How transcription proceeds in a large artificial heterochromatin in human cells.

Heterochromatin is critical for genome integrity, and recent studies have suggested the importance of transcription in heterochromatin for maintaining its silent state. We previously developed a method to generate a large homogeneously staining region (HSR) composed of tandem plasmid sequences in human cells that showed typical heterochromatin characteristics. In this study, we examined transcription in the HSR. We found that transcription of genes downstream to no-inducible SRalpha promoter was restricted to a few specific points inside the large HSR domain. Furthermore, the HSR localized to either to the surface or to the interior of the nucleolus, where it was more actively transcribed. The perinucleolar or intranucleolar locations were biased to late or early S-phase, and the location depended on either RNA polymerase II/III or I transcription, respectively. Strong activation of the inducible TRE promoter resulted in the reversible loosening of the HSR domain and the appearance of transcripts downstream of not only the TRE promoters, but also the SRalpha promoters. During this process, detection of HP1alpha or H3K9Me3 suggested that transcription was activated at many specific points dispersed inside large heterochromatin. The transcriptional rules obtained from studying artificial heterochromatin should be useful for understanding natural heterochromatin.