Showing papers on "Nucleolus published in 1989"

U3, U8 and U13 comprise a new class of mammalian snRNPs localized in the cell nucleolus

Abstract: Using anti-(U3)RNP autoantibodies, we have isolated and characterized two additional small nucleolar RNAs from HeLa cells, which are less abundant than U3 RNA. Both RNAs possess a trimethylguanosine cap as judged by precipitation with anti-TMG antibody, but are not precipitated by either anti-Sm or anti-La antibodies. In addition, both RNAs are not precipitable by anti-Th serum, which recognizes another nucleolar RNP autoantigen. Sequence analysis revealed that one of these RNAs, 136 nucleotides long, is the human U8 homolog; while the other, 105 nucleotides long, represents a novel species which we designate U13. Both RNAs share with U3 two conserved sequences (boxes C and D). The role of one or both of these boxes in binding the common 34 kd antigenic protein, otherwise known as fibrillarin, is discussed. Fractionation of HeLa cells revealed that U8 and U13, like U3, reside in the nucleolus. In glycerol gradients both RNAs cosediment with larger structures possibly representing ribosomal precursors. We propose that U3, U8 and U13 comprise a new subset of mammalian snRNPs whose roles in ribosome biogenesis are discussed.

A yeast nucleolar protein related to mammalian fibrillarin is associated with small nucleolar RNA and is essential for viability.

Abstract: In order to study the structural and functional organization of the eukaryotic nucleolus, we have started to isolate and characterize nucleolar components of the yeast Saccharomyces cerevisiae. We have identified a major 38 kd nucleolar protein (NOP1), which is located within nucleolar structures resembling the dense fibrillar region of mammalian nucleoli. This 38 kd protein is conserved in evolution since affinity-purified antibodies against the yeast protein stain the nucleolus of mammalian cells in indirect immunofluorescence microscopy and the yeast protein is decorated by antibodies directed against human fibrillarin. Affinity-purified antibodies against the yeast NOP1 efficiently precipitate at least seven small nuclear RNAs involved in rRNA maturation. We have cloned the gene encoding the yeast NOP1 protein. Haploid cells carrying a disrupted copy of the gene are not viable, showing that NOP1 is essential for cell growth. The gene codes for a 34.5 kd protein which contains glycine/arginine rich sequence repeats at the amino terminus similar to those found in other nucleolar proteins. This suggests that NOP1 is in association with small nucleolar RNAs, required for rRNA processing and likely to be the homologue of the mammalian fibrillarin.

Nucleologenesis and the onset of transcription in the eight-cell bovine embryo: fine-structural autoradiographic study.

Abstract: Eight-cell cow embryos were isolated and cultured in vitro in a medium enriched with 200 μCi of [5-3H]uridine for 20 min. Epon ultrathin sections of the embryos were investigated for the nucleolar morphology and for the appearance and localization of the sites of [5-3H]uridine incorporation by means of electron microscopic autoradiography. In addition to this, a general pattern of replicated embryonal DNA distribution was revealed by [methyl-3H]thymidine incorporation and light microscopic autoradiography. The essential phases of the transformation of the small nucleous precursor body (NPB) into a vast, functionally fully active nucleolus, characterized by typical nucleolar substructural components, are taking place within the eight-cell stage. This process differed in its morphology from the nucleologenetic process in early embryogenesis of other mammals, especially of that in the mouse. The first sign of NPB, transformation was the appearance of a large central vacuole followed later on by perinucleolar chromatin penetration into NPB, documented by both morphology and [3H]thymidine autoradiography. In some cases, concentration of dense fibrillar material forming clumps or stalks was seen in the central vacuole. The following rapid nucleolar development was characterized by the formation of secondary vacuoles concomitant with the onset of [5-3H]uridine incorporation into the dense fibrillar component and with the appearance of the first granules in the otherwise fibrillar structure of the nucleolus. During the late eight-cell stage, the still-rounded nucleolus developed features of a reticulated nucleolus known from somatic cells intensively synthesizing rRNA: a dense fibrillar component with associated labeling encircling fibrillar centers and a well-developed granular component. The labeled dense fibrillar component was observed mostly in the central area of the nucleolus; early embryonic NPB dense fibrous material not involved in transcription was disappearing rapidly. At the transition to the 16-cell stage the nucleoli lost their rounded shape because of the accumulation of a large amount of granular component, and they occupied a considerable part of the nucleus. In conclusion, the appearance of the nucleolar vacuole in eight-cell cow embryo is the starting point for following morphogenetic events linked with the onset of transcription.

