Abstract
After microinjection of antibodies against RNA polymerase I into the nuclei of cultured rat kangaroo (PtK2) and rat (RVF-SMC) cells alterations in nucleolar structure and composition were observed. These were detected by electron microscopy and double-label immunofluorescence microscopy using antibodies to proteins representative of the three major components of the nucleolus. The microinjected antibodies produced a progressive loss of the material of the dense fibrillar component (DFC) from the nucleoli which, at 4 h after injection, were transformed into bodies with purely granular component (GC) structure with attached fibrillar centers (FCs). Concomitantly, numerous extranucleolar aggregates appeared in the nucleoplasm which morphologically resembled fragments of the DFC and contained a protein (fibrillarin) diagnostic for this nucleolar structure. These observations indicate that the topological distribution of the material constituting the DFC can be experimentally influenced in interphase cells, apparently by modulating the transcriptional activity of the rRNA genes. These effects are different from nucleolar lesions induced by inhibitory drugs such as actinomycin D-dependent “nucleolar segregation”. The structural alterations induced by antibodies to RNA polymerase I resemble, however, the initial events of nucleolar disintegration during mitotic prophase.
Similar content being viewed by others
Abbreviations
- FC :
-
Fibrillar center
- DFC :
-
dense fibrillar component
- GC :
-
granular component
- AMD :
-
actinomycin D
References
Ansorge W (1982) Improved system for capillary microinjection into living cells. Exp Cell Res 140:31–37
Benavente R, Krohne G (1985) Change of karyoskeleton during spermatogenesis of Xenopus: expression of lamin LIV, a nuclear lamina protein specific for the male germ line. Proc Natl Acad Sci USA 82:6176–6180
Benavente R, Krohne G (1986) Involvement of nuclear lamins in postmitotic reorganization of chromatin as demonstrated by microinjection of lamin antibodies. J Cell Biol 103:1847–1854
Benavente R, Krohne G, Franke WW (1985) Cell type-specific expression of nuclear lamina proteins during development of Xenopus laevis. Cell 41:177–190
Benavente R, Rose KM, Reimer G, Hügle-Dörr B, Scheer U (1987) Inhibition of nucleolar reformation after microinjection of antibodies to RNA polymerase I into mitotic cells. J Cell Biol 105:1483–1491
Bernhard W (1971) Drug-induced changes in the interphase nucleus. In: Clementmi F, Ceccarelli B (eds) Advances in cytopharmacology, vol 1. Raven Press, New York, pp 49–67
Bouteille M, Hernandez-Verdun D, Dupuy-Coin AM, Bourgeois CA (1982) Nucleoli and nucleolar-related structures in normal, infected and drug-treated cells. In: Jordan EG, Cullis CA (eds) The nucleolus. Cambridge University Press, Cambridge, pp 179–211
Busch H, Smetana K (1970) The nucleolus. Academic Press, New York, pp 1–626
Daskal Y (1979) Drug effects on nucleolar and extranucleolar chromatin. In: Busch H, Crooke ST, Daskal Y (eds) Effects of drugs on the cell nucleus. Academic Press, New York, pp 107–124
De la Torre C, Gimenez-Martin G (1982) The nucleolar cycle. In: Jordan EG, Cullis CA (eds) The nucleolus. Cambridge University Press, Cambridge, pp 153–177
Eckert WA, Franke WW, Scheer U (1975) Nucleocytoplasmic translocation of RNA in Tetrahymena pyriformis and its inhibition by actinomycin D and cycloheximide. Exp Cell Res 94:31–46
Franke WW, Grund C, Osborn M, Weber K (1978) The intermediate-sized filaments in rat kangaroo PtK2 cells. I. Morphology in situ. Cytobiologie 17:365–391
Franke WW, Schmid E, Vandekerckhove J, Weber K (1980) A permanently proliferating rat vascular smooth muscle cell with maintained expression of smooth muscle cell characteristics, including actin of the vascular smooth muscle type. J Cell Biol 87:594–600
