Expression of 7q TERRA. (A) Distribution of TERRA reads from 7q, 12q, 29 bp-like-containing subtelomeres and from repeat-devoid subtelomeres in the indicated cell lines. Only unique reads were considered for the analysis. (B) Northern blot hybridizations of total and nuclear RNA from U2OS, HeLa, and HEK293T cells using probes for the 7q/12q TERRA or UUAGGG repeats (total TERRA). The positions of 28S and 18S ribosomal RNAs are indicated on the right. GAPDH and U6 were used as loading controls. RNaseA treatments were used to assure that signals derived from RNA and not genomic DNA contaminations. (C) Total and nuclear RNA from U2OS cells was incubated with RNaseH in the presence of oligonucleotides complementary to a sequence from 7q/12q TERRA or UUAGGG repeats (Tel). Oligonucleotides complementary to a GAPDH sequence were also included as indicated and served as a control for the RNaseH treatment. Products were analyzed by northern blotting as in B. (D) U2OS cells were treated with actinomycin D for the indicated times. Total RNA was extracted and analyzed by northern blotting as in C. The long-lived GAPDH mRNA was used as a loading control. Half-lives and R2 values were calculated using a scatter plot analysis after the 7q/12q or the TERRA signals were normalized to the GAPDH signals. Data points are from two independent experiments and the values for time 0 are set to 1.