Human antral follicles: Oocyte nucleus and the karyosphere formation (electron microscopic and autoradiographic data)

Abstract: The functional organization of the nucleus in the oocytes from human antral follicles was examined by morphological and autoradiographic analysis methods at the light and electron microscopic level. According to the position of the nucleus, the level of its transcriptional activity, and the pattern of distribution of structures in it, oocytes fall into two groups. In the first one, the oocytes with the nucleus in the central position are characterized by the distribution of numerous structures all over the nucleus or by a different extent of aggregation of chromatin around the nucleolus. The nuclei of these oocytes are characterized by [3H]uridine incorporation, the label being localized over purely fibrillar, agranular nucleoli and over dispersed fibrillar chromatin adjacent to either the regions of densely packed chromatin or fibrillar-granular material of the nucleolus-like bodies. The latter, the same as condensed chromatin, do not incorporate [3H]uridine. In the second group, the nuclei are displaced towards the oocyte's periphery, and chromosomes surround the nucleolus as a continuous mass closely adjacent to its surface, thus forming a karyosphere. The karyosphere formation takes place on the background of cessation of nuclear transcriptional activity. A fully formed karyosphere represents a complex of closely associated inactivated structures: Nucleolus, chromosomes, and nucleolus-like bodies. The karyosphere nucleolus bears no granules and consists of densely packed finely fibrillar material (fibrils 3 nm thick). Two zones (central and peripheral) can be distinguished in a nucleolus. Nucleolus-like bodies, consisting of granules 20 nm in diameter embedded in finely fibrillar material, are often associated with chromosomes. In this study, data obtained by observations on the loss of association between the oocyte (with karyosphere) and corona radiata cells are evaluated. The relation of the karyosphere formation to the atresia process and the duration of karyosphere existence in human antral follicles are also discussed.

Protein kinase NII and the regulation of rDNA transcription in mammalian cells.

Abstract: Transcription of ribosomal RNA genes is generally accepted to correlate with cell growth. Using primary cultures of adult bovine aortic endothelial (ABAE) cells, we have shown that transcription of rDNA in confluent cells falls to 5% of the transcription level in growing cells. Protein kinase NII appears to be a limiting factor to promote rDNA transcription in isolated nuclei of confluent cells. Protein kinase NII was detected by immunocytochemistry in the cytoplasm, nuclei and nucleoli of growing cells while it was no longer present in nucleoli of confluent cells. The kinase activity, in isolated nuclei, was estimated by endogenous phosphorylation of a specific substrate, nucleolin. A 10% residual activity was present in confluent cell nuclei compared to growing cell nuclei. Concomitantly, the transcription 'in vitro' of rDNA in the corresponding nuclei was also highly reduced (by 85%). Addition of exogenous protein kinase NII to confluent cell nuclei induced a strong increase in the phosphorylation of specific proteins including nucleolin. In parallel, the transcription of rDNA was increased by a factor of 5, to nearly the level observed in nuclei prepared from growing cells. These data suggest that, in confluent cells, factors necessary for rDNA transcription machinery are present but inactive in the nucleolus and that the phosphorylation of one or several of these factors (nucleolin, topoisomerase I,...) by protein kinase NII is a key event in the regulation of rDNA transcription.

Simultaneous immunoelectron microscopic visualization of protein B23 and C23 distribution in the HeLa cell nucleolus.

Abstract: The intranucleolar distribution of phosphoproteins B23 and C23 was visualized simultaneously by post-embedding immunoelectron microscopy in HeLa cell nucleoli, using specific antibodies The data show that proteins B23 and C23 co-localize to the same nucleolar compartments, ie, the dense fibrillar component and the granular component Neither of the two antibodies is significantly associated with the fibrillar centers in these cells, although the fibrillar centers appear positive after silver staining These findings suggest that other unidentified components must be responsible for the silver staining observed in the fibrillar centers of interphase nucleoli The results are discussed in the light of previously reported data obtained by preembedding immunolabeling techniques and by silver staining, which both suggested a localization of protein C23 inside the fibrillar centers

Herpes simplex virus alpha protein ICP27 possesses separable positive and negative regulatory activities.

