A three-dimensional view of precursor messenger RNA metabolism within the mammalian nucleus.

A quantitative three-dimensional analysis of nuclear components involved in precursor messenger RNA metabolism was performed with a combination of fluorescence hybridization, immunofluorescence, and digital imaging microscopy. Polyadenylate [poly(A)] RNA-rich transcript domains were discrete, internal nuclear regions that formed a ventrally positioned horizontal array in monolayer cells. A dimmer, sometimes strand-like, poly(A) RNA signal was dispersed throughout the nucleoplasm. Spliceosome assembly factor SC-35 localized within the center of individual domains. These data support a nuclear model in which there is a specific topological arrangement of noncontiguous centers involved in precursor messenger RNA metabolism, from which RNA transport toward the nuclear envelope radiates.

[1]  I. Raška,et al.  Immunological and ultrastructural studies of the nuclear coiled body with autoimmune antibodies. , 1991, Experimental cell research.

[2]  E. Tan,et al.  Differentiation of antinuclear antibodies giving speckled staining patterns in immunofluorescence , 1972 .

[3]  J. Lawrence,et al.  Higher level organization of individual gene transcription and RNA splicing. , 1993, Science.

[4]  R. Pepperkok,et al.  Transcription-dependent colocalization of the U1, U2, U4/U6, and U5 snRNPs in coiled bodies , 1992, The Journal of cell biology.

[5]  H. Weintraub,et al.  Localization of DNAase I-sensitive sequences to specific regions of interphase nuclei , 1985, Cell.

[6]  D. Spector,et al.  Immunoelectron microscopic localization of snRNPs , 1983, Biology of the cell.

[7]  D. Spector.  Higher order nuclear organization: three-dimensional distribution of small nuclear ribonucleoprotein particles. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[8]  O. Miller,et al.  Correlation of hnRNP structure and nascent transcript cleavage , 1981, Cell.

[9]  G. Leser,et al.  Ultrastructural distribution of nuclear ribonucleoproteins as visualized by immunocytochemistry on thin sections , 1984, The Journal of cell biology.

[10]  J. Lawrence,et al.  Preservation of specific RNA distribution within the chromatin-depleted nuclear substructure demonstrated by in situ hybridization coupled with biochemical fractionation , 1991, The Journal of cell biology.

[11]  J. Lawrence,et al.  Sensitive, high-resolution chromatin and chromosome mapping in situ: Presence and orientation of two closely integrated copies of EBV in a lymphoma line , 1988, Cell.

[12]  T. Puck,et al.  The spatial distribution of exposed nuclear DNA in normal, cancer, and reverse-transformed cells. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[13]  J. Gall,et al.  Spliceosomes and snurposomes , 1991, Science.

[14]  J. Lawrence,et al.  Discrete nuclear domains of poly(A) RNA and their relationship to the functional organization of the nucleus , 1991, The Journal of cell biology.

[15]  W. A. Carrington.  Image restoration in 3-D microscopy with limited data , 1990, Photonics West - Lasers and Applications in Science and Engineering.

[16]  K. Pienta,et al.  The role of the nuclear matrix in the organization and function of DNA. , 1986, Annual review of biophysics and biophysical chemistry.

[17]  T. Maniatis,et al.  Associations between distinct pre‐mRNA splicing components and the cell nucleus. , 1991, The EMBO journal.

[18]  S. Fakan,et al.  The ultrastructural visualization of nucleolar and extranucleolar RNA synthesis and distribution. , 1980, International review of cytology.

[19]  G. Leser,et al.  Immunoelectron microscope visualization of nuclear ribonucleoprotein antigens within spread transcription complexes , 1986, Journal of Cell Biology.

[20]  N. Ringertz,et al.  Intranuclear localization of snRNP antigens , 1986, The Journal of cell biology.

[21]  J B Lawrence,et al.  Fluorescent detection of nuclear RNA and DNA: implications for genome organization. , 1991, Methods in cell biology.

[22]  D. Spector,et al.  U1 and U2 small nuclear RNAs are present in nuclear speckles , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[23]  Y. Wang,et al.  Localization of pre-messenger RNA at discrete nuclear sites. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[24]  S A Berman,et al.  Localization of an acetylcholine receptor intron to the nuclear membrane. , 1990, Science.

[25]  G. Blobel,et al.  Gene gating: a hypothesis. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[26]  L. Maquat.  Nuclear mRNA export. , 1991, Current opinion in cell biology.

[27]  T. Maniatis,et al.  Factor required for mammalian spliceosome assembly is localized to discrete regions in the nucleus , 1990, Nature.

[28]  J. Nevins.  The pathway of eukaryotic mRNA formation. , 1983, Annual review of biochemistry.

[29]  D. Spector,et al.  Differences in snRNP localization between transformed and nontransformed cells. , 1992, Molecular biology of the cell.

[30]  M. Rosbash,et al.  A yeast splicing factor is localized in discrete subnuclear domains. , 1992, The EMBO journal.

[31]  R. Benavente,et al.  Functional and dynamic aspects of the mammalian nucleolus , 1990, BioEssays : news and reviews in molecular, cellular and developmental biology.

[32]  R. Pepperkok,et al.  Mammalian nuclei contain foci which are highly enriched in components of the pre‐mRNA splicing machinery. , 1991, The EMBO journal.

[33]  J. Gall,et al.  The sphere organelle contains small nuclear ribonucleoproteins. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[34]  R. Pepperkok,et al.  In vivo detection of snRNP‐rich organelles in the nuclei of mammalian cells. , 1991, The EMBO journal.

[35]  Robert H. Singer,et al.  Highly localized tracks of specific transcripts within interphase nuclei visualized by in situ hybridization , 1989, Cell.