Shelterin Components Modulate the Phase-Separation Propensity of Telomeres

Abstract

Telomeres are nucleoprotein complexes that provide a mechanism to shelter the ends of chromosomes and are essential to maintain chromosome stability in Eukaryotes. In cells, individual telomeres are bound by the six-protein shelterin complex (TRF2-hRap1-Tin2-TRF1-TPP1-Pot1) and form distinct condensed globules of finite size. Interestingly, incells that maintain their telomeres via an alternative lengthening mechanism, multiple telomeric DNAs and shelterin proteins coalesce in membrane-less promyelocytic leukemia (PML) nuclear bodies. The mechanism controlling this dual nature of telomere organization is currently not understood. Here, we propose that shelterin proteins modulate the phase-separation propensity of telomeres, leading to either the condensation of single DNA chains or the coalescence of multiple ones. In particular, we use a combination of optical and fluorescence microscopy and single-molecule experiments to quantify the contribution of shelterin components to phase-separation and condensation of single DNA chains. Our experiments reveal that TRF2 promotes phase-separation of TRF2-DNA mixtures. Even though the N-terminal basic domain and the dimerization domain of TRF2 are sufficient to promote phase-separation, the C-terminal DNA-binding domain imparts specificity for telomeric DNA repeats. Moreover, addition of shelterin components, such as hRap1, provides a fine-tuning of the properties of the resulting dense and de-mixed phase.