Abstract: The HSV-1 alpha (immediate-early) protein ICP27 expressed in transfected cells can activate the expression of certain HSV-1 promoters as well as inhibit the transactivated expression of others. We constructed a set of plasmids encoding mutant ICP27 molecules truncated at their carboxyl termini and used transfection assays to determine the functional properties of the mutant proteins. A polypeptide containing the amino-terminal 263 amino acid residues of ICP27 retained partial ability to activate gene expression but was unable to inhibit transactivation. Mutant proteins possessing 406 or 504 amino acids of ICP27 were unable to activate gene expression but retained full ability to inhibit transactivation. These results define two separable regulatory activities of ICP27, one positive and one negative, which can modulate gene expression in transfected cells. Immunoblot and immunofluorescence experiments were used to study the immunological reactivities and intracellular localizations of the mutant proteins. All proteins possessing the amino-terminal 263 amino acids of ICP27 reacted with an ICP27-specific monoclonal antibody and were localized to the cell nucleus. The mutant proteins, however, exhibited a number of phenotypes with regard to intranuclear localization. A mutant possessing 504 residues of ICP27 was similar to the wild-type protein in apparently localizing to all regions of the nucleus. A mutant containing 406 residues of ICP27, on the other hand, was mostly excluded from the nucleolar regions, while a 263-residue mutant was localized predominantly in the nucleoli. Thus, some aspect of ICP27 structure or function can dramatically affect its intranuclear distribution.

Nucleolar targeting signal of human T-cell leukemia virus type I rex-encoded protein is essential for cytoplasmic accumulation of unspliced viral mRNA.

Abstract: The posttranscriptional regulator (rex) of human T-cell leukemia virus type I is known to be located predominantly in the cell nucleolus and to induce the accumulation of gag and env viral mRNAs. The N-terminal 19 amino acids of rex-encoded protein (Rex) has been shown to be sufficient to direct hybrid proteins to the cell nucleolus. We have studied the function of the nucleolar targeting signal (NOS) of rex by using full-length proviral DNA and mutant rex expression plasmids. Partial deletions of the NOS sequence abolished the accumulation of unspliced cytoplasmic mRNA, although the gene products of rex mutants were found in the nucleoplasm. These results indicate that NOS sequence, or nucleolar localization of Rex, is essential for Rex function.

Nucleolin from Xenopus laevis: cDNA cloning and expression during development.

Abstract: Nucleolin is a key nucleolar protein in higher eukaryotic cells and is involved directly in ribosome biogenesis. Using an antiserum raised against hamster nucleolin, the homologous protein was detected in nucleoli of Xenopus laevis hepatocytes as well as in the amplified nucleoli of oocytes. A cDNA encoding Xenopus nucleolin has been isolated and sequenced. The deduced protein sequence reveals similar domains in Xenopus and in mammals, but they have undergone separate evolutions. In particular, each of the four RNA-binding domains has evolved differently--the carboxy-proximal domain is twice as conserved (87%) as the amino-proximal domain (42%). These data shed some light on the possible roles of each domain. The expression of nucleolin has been followed throughout oogenesis and embryogenesis. The appearance of nucleolin during early development precedes the transcription of rDNA and the synthesis of ribosomal proteins. The maximal accumulation of nucleolin at gastrulation coincides with nucleolar reformation. Furthermore, when ribosomal synthesis is activated during oogenesis and embryogenesis, peptides immunorelated to nucleolin appear and accumulate. The results suggest that nucleolin plays a role not only in ribosome assembly but also in nucleologenesis.

Genome reactivation in developing early pig embryos: an ultrastructural and autoradiographic analysis.

