Figure 7. TERRA maintains telomere integrity in ES cells.

A. DNA FISH analysis using telomeric repeat probes of ES cell metaphase spreads. Telomeric DNA, green. Metaphase chromosomes, blue. A single cell is shown in each panel. ES cells were harvested after 24-hr knockdown with TERRA LNA versus control (Scr and sense) LNAs. Arrows, loss of telomeric cap on one (heterogeneous) or both sister chromatids. Asterisks, insertion or duplication. R, ring or fused telomeres.

B. Magnified examples of each aberrant telomere phenotype. DNA FISH performed using C-rich telomeric PNA probes (TelC). Black and white images shown for the TelC and DAPI signals are merged in the color image.

C. Magnified examples of each aberrant telomere phenotype. DNA FISH performed using G-rich telomeric PNA probes (TelG). Black and white images shown for the TelG and DAPI signals are merged in the color image.

D. Quantitation of each telomeric pathology. Two biological replicates of each knockdown were averaged. Error bars are S.D. ***, P< 0.001, compared to Scr KD. P values determined by twotailed Chi-square analysis. Number of counted chromosomes is indicated.

E. Summary: The TERRA interactome revealed by combining CHIRT, iDRiP, and LNA-mediated knockdown enabled determination of TERRA function. TERRA plays a critical role in telomere function by antagonizing ATRX, controlling telomerase activity, and maintaining telomeric integrity. Chromosome image adapted from http://med.stanford.edu/content/dam/smnews/images/2015/01/telomeres.jpg.