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. Author manuscript; available in PMC: 2022 Apr 24.
Published in final edited form as: Dev Cell. 2022 Jan 24;57(2):277–290.e9. doi: 10.1016/j.devcel.2021.12.017

Figure 2: Telomeric DNA acts as an oligomerizing scaffold to promote TRF1 and TRF2-mediated condensation.

Figure 2:

A. Example images of Cy3-TRF2 in the presence and absence of 8ds3ss telomeric DNA. B. The total volume of TRF2 condensates settled per micron squared area on the coverslip in the presence or absence of 2.5 μM 8ds3ss (mean ± SD, n = 30 with three technical replicates). Linear fit (solid line) reveals csat (± SE). C. The minimum TRF2 concentration that exhibits phase separation with a variable number of dsTEL repeats per DNA substrate. The total concentration of dsTEL tracts was fixed to 20 μM. D. TRF2 has a reentrant phase behavior as a function of 8ds3ss concentration. E. The total volume of TRF1 or TRF2 condensates settled per micron squared area on the coverslip under variable 8ds3ss concentration (mean ± SD, n = 20 with two technical replicates). F. Example images of Cy3-TRF1 in the presence and absence of 8ds3ss. G. The total volume of TRF1 condensates settled per micron squared area on the coverslip in the presence or absence of 2.5 μM 8ds3ss (mean ± SD, n = 30 with three technical replicates). Linear fits (solid lines) reveal csat (± SE). H. An increase in 8ds3ss concentration inhibits TRF1 phase separation. I. Fusion of TRF2 (22 μM, left) and TRF1 (44 μM, right) droplets formed in the presence of 2.5 μM 8ds3ss. J. U2OS expressing sspB-mCherry-TRF2. TRF corelets were locally activated at a single telomere (solid circle) or away from any telomere (dotted circle). K. (Top) Example images show first and last frames of locally activated TRF1, TRF1A75P, and TRF2 at telomeres (left; n = 3, 11, 10 cells analyzed, respectively) and away from telomeres (right; n = 3, 3, 4 cells analyzed, respectively) in U2OS cells. (Bottom) Quantification of change in intensity upon local activation, at and away from existing telomeres for WT TRF1, TRF1A75P, and WT TRF2. The intensity of each locally activated telomere or region was normalized to the average intensity of all other telomeres within the same activated cell. P-values were quantified by one-way ANOVA with multiple comparisons. See also Figures S2 and S3, Table S1, and Movie S4.