Summary Mammalian telomeres are associated with shelterin, the telomere specific protein complex that solves the end-protection problem. The telomeric shelterin binding sites, TTAGGG repeats, are maintained by telomerase, which solves the end-replication problem. We report that the TTAGGG repeat arrays of human and mouse telomeres challenge the DNA replication machinery, giving rise to replication-dependent defects that resemble those of the aphidicolin-induced common fragile sites. Conditional gene deletion experiments showed that efficient duplication of telomeric DNA requires the shelterin component TRF1. In the absence of TRF1, telomeres activate the ATR kinase in S phase and show a fragile site phenotype in metaphase. SMARD showed that TRF1 promotes efficient replication of TTAGGG repeats and prevents fork stalling. Two helicases that can remove G4 DNA structures, BLM and RTEL1, were required to repress the fragile telomere phenotype. These results identify a second telomere replication problem that is solved by the shelterin component TRF1.

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SynopsisData on the decrease of the DNA melting temperature T , with the salt concentration i M are report,ed and discussed. The electrost.atic free energy change in the helix-coil transition, AF,, is related to the potential, $, which represents the electrostatic repulsion between the phosplmte charges; $ is calculated as a function of M and of the dist,ances bet,ween the charges of the two strands. The Debye-Huckel approximation is shown to overest,imate $. I t is suggested that the high local concentration of the counterione in the immediate vicinity of the fixed charges screen these charges from interacting with other fixed charges, to the extent, that the system behaves as if the fixed ions carry :L redriced charge. The notioii of a redriced charge represents in a single parameter the deviation of the 1)ehye-Hiickel approximation from the true potential. A plot, of T , versris AHu is Calculated from the slope and Our calculations support t,he hypothesis that the change of T , with salt concentration is due to changes in the screened iriteractioris between the fixed phosphate charges. In analyzing the results of these caldations, we are able on the one hand t,o indicate some of the limitations of the theoretical model and, on the other hand, draw some conchisions about the order of magnitude of the nonelectrostat,ic interaction energy of format,ion of the double helix. ~~ -~~ gives a straight line as predict,ed. erit with experimentally determined vahies.

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