This laboratory has previously isolated a fraction from rat liver nuclei consisting of nuclear pore complexes associated with the proteinaceous lamina which underlies the inner nuclear membrane. Using protein eluted from sodium dodecyl sulfate (SDS) gels, we have prepared antibodies in chickens to each of the three predominant pore complex-lamina bands. Ouchterlony double diffusion analysis shows that each of these individual bands cross-reacts strongly with all three antisera. In immunofluorescence localization performed on tissue culture cells with these antibodies, we obtain a pattern of intense staining at the periphery of the interphase nucleus, with little or no cytoplasmic reaction. Electron microscope immunoperoxidase staining of rat liver nuclei with these antibodies labels exclusively the nuclear periphery. Furthermore, reaction occurs in areas which contain the lamina, but not at the pore complexes. While our isolation procedure extracts the internal contents of nuclei completely, semiquantitative Ouchterlony analysis shows that it releases negligible amounts of these lamina antigens. Considered together, our results indicate that these three bands represent major components of a peripheral nuclear lamina, and are not structural elements of an internal "nuclear protein matrix." Fluorescence microscopy shows that the perinuclear interphase localization of these lamina proteins undergoes dramatic changes during mitosis. Concomitant with nuclear envelope disassembly in prophase, these antigens assume a diffuse localization throughout the cell. This distribution persists until telophase, when the antigens become progressively and completely localized at the surface of the daughter chromosome masses. We propose that the lamina is a biological polymer which can undergo reversible disassembly during mitosis.

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Immunocytochemical localization of the major polypeptides of the nuclear pore complex-lamina fraction. Interphase and mitotic distribution.

Identification and characterization of the packaging proteins of core 40S hnRNP particles

NUCLEAR RIBONUCLEOPROTEIN PARTICLES . Evidence for, and Isolation of, hnRNP Particles . General Structural Features of hnRNP Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. Proteins of hnRNP Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ...... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Arrangement of Proteins in hnRNP Monomer Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. Arrangement of hnRNA in hnRNP Particles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. Assembly and Disassembly of hnRNP Particles .. Functions of hnRNP Particles and hnRNP Proteins . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Structure and Function of Nuclear and Cytoplasmic Ribonucleoprotein Particles

Proteins associated with heterogeneous nuclear RNA in eukaryotic cells.

Publisher Summary This chapter discusses the recent studies in order to draw a contemporary picture of the structure of transcribing chromatin. The basic structure of “active” chromatin, unique, repeating, altered nucleosomal structure, is discussed in this chapter. Both micrococcal nuclease and DNase II digestion studies strongly suggest that “active” chromatin is organized in repeating subunits that share at least some properties, with bulk nucleosomes. Isolation of pure chromatin fragments containing “active” ribosomal and nonribosomal genes and careful studies of their composition and organization are required before understanding the basis for the variability of “active” chromatin structure. From the conflicting results available at the present time, it is obviously impossible to conclude whether or not histone H1 (and its variants) is present in “active” chromatin. Again, there is a need for careful well-controlled studies and again, the use of antibodies directed against the various H1 variants should be very useful in elucidating whether they are associated with the isolated “active” subunits. as they are with the isolated nucleosomes of bulk chromatin.

Structure of Transcribing Chromatin

DNA-like RNA (dRNA) is found in the nucleus as a complex with globular protein particles called informofers. Intact informofers may be detached from the dRNA and can form complexes in vitro and in vivo with different kinds of dRNA, including virus-specific RNA synthesized in adenovirus-infected cells.

Structure and function of informofers.

Transcription of the human adenovirus 5 genome in transformed rat embryo cells (DFK3) was investigated using two different approaches. Preferential digestion of transcribed viral sequences by DNase I was analysed using kinetics of renaturation of 32P-labeled Ad5 HpaI restriction fragments in the presence of material which was stable after nuclease treatment. The second approach was the hybridization of 32P-labeled nuclear RNA from transformed cells with Ad5 restriction fragments which were attached to a nitrocellulose filter. These two methods gave similar results. It was found that not all integrated regions of the Ad5 genome are active in transformed cells. 2,5 copies of the HpaI-E fragment of Ad5 DNA were found in transformed DFK3 cell line. Nuclear RNA from these cells hybridized to HpaI-E fragment of Ad5 DNA, but only about half of sequences of the integrated HpaI-E fragment was sensitive to DNase I digestion.

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E. S. Zalmanzon

Papers

Structure and function of informofers.

Studies of the transcription of viral genome in adenovirus 5 transformed cells