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

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.

Clinical features of ATRX or DAXX mutated neuroblastoma

“…Interestingly, all ALT activated NBLs had these alterations. Therefore, loss of function due to ATRX or DAXX alterations may contribute to ALT activation in NBLs, and it is possible that this is a direct effect of defects of histone H3.3 deposition at telomeres [26,27]. Although ATRX is located in chromosome X, both males and females were found to have ATRX mutations, consistent with previous work [10].…”

Genetic Inactivation of ATRX Leads to a Decrease in the Amount of Telomeric Cohesin and Level of Telomere Transcription in Human Glioma Cells

“…As illustrated above, telomerasepositive cultured cells, just like telomerase-positive tumors, exhibited nuclear ATRX, as seen by immunofluorescence, whereas ALT-positive tumors and cultured cells exhibited total loss of nuclear ATRX associated with mutations in ATRX (10,24). This is also illustrated in Fig.…”

“…Thus, in several clinical studies, mutations in ATRX were found to be tightly associated with the ALT pathway in various types of tumors (10,11,(20)(21)(22)(23). This correlation has also been observed in cultured cells (24,25). To understand how genetic inactivation of ATRX might interfere with telomeric pathways in tumor cells and, in particular, favor the ALT pathway, we used glioma cells in culture because gliomas have been reported to be among the 5 to 15% of tumors that exhibited ALT-maintained telomeres (9).…”

Assembly of telomeric chromatin to create ALTernative endings

“…The same tumors also invariably lacked detectable expression of ATRX and often DAXX [14]. This remarkable correlation between ALT and deregulated ATRX expression was further substantiated by the absence of detectable ATRX protein in 22 ALT cell lines and hybrid cell lines that displayed ALT activity [20,24]. Therefore, ATRX has emerged as the strongest candidate identified to date for the suppression of ALT.…”