Goessens G (1984) Nucleolar structure. Int Rev Cytol 87:107–158
Goessens G, Lepoint A (1974) The fine structure of the nucleolus during interphase and mitosis in Ehrlich tumor cells cultivated in vitro. Exp Cell Res 87:63–72
Hadjiolov AA (1985) The nucleolus and ribosome biogenesis. Cell Biol Monogr 12:1–268
Hadjiolova K, Rose KM, Scheer U (1986) Immunolocalization of nucleolar proteins after D-galactosamine-induced inhibition of transcription in rat hepatocytes. Exp Cell Res 165:481–493
Hügle B, Hazan R, Scheer U, Franke WW (1985) Localization of ribosomal protein S1 in the granular component of the interphase nucleolus and its distribution during mitosis. J Cell Biol 100:873–886
Kostura M, Craig N (1986) Treatment of Chinese hamster ovary cells with the transcriptional inhibitor actinomycin D inhibits binding of messenger RNA to ribosomes. Biochemistry 25:6384–6391
Krohne G, Benavente R (1986) The nuclear lamins. A multigene family of proteins in evolution and differentiation. Exp Cell Res 162:1–10
Mercer WE, Avignolo C, Galanti N, Rose KM, Hyland JK, Jacob ST, Baserga R (1984) Cellular DNA replication is independent of the synthesis or accumulation of ribosomal RNA. Exp Cell Res 150:118–130
Monneron A (1971) Action of some drugs on liver nuclei and polysemes. In: Clementini F, Ceccarelli B (eds) Advances in cytopharmacology, vol 1. Raven Press, New York, pp 131–144
Narayan KS, Steele WJ, Busch H (1966) Evidence that the granular and fibrillar components of nucleolus contain 28 and 6S RNA, respectively. Exp Cell Res 43:483–492
Ochs RL, Lischwe M, Spohn WH, Busch H (1985) Fibrillarin: a new protein of the nucleolus identified by autoimmune sera. Biol Cell 54:123–134
Perry RP (1966) On ribosome biogenesis. Natl Cancer Inst Monogr 23:527–545
Reimer G, Rose KM, Scheer U, Tan EM (1987a) Antibody to RNA polymerase I in scleroderma sera. J Clin Invest 79:65–72
Reimer G, Raska I, Tan EM, Scheer U (1987b) Human autoantibodies: probes for nucleolus structure and function. Virchows Arch B 54:131–143
Reimer G, Pollard KM, Pennig CA, Ochs RL, Lischwe MA, Busch H, Tan EM (1987c) Monoclonal antibody from NZB/NZW F1 mouse and some human scleroderma sera target a Mr 34,000 nucleolar protein of the U3-ribonucleoprotein particle. Arthritis Rheum 30:793–800
Rose KM, Stetler DA, Jacob ST (1981) Protein activity of RNA polymerase I purified from rat hepatoma: probable function of Mr 42,000 and 24,000 polypeptides. Proc Natl Acad Sci USA 78:2833–2837
Scheer U, Raska I (1987) Immunocytochemical localization of RNA polymerase I in the fibrillar centers of nucleoli. Chromosomes Today 9:284–294
Scheer U, Rose KM (1984) Localization of RNA polymerase I in interphase cells and mitotic chromosomes by light and electron microscopic immunocytochemistry. Proc Natl Acad Sci USA 81:1431–1435
Schlegel RA, Miller LS, Rose KM (1985) Reduction of RNA synthesis following red cell-mediated microinjection of antibodies to RNA polymerase I. Cell Biol Int Rep 9:341–350
Schmidt-Zachmann MS, Hügle B, Scheer U, Franke WW (1984) Identification and localization of a novel nucleolar protein of high molecular weight by a monoclonal antibody. Exp Cell Res 153:327–346
Simard R, Langelier Y, Mandeville R, Maestracci N, Royal A (1974) Inhibitors as tools in elucidating the structure and function of the nucleus. In: Busch H (ed) The cell nucleus, vol 1. Academic Press, New York, pp 447–487
Smetana K, Busch H (1974) The nucleolus and nucleolar DNA. In: Busch H (ed) The cell nucleus, vol. 1. Academic Press, New York, pp 75–147
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Benavente, R., Reimer, G., Rose, K.M. et al. Nucleolar changes after microinjection of antibodies to RNA polymerase I into the nucleus of mammalian cells. Chromosoma 97, 115–123 (1988). https://doi.org/10.1007/BF00327368
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00327368