ALT cancer cells are specifically sensitive to lysine acetyl transferase inhibition.

Some cancer cells elongate their telomeres through the ALT (alternative lengthening of telomeres) pathway, which is based on homologous recombination for the addition of telomere repeats without telomerase activity. two homologous lysine acetyltransferases, exert opposite effects on the ALT pathway, inhibiting or favoring it respectively. Here we show that ALT cells are particularly sensitive to the inhibition of acetyltransferases activities using Anacardic Acid (AA).

Opposite effects of GCN5 and PCAF knockdowns on the alternative mechanism of telomere maintenance.

Cancer cells can use a telomerase-independent mechanism, known as alternative lengthening of telomeres (ALT), to elongate their telomeres. General control non-derepressible 5 (GCN5) and P300/CBP-associated factor (PCAF) are two homologous acetyltransferases that are mutually exclusive subunits in SAGA-like complexes. Here, we reveal that down regulation of GCN5 and PCAF had differential effects on some phenotypic characteristics of ALT cells.

A preclinical mouse model of glioma with an alternative mechanism of telomere maintenance (ALT).

Glioblastoma multiforme is the most aggressive primary tumor of the central nervous system. Glioma stem cells (GSCs), a small population of tumor cells with stem-like properties, are supposedly responsible for glioblastoma multiforme relapse after current therapies. In approximately thirty percent of glioblastoma multiforme tumors, telomeres are not maintained by telomerase but through an alternative mechanism, termed alternative lengthening of telomere (ALT), suggesting potential interest in developing specific therapeutic strategies.

Assessing cell cycle progression of neural stem and progenitor cells in the mouse developing brain after genotoxic stress.

Neurons of the cerebral cortex are generated during brain development from different types of neural stem and progenitor cells (NSPC), which form a pseudostratified epithelium lining the lateral ventricles of the embryonic brain. Genotoxic stresses, such as ionizing radiation, have highly deleterious effects on the developing brain related to the high sensitivity of NSPC. Elucidation of the cellular and molecular mechanisms involved depends on the characterization of the DNA damage response of these particular types of cells, which requires an accurate method to determine NSPC progression through the cell cycle in the damaged tissue.

In vivo importance of homologous recombination DNA repair for mouse neural stem and progenitor cells.

We characterized the in vivo importance of the homologous recombination factor RAD54 for the developing mouse brain cortex in normal conditions or after ionizing radiation exposure. Contrary to numerous homologous recombination genes, Rad54 disruption did not impact the cortical development without exogenous stress, but it dramatically enhanced the radiation sensitivity of neural stem and progenitor cells. This resulted in the death of all cells irradiated during S or G2, whereas the viability of cells irradiated in G1 or G0 was not affected by Rad54 disruption.

Rad51 and DNA-PKcs are involved in the generation of specific telomere aberrations induced by the quadruplex ligand 360A that impair mitotic cell progression and lead to cell death.

Functional telomeres are protected from non-homologous end-joining (NHEJ) and homologous recombination (HR) DNA repair pathways. Replication is a critical period for telomeres because of the requirement for reconstitution of functional protected telomere conformations, a process that involves DNA repair proteins. Using knockdown of DNA-PKcs and Rad51 expression in three different cell lines, we demonstrate the respective involvement of NHEJ and HR in the formation of telomere aberrations induced by the G-quadruplex ligand 360A during or after replication.

Different DNA-PKcs functions in the repair of radiation-induced and spontaneous DSBs within interstitial telomeric sequences.

Interstitial telomeric sequences (ITSs) in hamster cells are hot spots for spontaneous and induced chromosome aberrations (CAs). Most data on ITS instability to date have been obtained in DNA repair-proficient cells. The classical non-homologous end joining repair pathway (C-NHEJ), which is the principal double strand break (DSB) repair mechanism in mammalian cells, is thought to restore the morphologically correct chromosome structure.

Sequence-driven telomeric chromatin structure.

Interstitial (also called internal or intrachromosomal) telomeric sequences (ITS) are found in many organisms.(1) In hamsters, CHO cells show long (up to several Mbp) ITS(2) (Fig. 1A) which are over involved in spontaneous or radiation induced chromosome aberrations.

