Abstract
We report on the effect of different doses and times of incubation of the cytostatic drug actinomycin D (AMD) on nucleolar morphology, rRNA gene transcription and rDNA gene localization usingin situ hybridization and the immunocytochemical detection of the human upstream binding factor (UBF) at the electron microscopic level in HeLa cells. Low doses of AMD (0.001 μg/ml, 30 min) selectively block rRNA gene transcription but alter neither nucleolar morphology nor the localization of rDNA with respect to the nucleolar components. Treatment with high doses of AMD (0.05 μg/ml, 1 h) resulted in a retraction of the rDNA out of the nucleolus in addition to the well-known blocking or rDNA transcription, total nuclear transcription and nucleolar segregation. Under these conditions accumulations of rDNA were found in patches of chromatin at the nucleolar periphery. We conclude that the blocking of rRNA gene transcription and the changes in nucleolar morphology, both induced by AMD at different doses, are independent phenomena.
Similar content being viewed by others
References
Busch H, Smetana K (1970) Effects of drugs and other agents on the nucleolus. In: Busch H, Smetana K, ed.The Nucleolus. New York: Academic Press, pp 472–510.
Dadoune JP, Siffroi JP, Alfonsi MF (1994) Ultrastructural localization of rDNA and rRNA by in situ hybridization in the nucleolus of human spermatids.Cell Tissue Res 278: 611–616.
Hozák P, Schöfer C, Sylvester J, Wachtler F (1993) A study on nucleolar DNA — isolation of DNA from fibrillar components and ultrastructural localization of different DNA probes.J Cell Sci 104: 1199–1205.
Jimenez-Garcia LF, Segura-Valdez ML, Ochs RLet al. (1993) Electron microscopic localization of ribosomal DNA in rat liver nucleoli by nonisotopic in situ hybridization.Exp Cell Res 207: 220–225.
Jordan P, Carmo-Fonseca M (1995) Evidence that the preinitiation complex for RNA polymerase I transcription remains permanently assembled in vivo. ECBO Meeting 1995. Stuttgart: Wissenschaftliche Verlagsgesellschaft, Supplement Abstract, p 29.
Learned RM, Learned TK, Haltiner MM, Tjian RT (1986) Human rRNA transcription is modulated by the coordinate binding of two factors to an upstream control element.Cell 45: 847–857.
Mosgöller W, Schöfer C, Derenzini Met al. (1993) Distribution of DNA in human Sertoli cell nucleoli.J Histochem Cytochem 41: 1487–1493.
Perry RP, Kelley DE (1970) Inhibition of RNA synthesis by actinomycin D: characteristic dose-response of different RNA species.J Cell Physiol 76: 127–139
Puvion-Dutilleul F, Bachellerie JP, Puvion E (1991) Nucleolar organization of HeLa cells as studied by in situ hybridization.Chromosoma 100: 395–409.
Puvion-Dutilleul F, Mazan S, Nicoloso Met al. (1992) Alterations of nucleolar ultrastructure and ribosome biogenesis by actinomycin-D — implications for U3-snRNP function.Eur J Cell Biol 58: 149–162.
Rodrigo RM, Rendon MC, Torreblanca J, Garcia-Herdugo G, Moreno FJ (1992) Characterization and immunolocalization of RNA polymerase I transcription factor UBF with anti-NOR serum in protozoa, higher plant and vertebrate cells.J Cell Sci 103: 1053–1063.
Roussel P, André C, Masson G, Géraud G, Hernandez-Verdun D (1993) Localization of the RNA polymerase I transcription factor hUBF during the cell cycle.J Cell Sci 104: 327–337.
Sachs L (1978)Angewandte Statistik. Statistische Methoden und ihre Anwendungen. Berlin: Springer.
Schoefl GI (1964) The effect of actinomycin D on the fine structure of nucleoli.J Ultrastruct Res 10: 224–243.
Smith SD, Oriahi E, Lowe Det al. (1990a) Characterization of factors that direct transcription of rat ribosomal DNA.Mol Cell Biol 10: 3105–3116.
Smith SD, Oriahi E, Yang-Yen HFet al. (1990b) Interaction of RNA polymerase I transcription factors with a promoter in the nontranscribed spacer of rat ribosomal DNA.Nucleic Acids Res 18: 1677–1685.
Stahl A, Wachtler F, Hartung Met al. (1991) Nucleoli, nucleolar chromosomes and ribosomal genes in the human spermatocyte.Chromosoma 101: 231–244.
Sylvester JE, Whiteman DA, Podolsky Ret al. (1986) The human ribosomal RNA genes: structure and organization of the complete repeating unit.Hum Genet 73: 193–198.
Thiry M, Thiry-Blaise L (1989) In situ hybridization at the electron microscopic level: an improved method for the precise localization of ribosomal DNA and RNA.Eur J Cell Biol 50: 235–243.
Tres LL (1975) Nucleolar RNA synthesis of meiotic prophase spermatocytes in the human testis.Chromosoma 53: 141–151.
Wachtler F, Popp W, Schwarzacher HG (1987) Structural changes in nucleoli during inhibition of protein- and RNA-biosynthesis.Cell Tissue Res 247: 583–589.
Wachtler F, Hartung M, Devictor Met al. (1989) Ribosomal DNA is located and transcribed in the dense fibrillar component of human Sertoli cell nucleoli.Exp Cell Res 184: 61–71.
Wachtler F, Mosgöller W, Schwarzacher HG (1990) Electron microscopic in situ hybridization and autoradiography: Localization and transcription of rDNA in human lymphocyte nucleoli.Exp Cell Res 187: 346–348.
Wachtler F, Stahl A, Sylvester Jet al. (1991) Localization of rDNA in nucleoli of human cells as revealed by high resolution in situ hybridisation.Nucleus 34: 59–73.
Wachtler F, Schöfer C, Mosgöller Wet al. (1992) Human ribosomal RNA gene repeats are localized in the dense fibrillar component of nucleoli: light and electron microscopic in situ hybridization in human Sertoli cells.Exp Cell Res 198: 135–143.
Zatsepina OV, Voit R, Grummt Iet al. (1993) The RNA polymerase I-specific transcription initiation factor UBF is associated with transcriptionally active and inactive ribosomal genes.Chromosoma 102: 599–611.
Author information
Authors and Affiliations
Corresponding author
Additional information
This paper is dedicated to Professor Doctor Dr.h.c. Hans Georg Schwarzacher on the occasion of his becoming Professor Emeritus.
accepted for publication by J. S. (Pat) Heslop-Harrison
Rights and permissions
About this article
Cite this article
Schöfer, C., Weipoltshammer, K., Almeder, M. et al. Redistribution of ribosomal DNA after blocking of transcription induced by actinomycin D. Chromosome Res 4, 384–391 (1996). https://doi.org/10.1007/BF02257274
Received:
Revised:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02257274