The RNA 3′ cleavage factors CstF 64 kDa and CPSF 100 kDa are concentrated in nuclear domains closely associated with coiled bodies and newly synthesized RNA

Wouter Schul, Bastiaan Groenhout, Karel Koberna, Yoshio Takagaki, Andreas Jenny, Erik M M Manders, Ivan Raška, Roel Van Driel, Luitzen De Jong

Research output: Contribution to journalArticle

77 Citations (Scopus)

Abstract

The cleavage stimulation factor (CstF), and the cleavage and polyadenylation specificity factor (CPSF) are necessary for 3′-terminal processing of polyadenylated mRNAs. To study the distribution of 3′ cleavage factors in the nuclei of human T24 cells, monoclonal antibodies against the CstF 64 kDa subunit and against the CPSF 100 kDa subunit were used for immunofluorescent labelling. CstF 64 kDa and CPSF 100 kDa were distributed in a fibrogranular pattern in the nucleoplasm and, in addition, were concentrated in 1-4 bright foci. Double immunofluorescence labelling experiments revealed that the foci either overlapped with, or resided next to, a coiled body. Inhibition of transcription with α-amanitin or 5,6-dichloro-β-D-ribofuranosyl-benzimidazole (DRB) resulted in the complete co-localization of coiled bodies and foci containing 3′ cleavage factors. Electron microscopy on immunogold double-labelled cells revealed that the foci represent compact spherical fibrous structures, we named 'cleavage bodies', intimately associated with coiled bodies. We found that ∼20% of the cleavage bodies contained a high concentration of newly synthesized RNA, whereas coiled bodies were devoid of nascent RNA. Our results suggest that the cleavage bodies that contain RNA are those that are adjacent to a coiled body. These findings reveal a dynamic and transcription-dependent interaction between different subnuclear domains, and suggest a relationship between coiled bodies and specific transcripts.

Original languageEnglish (US)
Pages (from-to)2883-2892
Number of pages10
JournalEMBO Journal
Volume15
Issue number11
StatePublished - 1996
Externally publishedYes

Fingerprint

Cleavage Stimulation Factor
Cleavage And Polyadenylation Specificity Factor
Coiled Bodies
RNA Cleavage
RNA
Transcription
Labeling
Amanitins
Electron microscopy
Monoclonal Antibodies
Messenger RNA
Fluorescent Antibody Technique
Electron Microscopy
Processing
Experiments

Keywords

  • Cleavage factors
  • Coiled bodies
  • Nuclear bodies
  • PML

ASJC Scopus subject areas

  • Cell Biology
  • Genetics

Cite this

The RNA 3′ cleavage factors CstF 64 kDa and CPSF 100 kDa are concentrated in nuclear domains closely associated with coiled bodies and newly synthesized RNA. / Schul, Wouter; Groenhout, Bastiaan; Koberna, Karel; Takagaki, Yoshio; Jenny, Andreas; Manders, Erik M M; Raška, Ivan; Van Driel, Roel; De Jong, Luitzen.

In: EMBO Journal, Vol. 15, No. 11, 1996, p. 2883-2892.

Research output: Contribution to journalArticle

Schul, W, Groenhout, B, Koberna, K, Takagaki, Y, Jenny, A, Manders, EMM, Raška, I, Van Driel, R & De Jong, L 1996, 'The RNA 3′ cleavage factors CstF 64 kDa and CPSF 100 kDa are concentrated in nuclear domains closely associated with coiled bodies and newly synthesized RNA', EMBO Journal, vol. 15, no. 11, pp. 2883-2892.
Schul, Wouter ; Groenhout, Bastiaan ; Koberna, Karel ; Takagaki, Yoshio ; Jenny, Andreas ; Manders, Erik M M ; Raška, Ivan ; Van Driel, Roel ; De Jong, Luitzen. / The RNA 3′ cleavage factors CstF 64 kDa and CPSF 100 kDa are concentrated in nuclear domains closely associated with coiled bodies and newly synthesized RNA. In: EMBO Journal. 1996 ; Vol. 15, No. 11. pp. 2883-2892.
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