple the functional state of telomeres to the long-range organization of chromosomes and gene regulation networks by binding to extratelomeric sequences.
The nuclear receptor family comprises ligand-dependent and orphan receptors. To the latter group belong the estrogen receptor-related receptors (ERRs) for which conflicting results have been published concerning the nature (constitutive or liganded) of their transcriptional activities. ERRs interfere in various ways, positively and negatively, with estrogen signaling. Moreover recent data analyzing ERR expression in human breast tumors have proposed ERR and ERR as prognostic markers of these cancers. The identification of modulators (positive or negative) of ERR activities would therefore be highly useful in our understanding of estrogen-related pathologies. The purpose of this review is to summarize our knowledge of the nature of ERR activities and progresses in identifying synthetic ERR modulators.
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