Expression of cell cycle biomarkers and telomere length in papillary thyroid carcinoma: a comparative study between radiation-associated and spontaneous cancers.

Objective: Radiation exposure during childhood is the only well-established risk factor for papillary thyroid carcinoma (PTC). To better define the biologic profile of radiation-induced and sporadic PTC, we compared in these two groups of PTC the expression of cell cycle regulatory proteins and telomere length.

Methods: Cell cycle markers (cyclin A, B1, D1, E, and Ki67) were evaluated on 100 PTC specimens (26 radiation-induced and 74 sporadic PTCs).

Aquaporin 1 is overexpressed in lung cancer and stimulates NIH-3T3 cell proliferation and anchorage-independent growth.

The aquaporins represent a family of transmembrane water channel proteins that play a major role in trans-cellular and transepithelial water movement. Most tumors have been shown to exhibit high vascular permeability and interstitial fluid pressure, but the transport pathways for water within tumors remain unknown. Here, we tested 10 non-small cell lung cancer cell lines of various origins by reverse transcriptase-polymerase chain reaction and Western blot analysis and identified clear expression of aquaporin 1 (AQP1) in seven cell lines.

Impact of the KU80 pathway on NHEJ-induced genome rearrangements in mammalian cells.

Using a substrate measuring deletion or inversion of an I-SceI-excised fragment and both accurate and inaccurate rejoining, we determined the impact of non-homologous end-joining (NHEJ) on mammalian chromosome rearrangements. Deletion is 2- to 8-fold more efficient than inversion, independent of the DNA ends structure. KU80 controls accurate rejoining, whereas in absence of KU mutagenic rejoining, particularly microhomology-mediated repair, occurs efficiently.

Interstitial telomeric repeats are not preferentially involved in radiation-induced chromosome aberrations in human cells.

Telomeric repeat sequences, located at the end of eukaryotic chromosomes, have been detected at intrachromosomal locations in many species. Large blocks of telomeric sequences are located near the centromeres in hamster cells, and have been reported to break spontaneously or after exposure to ionizing radiation, leading to chromosome aberrations. In human cells, interstitial telomeric sequences (ITS) can be composed of short tracts of telomeric repeats (less than twenty), or of longer stretches of exact and degenerated hexanucleotides, mainly localized at subtelomeres.

Human fibroblasts expressing hTERT show remarkable karyotype stability even after exposure to ionizing radiation.

Ectopic expression of telomerase results in an immortal phenotype in various types of normal cells, including primary human fibroblasts. In addition to its role in telomere lengthening, telomerase has now been found to have various functions, including the control of DNA repair, chromatin modification, and the control of expression of genes involved in cell cycle regulation. The investigations on the long-term effects of telomerase expression in normal human fibroblast highlighted that these cells show low frequencies of chromosomal aberrations.

Telomeres and chromosomal instability.

Telomeres are distinctive structures, composed of a repetitive DNA sequence and associated proteins, which enable cells to distinguish chromosome ends from DNA double-strand breaks. Telomere alterations, caused by replication-mediated shortening, direct damage or defective telomere-associated proteins, usually generate chromosomal instability, which is observed in senescence and during the immortalization process. In cancer cells, this chromosome instability could be extended by their ability to 'repair' chromosomes and terminate in break-fusion-bridge cycles.

Telomere-driven genomic instability in cancer cells.

Telomeres, the ends of linear chromosomes, play a major role in the maintenance of genome integrity. Telomerase or alternative lengthening of telomeres (ALT) mechanisms exist in most cancer cells in order to stabilize telomere length by the addition of telomeric repeats. Telomere loss can be dramatically mutagenic.

The influence of interstitial telomeric sequences on chromosome instability in human cells.

Although most telomere repeat sequences are found at the ends of chromosomes, some telomeric repeat sequences are also found at intrachromosomal locations in mammalian cells. Several studies have found that these interstitial telomeric repeat sequences can promote chromosome instability in rodent cells, either spontaneously or following ionizing radiation. In the present study we describe the extensive cytogenetic analysis of three different human cell lines with plasmids containing telomeric repeat sequences integrated at interstitial sites.