Abstract: The onset of RNA synthesis in developing early pig embryos from 1-cell to 8-cell and morula stages was studied using high-resolution autoradiography of (5-3H)uridine incorporation. No transcriptional activity was detected in nuclei of 1- and 2- cell stage embryos with this technique. In these embryos nucleolus-like bodies (NLB) consist of sharply delineated, round, electron dense fibrillar masses. In the 4-cell stage embryos, the first uridine-3H incorporation in the nucleoplasm was detected and localized mainly near the regions of condensed chromatin. The first signs or reticulation and chromatin association were observed at the periphery of NLBs. In the next cell cycle (5- to 8-cell embryos) uridine-3H labelling was detected in the nucleoplasm and nucleoli. In these embryos, nucleoli consist of a central dense fibrillar mass without any transcriptional activity and fibrillo-granular cortex over which label was localized. The degree of functional restructure of nucleoli was variable within one blastomere or among different blastomeres, some nucleoli being more reticulated and showing more transcriptional activity than others. Fully developed nucleoli were present in early morulae. Electron dense unidentified structures described here as small dense roundshaped bodies (RDB) often surrounded by blocks of large chromatin granules were observed in intact 2-cell and alpha-amanitin treated 4-cell stage embryos. These structures did not show any transcriptional activity

The sphere organelle contains small nuclear ribonucleoproteins.

Abstract: We show by immunofluorescence microscopy of amphibian oocyte nuclei that small nuclear ribonucleoproteins (snRNPs) occur in lampbrush chromosome loops, in a few dozen extrachromosomal organelles previously described as "spheres," and in thousands of smaller granules. Spheres are variable in size (up to approximately 20 microns in diameter in the newt Notophthalmus and approximately 10 microns in the frog Xenopus) and are easily distinguishable from nucleoli by morphology and composition. Spheres occur both free in the nucleoplasm and attached to specific chromosome loci, the sphere organizers. Oocyte nuclei of a cricket and a spider contain essentially similar organelles, suggesting that spheres may be common throughout the animal kingdom. We suggest that spheres play a role in the assembly of snRNP complexes for the nucleus comparable to the way that nucleoli assemble ribosomal RNP complexes for the cytoplasm.

Higher order structure is present in the yeast nucleus: autoantibody probes demonstrate that the nucleolus lies opposite the spindle pole body

Abstract: A panel of sera from 892 autoimmune patients was screened by indirect immunofluorescence on mammalian cells. Seventy-three sera were identified that recognize the nucleolus. Three of these sera appear to stain the nucleolus in yeast, suggesting that they recognize highly conserved antigens. These three sera also immunoprecipitate mammalian U3 snRNA-containing particles, which reside in the nucleolus and have been implicated in rRNA processing. Double immunofluorescence experiments with anti-nucleolus and anti-tubulin antibodies revealed a novel form of non-random nuclear organization in yeast. The spindle pole body and the nucleolus-both of which are associated with the nuclear envelope-preferentially localize at opposite ends of the nucleus. Organization of these and other components into specific regions of the nucleus may be important for optimizing their proper function.

Essential roles of the RNA polymerase I largest subunit and DNA topoisomerases in the formation of fission yeast nucleolus.

Abstract: A temperature-sensitive lethal mutant nuc1-632 of Schizosaccharomyces pombe shows marked reduction in macromolecular synthesis and a defective nuclear phenotype with an aberrant nucleolus, indicating a structural role of the nuc1+ gene product in nucleolar organization. We cloned the nuc1+ gene by transformation and found that it appears to encode the largest subunit of RNA polymerase I. We raised antisera against nuc1+ fusion polypeptides and detected a polypeptide (approximately 190 kD and 2 x 10(4) copies/cell) in the S. pombe nuclear fraction. By immunofluorescence microscopy, anti-nuc1+ antibody revealed intense staining at a particular nuclear domain previously defined as the nucleolus. The nucleolar immunofluorescence by anti-nuc1+ was faded in nuc1-632 at restrictive temperature and dramatically diminished in the absence of DNA topoisomerases I and II. Thus active RNA polymerase I appears to be required for the formation of the nucleolus as its major component, and DNA topoisomerases appear to be required for the folding of rDNA and RNA polymerase I molecules into the functional organization of nucleolar genes.

Detection of dengue 4 virus core protein in the nucleus. I. A monoclonal antibody to dengue 4 virus reacts with the antigen in the nucleus and cytoplasm.

