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J Biol Chem . 2020 Jul 3;

Role of folding kinetics of secondary structures in telomeric G-overhangs in the regulation of telomere maintenance in Saccharomyces cerevisiae

Liron Malki, Ofer Sarig, Nicole Cesarato, Janan Mohamad, Talia Canter, Sari Assaf, Mor Pavlovsky, Dan Vodo, Yossi Anis, Ofer Bihari, Kiril Malovitski, Andrea Gat, Holger Thiele, Bethany E Perez White, Liat Samuelov, Arti Nanda, Amy S Paller, Regina C Betz, Eli Sprecher, Xueyi Shen, David M Howard, Mark J Adams, W David Hill, Toni-Kim Clarke, Major Depressive Disorder Working Group of the Psychiatric Genomics Consortium, Ian J Deary, Heather C Whalley, Andrew M McIntosh, Samuel M Gonçalves, Cláudio Duarte-Oliveira, Cláudia F Campos, Vishukumar Aimanianda, Rob Ter Horst, Luis Leite, Toine Mercier, Paulo Pereira, Miguel Fernández-García, Daniela Antunes, Cláudia S Rodrigues, Catarina Barbosa-Matos, Joana Gaifem, Inês Mesquita, António Marques, Nuno S Osório, Egídio Torrado, Fernando Rodrigues, Sandra Costa, Leo Ab Joosten, Katrien Lagrou, Johan Maertens, João F Lacerda, António Campos Jr, Gordon D Brown, Axel A Brakhage, Coral Barbas, Ricardo Silvestre, Frank L van de Veerdonk, Georgios Chamilos, Mihai G Netea, Jean-Paul Latgé, Cristina Cunha, Agostinho Carvalho, Katarina Jurikova, Martin Gajarsky, Mona Hajikazemi, Jozef Nosek, Katarina Prochazkova, Katrin Paeschke, Lukas Trantirek, Lubomir Tomaska

The ends of eukaryotic chromosomes typically contain a 3' ssDNA G-rich protrusion (G-overhang). This overhang must be protected against detrimental activities of nucleases and of the DNA damage response machinery and participates in the regulation of telomerase, a ribonucleoprotein complex that maintains telomere integrity. These functions are mediated by DNA-binding proteins, such as Cdc13 in Saccharomyces cerevisiae, and the propensity of G-rich sequences to form various non-B DNA structures. Using CD and NMR spectroscopies, we show here that G-overhangs of S. cerevisiae form distinct Hoogsteen pairing-based secondary structures, depending on their length. Whereas short telomeric oligonucleotides form a G-hairpin, their longer counterparts form parallel and/or antiparallel G-quadruplexes (G4s). Regardless of their topologies, non-B DNA structures exhibited impaired binding to Cdc13 in vitro as demonstrated by electrophoretic mobility shift assays. Importantly, whereas G4 structures formed relatively quickly, G-hairpins folded extremely slowly, indicating that short G-overhangs, which are typical for most of the cell cycle, are present predominantly as single-stranded oligonucleotides and are suitable substrates for Cdc13. Using ChIP, we show that the occurrence of G4 structures peaks at the late S phase, thus correlating with the accumulation of long G-overhangs. We present a model of how time- and length-dependent formation of non-B DNA structures at chromosomal termini participates in telomere maintenance.

PMID: 32385108