Multiple chromosomal mechanisms generate an EWS/FLI1 or an EWS/ERG fusion gene in Ewing tumors.

The t(11;22)(q24;q12) translocation is found in about 85% of Ewing tumors and leads in all cases to an EWS/FLI1 fusion gene. In a few instances, complex translocations, involving chromosomes 11, 22 and a third chromosome or other variant translocations not involving chromosome 11 also have been reported. These rearrangements generate an active fusion gene between the EWS gene and either the human FLI1 gene or other members of the ETS gene family: the ERG gene localized in band 21q22.

Cloning and characterization of hOGG1, a human homolog of the OGG1 gene of Saccharomyces cerevisiae.

The OGG1 gene of Saccharomyces cerevisiae encodes a DNA glycosylase activity that is a functional analog of the Fpg protein from Escherichia coli and excises 7,8-dihydro-8-oxoguanine (8-oxoG) from damaged DNA. The repair of this ubiquitous kind of oxidative damage is essential to prevent mutations both in bacteria and in yeast. A human cDNA clone carrying an ORF displaying homology to the yeast protein was identified.

A 10-cM YAC contig spanning GLC1A, the primary open-angle glaucoma locus at 1q23-q25.

Primary open-angle glaucoma is a complex of ocular disorders characterized by irreversible lesions of the optic nerve, open angle of the anterior chamber of the eye and elevated intraocular pressures. GLC1A, a locus involved in one form of this disease, has been mapped to an approximately 9-cM interval within 1q23-q25, between markers D1S445 and D1S416/D1S480. A 10-cM yeast artificial chromosome (YAC) contig spanning the whole region is described.

Oncogenic conversion of a novel orphan nuclear receptor by chromosome translocation.

A recurrent t(9;22) (q22;q12) chromosome translocation has been described in extraskeletal myxoid chondrosarcoma (EMC). Fluorescent in situ hybridization experiments performed on one EMC tumour indicated that the chromosome 22 breakpoint occurred in the EWS gene. Northern blot analysis revealed an aberrant EWS transcript which is cloned by a modified RT-PCR procedure.

In situ hybridization of fluorescent probes on chromosomes, nuclei or stretched DNA: applications in physical mapping and characterization of genomic rearrangements.

During the last few years, various technologies and applications of fluorescence in situ hybridization (FISH) have been developed. Hybridization on nuclei allows an increase in the resolution of the technique. It also permits the characterization of some chromosomal abnormalities such as trisomies, monosomies or translocations in pathological cells when it is difficult to obtain metaphases.

A human homolog of the S. cerevisiae HIR1 and HIR2 transcriptional repressors cloned from the DiGeorge syndrome critical region.

The DiGeorge syndrome (DGS) is a developmental disorder affecting derivatives of the third and fourth pharyngeal pouches. DGS patients present an interstitial deletion in one of their two chromosomes 22. Cosmid DAC30 was mapped to the DGS smallest critical region.

Cloning of a balanced translocation breakpoint in the DiGeorge syndrome critical region and isolation of a novel potential adhesion receptor gene in its vicinity.

Deletions of the 22q11.2 have been associated with a wide range of developmental defects (notably DiGeorge syndrome, velocardiofacial syndrome, conotruncal anomaly face syndrome and isolated conotruncal cardiac defects) classified under the acronym CATCH 22. A DiGeorge syndrome patient bearing a balanced translocation whose breakpoint maps within the critical region has been previously described.

Interphase molecular cytogenetics of Ewing's sarcoma and peripheral neuroepithelioma t(11;22) with flanking and overlapping cosmid probes.

The translocation, t(11;22)(q24;q12), recurrently observed in Ewing's sarcoma and in peripheral neuroepithelioma has been recently cloned. The analysis of a series of ES/PNE has revealed that the chromosome 22 breakpoints are clustered in a small region of 7 kb, called EWSR1, and that those on chromosome 11 are spread over a larger region of 40-50 kb, called EWSR2. Cosmids from loci flanking or overlapping these two regions have been obtained.