Long interstitial (TTAGGG)n arrays do not colocalize with repressive chromatin modifications in Chinese hamster cells

Interstitial tandem (TTAGGG)n repeats (ITRs) in Chinese hamster (CH) cells are mostly arranged into the long (>100kb) continuous pericentric arrays which do not contain HindIII sites (Faravelli M, Azzalin CM, Bertoni L, Chernova O, Attolini C, Mondello C, et al. Molecular organization of internal telomeric sequences in Chinese hamster chromosomes, Gene 2002;283,11-16), are free of protein-coding genes, and can formally be classified as heterochromatin. ITRs dynamically interact with (TTAGGG)n-binding protein TRF1 and can be visualized using Green Fluorescent Protein (GFP)-tagged TRF1. Here we examined whether mobility of GFP-TRF1 associated with ITRs in CH cells is affected by inhibitors of transcription and whether ITRs colocalize with known repressive chromatin modifications hallmarked by histone H3 trimethylated at lysine-9 (H3K9m3) or lysine-27 (H3K27m3). We found that GFP-TRF1 bodies do not colocalize in the nuclei of V79 cells with H3K9m3 or H3K27m3 indicating that ITRs do not represent typical constitutive or facultative heterochromatin. Mobility of ITR-bound GFP-TRF1 is suppressed by inhibitors of transcription consistent with the view that TRF1 is exchanged during transcription of ITRs. However, GFP-TRF1 bodies do not colocalize with nuclear hubs of intensive transcription detected through in vivo incorporation of 5-bromouridine triphosphate. Using real-time PCR, we also examined transcription of unique sequences adjacent to (TTAGGG)n arrays in CH genome and found that they are transcribed, indicating that these arrays do not generally inhibit transcription in cis. Together, our results suggest that ITRs represent a special kind of moderately transcribed heterochromatin which possible function remains to be established.