Abstract
Many tumors maintain chromosome-ends through a telomerase-independent, DNA-templated mechanism called alternative lengthening of telomeres (ALT). While ALT occurs in only a subset of tumors, it is strongly associated with mutations in the genes ATRX and DAXX, which encode components of an H3.3 histone chaperone complex. The role of ATRX and DAXX mutations in potentiating the mechanism of ALT remains incompletely understood. Here we characterize an osteosarcoma cell line, G292, with wild-type ATRX but a unique chromosome translocation resulting in loss of DAXX function. While ATRX and DAXX form a complex in G292, this complex fails to localize to nuclear PML bodies. We demonstrate that introduction of wild type DAXX suppresses the ALT phenotype and restores the localization of ATRX/DAXX to PML bodies. Using an inducible system, we show that ALT-associated PML bodies are disrupted rapidly following DAXX induction and that ALT is again restored following withdrawal of DAXX.
Highlights
The unlimited proliferative capacity of cancer cells is closely linked to maintenance of telomeres
While mutations in ATRX and DAXX are both strongly associated with alternative lengthening of telomeres (ALT), prior mechanistic studies have focused primarily on ATRX
We here describe a DAXX mutation in an ALT positive osteosarcoma cell line, G292, and demonstrate that maintenance of ALT is dependent on ongoing DAXX deficiency
Summary
The unlimited proliferative capacity of cancer cells is closely linked to maintenance of telomeres. Active mechanisms must both counteract the shortening of telomeres with each cell cycle (the end-replication problem) and prevent telomere ends from inducing double strand break DNA damage repair pathways (the end-protection problem). When complexed with ATRX, DAXX directly binds H3.3 and is required for its deposition at telomeric regions, but whether this is the sole function responsible for DAXX suppression of ALT remains unclear[10]. Upon reintroduction of full length DAXX in G292, telomere maintenance by ALT is rapidly suppressed, resulting in loss of C-circles and ALT-associated PML bodies (APBs), demonstrating that continued DAXX deficiency is essential to maintain ALT in this system. We conclude that DAXX is required for correct ATRX localization and suggest that G292-iDAXX is a useful model system for study of the ALT mechanism
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