Association of DNA topoisomerase I and RNA polymerase I: a possible role for topoisomerase I in ribosomal gene transcription

Abstract

Abstract. RNA polymerase I preparations purified from a rat hepatoma contained DNA topoisomerase activity. The DNA topoisomerase associated with the polymerase had an Mr of 110000, required Mg2+ but not ATP, and was recognized by anti-topoisomerase I antibodies. When added to RNA polymerase I preparations containing topoisomerase activity, anti-topoisomerase I antibodies were able to inhibit the DNA relaxing activity of the preparation as well as RNA synthesis in vitro. RNA polymerase II prepared by analogous procedures did not contain topoisomerase activity and was not recognized by the antibodies. The topoisomerase I: polymerase I complex was reversibly dissociated by column chromatography on Sephacryl S200 in the presence of 0.25 M (NH 4 hS04. Topoisomerase I was immunolocalized in the transcriptionally active ribosomal gene complex containing RNA polymerase I in situ. These data indicate that topoisomerase I and RNA polymerase I are tightly complexed both in vivo and in vitro, and suggest a role for DNA topoisomerase I in the transcription of ribosomal genes.

Identification of a mammalian RNA polymerase I holoenzyme containing components of the DNA repair/replication system

“…In addition, TopoI co-localizes with RPI in immunohistochemical assays and co-purifies with RPI during biochemical purification of the enzyme (23,24). These observations suggest a specific role for TopoI in rDNA transcription.…”

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

“…12. Immunofluorescence microscopicallocalization of DNA topoisomerase I on cultured rat cells (for details see Rose et al, 1988). Besides a strong nucleolar signal, a diffuse staining of the whole nucleoplasm is noted ((a' ) the corresponding phase contrast micrograph is shown in (a)).…”

“…10; Scheer and Rose, 1984) and topoisomerase I ( Fig. 12; Rose et al, 1988; see also Guldner et al, 1986) remain bound to the NORs during mitosis. Apparently rRNA genes together with the essential transcriptional machinery (perhaps in the form of 'frozen' transcriptional complexes) are transmitted from one cell generation to the next one, thereby ensuring a rapid reactivation of rRNA transcription following mitosis.…”

“…A number of studies have indicated that this enzyme which interconverts topological isomers of DNA by breaking and resealing one DNA strand, is required for proper transcription ofrRNA genes (e.g. Garg et al, 1987;Egyhazi and Durban, 1987;Rose et al, 1988;Zhang et al, 1988;Kim and Wang, 1989). Topoisomerase I has been shown to be associated with RNA polymerase I and is located within the transcribed region of the rRNA genes but absent from the spacer region (Zhang et al, 1988).…”

“…Immunogold electron microscopical localization of DNA to po isomerase I (preembedding protocol). A frozen section (5 Jlm) of rat liver was incubated with affinity purified antibodies to topoisomerase I (see Rose et al, 1988) followed by secondary antibodies coupled to 5 nm gold particles. After fixation and Epon embedding, ultrathin sections were cut.…”