Loss of ATRX, Genome Instability, and an Altered DNA Damage Response Are Hallmarks of the Alternative Lengthening of Telomeres Pathway

Abstract

The Alternative Lengthening of Telomeres (ALT) pathway is a telomerase-independent pathway for telomere maintenance that is active in a significant subset of human cancers and in vitro immortalized cell lines. ALT is thought to involve templated extension of telomeres through homologous recombination, but the genetic or epigenetic changes that unleash ALT are not known. Recently, mutations in the ATRX/DAXX chromatin remodeling complex and histone H3.3 were found to correlate with features of ALT in pancreatic neuroendocrine cancers, pediatric glioblastomas, and other tumors of the central nervous system, suggesting that these mutations might contribute to the activation of the ALT pathway in these cancers. We have taken a comprehensive approach to deciphering ALT by applying genomic, molecular biological, and cell biological approaches to a panel of 22 ALT cell lines, including cell lines derived in vitro. Here we show that loss of ATRX protein and mutations in the ATRX gene are hallmarks of ALT–immortalized cell lines. In addition, ALT is associated with extensive genome rearrangements, marked micronucleation, defects in the G2/M checkpoint, and altered double-strand break (DSB) repair. These attributes will facilitate the diagnosis and treatment of ALT positive human cancers.

Author Summary

Telomeres, the protective elements at the ends of chromosomes, need to be maintained for cells to proliferate indefinitely. In many human cancers, the telomeric DNA is replenished by telomerase. However, a second pathway for telomere maintenance, referred to as the ALT pathway, has increasingly been recognized in human cancers. The genetic basis for activation of ALT is not known, but recent data have implicated a chromatin remodeling complex (ATRX/DAXX) and the histone variant H3.3 as players in the repression of ALT. We have examined a large panel of ALT cell lines for their genetic and cell biological features and found that loss of ATRX is a common event in the genesis of ALT lines. In addition, we document that ALT cell lines frequently undergo chromosomal changes and are impaired in their ability to detect and repair damage in their DNA. These hallmarks of ALT are expected to facilitate the detection of ALT–type tumors in the clinic and may lead to ALT–specific treatments.