Poster abstracts
Poster number 158 submitted by Ruwani Madushika Dalath
TERRA visualization with gamma PNA probes reveals a role of APB formation in localizing TERRA to ALT telomeres
Ruwani Madushika Dalath (Department of Biological Sciences, Carnegie Mellon University), Mackenzie Riley (Department of Chemistry, Carnegie Mellon University), Meng Xu, Rongwei Zhao (Department of Biological Sciences, Carnegie Mellon University), Nisha Hasija, Colin Martin (Department of Chemistry, Carnegie Mellon University), Bruce Armitage* (Department of Chemistry, Carnegie Mellon University), Huaiying Zhang* (Department of Biological Sciences, Carnegie Mellon University)
Abstract:
Alternative lengthening of telomeres (ALT) is a homology-directed repair mechanism utilized by a subset of aggressive cancers that maintains the telomere length independent of telomerase activity(Bryan et al., 1997). One of the key features of ALT cancer cells is the upregulation of a long noncoding RNA called Telomeric Repeat Containing RNA (TERRA) that contributes to both DNA damage and repair at telomeres to facilitate the ALT pathway(Azzalin et al., 2007a). Although TERRA is known to act in trans, the mechanism that governs its localization to ALT telomeres remains elusive. To address this, we developed a novel fluorescence in situ hybridization (FISH) protocol using gamma PNA, a second-generation peptide nucleic acid probe, engineered for high-efficiency TERRA detection with strong selectivity against telomeric DNA. This approach allowed us to show that TERRA localization to ALT telomeres depends on the formation of ALT-associated PML bodies (APBs). Disruption of APBs, using both genetic knockdowns and chemical inhibitors, significantly reduces TERRA accumulation at telomeres. Conversely, enforced APB formation through a DNA damage-independent chemical dimerization system is sufficient to enhance TERRA localization to telomeres. Collectively, our study introduces an improved method for TERRA detection and uncovers a mechanistic link between APB formation and TERRA recruitment at ALT telomeres, offering new insights into the regulation of ALT activity in cancers.
References:
1. Bryan, T. M., Englezou, A., Dalla-Pozza, L., Dunham, M. A., & Reddel, R. R. (1997). Evidence for an alternative mechanism for maintaining telomere length in human tumors and tumor-derived cell lines. Nature Medicine, 3(11), 1271–1274. https://doi.org/10.1038/nm1197-1271
2. Azzalin, C. M., Reichenbach, P., Khoriauli, L., Giulotto, E., & Lingner, J. (2007a). Telomeric Repeat–Containing RNA and RNA Surveillance Factors at Mammalian Chromosome Ends. Science, 318(5851), 798–801. https://doi.org/10.1126/science.1147182
Keywords: TERRA, PNA, PML