Oral Presentation 45th Lorne Genome Conference 2024

Nuclear F-actin and the DNA damage response regulate telomerase recruitment in human cells (#22)

Tracy M Bryan 1 , Ashley Harman 1 , Anthony Cesare 1 , Noa Lamm 1
  1. Children's Medical Research Institute, Westmead, NSW, Australia

To proliferate indefinitely, cancer cells require a mechanism to maintain the ends of their chromosomes (telomeres). In ~85% of cancer cells, telomeres are elongated by the ribonucleoprotein telomerase. While the biochemical mechanism of telomerase is reasonably well characterised, less is known about how it is regulated, including its recruitment to telomeres.  We have previously linked the DNA damage response (DDR) to telomerase recruitment, specifically via the regulatory kinase ATR, which coordinates the cellular response to replication stress1. Furthermore, nuclear filamentous actin (F-actin) is important for DNA replication dynamics following replication stress2. Given the convergence of these phenomena, we hypothesise that there is controlled interplay between DNA replication, the DDR, and telomere maintenance, which together result in cancer cell immortalisation. We aimed to elucidate whether actin polymerisation is important for telomerase recruitment, and how F-actin brings telomeres and telomerase together.

Here we have demonstrated, via fluorescence in situ hybridisation (FISH), that nuclear F-actin is important for telomerase recruitment, as inhibition of actin polymerisation decreases the presence of telomerase at telomeres. This occurs in an ATR and mTOR-dependent manner, employs other regulators of actin structure and function (e.g. WASP and ARP2/3) and involves myosin. We also explored whether the nuclear F-actin network serves as tracks to bring telomeres and telomerase together. Using FISH we found that telomeres interacting with telomerase are located closer to F-actin than are other telomeres or unbound telomerase. We also examined this via live-cell microscopy and observed that telomerase recruitment to telomeres largely occurs on or adjacent to F-actin filaments.

These results demonstrate that telomerase is recruited to telomeres by F-actin via activation of a DDR. This supports a model of cancer cell immortalisation in which DNA replication and the DDR are crucial factors which dictate the timing and occurrence of telomere maintenance.

  1. Tong, A.S., Stern, J.L., Sfeir, A., Kartawinata, M., de Lange, T., Zhu, X.D., and Bryan, T.M. (2015). ATM and ATR Signaling Regulate the Recruitment of Human Telomerase to Telomeres. Cell reports 13, 1633-1646.
  2. Lamm, N., Read, M.N., Nobis, M., Van Ly, D., Page, S.G., Masamsetti, V.P., Timpson, P., Biro, M., and Cesare, A.J. (2020). Nuclear F-actin counteracts nuclear deformation and promotes fork repair during replication stress. Nat Cell Biol 22, 1460-1470.