Abstract: A mouse monoclonal antibody (MAb) to dengue 4 (DEN-4) virus reacted with the antigen in the nucleus as well as in the cytoplasm of DEN-4-infected mammalian and mosquito cells, as demonstrated by the peroxidase-antiperoxidase staining method. The intranuclear antigen appeared to accumulate at the nucleoli, forming spots, whereas the cytoplasmic antigen appeared to be localized mainly in large perinuclear foci in the infected cells. The MAb-reactive antigen was produced in the presence of actinomycin D, which caused the accumulation in the nucleus to be altered to a dispersed pattern. Radioimmunoprecipitation analysis of [35S]methionine-labelled purified virions and Western blot analysis of the antigens prepared from the infected mammalian and mosquito cells showed that the MAb was directed against the DEN-4 virus core protein (Mr 15.5K). These results indicated that the DEN-4 virus core protein was partially transported, soon after its synthesis in the cytoplasm, into the nucleus and accumulated at the nucleoli.

High-resolution autoradiographic studies of comparative nucleologenesis and genome reactivation during early embryogenesis in pig, man and cattle.

Abstract: Using high-resolution autoradiography, simultaneous studies of ultrastructure and nucleic acid dynamics were performed during nucleologenesis in early porcine, human and bovine embryos. In contrast to the early genome activation known to occur during the second cell cycle in the mouse, the onset of rRNA synthesis detected by (5-3H) uridine incorporation in the nucleolar compartment is delayed by cleavage of one cell cycle (to the third cell cycle) in the early pig embryo and by two cell cycles (to the fourth cell cycle) in human and cattle embryos. Extranucleolar RNA synthesis, as detected by nucleoplasm labelling, generally started shortly before rRNA synthesis. The timing of nucleolar labelling was well correlated with the penetration of embryonic DNA into the nucleolus precursor body and with nucleolus structure differentiation. Ultrastructural and/or autoradiographic techniques are suggested for the study of the onset of embryonic transcription, e.g. in embryos "reconstructed" by micromanipulation.

Maturation at high frequency of germinal-vesicle-stage mouse oocytes after cryopreservation: alterations in cytoplasmic, nuclear, nucleolar and chromosomal structure and organization associated with vitrification

Abstract: After thawing and culture in vitro, greater than 90% of germinal-vesicle (GV)-stage mouse oocytes cryopreserved by vitrification, were capable of resuming meiosis and undergoing normal chromosomal and cytoplasmic maturation to metaphase II. This high frequency of development occurred against a background of profound alterations in the structure and organization of the cytoplasm, nucleus, nucleolus and chromatin during the dehydration stage of vitrification. Most, but not all, cytoplasmic and nuclear perturbations returned to a normal state during post-thaw culture. However, the results clearly demonstrate that vitrification is associated with chromosomal and cellular disorders that could adversely affect development after fertilization. Irreversible changes of potential developmental significance observed after vitrification at the GV stage include (i) premature chromosomal condensation, (ii) mixing of nucleoplasmic and cytoplasmic components prior to GV breakdown, and (iii) externalization of chromatin fragments into the cytoplasm after reformation of the oocyte nucleus, which shows the potential for the generation of fertilizable oocytes containing deleted segments of DNA.

Interactions of minute virus of mice and adenovirus with host nucleoli.

Abstract: Biochemical evidence is presented that both minute virus of mice (MVM) and adenovirus interact with the nucleolus during lytic growth and that MVM can also target specific changes involving nucleolar components in adenovirus-infected cells. These virus-nucleolus interactions were studied by analysis of intranuclear compartmentalization of both viral DNAs and host nucleolar proteins: (i) MVM in mouse cells (its normal host) replicates its DNA in the host nucleoli; (ii) specific nucleolar proteins as well as small nuclear ribonucleoprotein antigens are recompartmentalized to multiple intranuclear foci in adenovirus-infected HeLa cells; and (iii) when adenovirus helps MVM DNA replication in a nonpermissive human cell (HeLa), the MVM DNA is also recompartmentalized for synthesis. The data suggest mechanisms for disruption of nucleolar function common to oncogenic or oncolytic virus lytic growth and cell transformation. Images

Immunolocalization of DNA at nucleolar structural components in onion cells.

Abstract: Intranucleolar DNA, including ribosomal DNA (rDNA), was localized in situ in proliferating onion cells under the electron microscope using an anti-DNA monoclonal antibody and a postembedding indirect immunogold procedure. In the interphase nucleolus of this species, characterized by a very high amount of rRNA genes, we found DNA concentrated mostly in fibrillar centres (FCs) and in the region of the dense fibrillar component (DFC) immediately surrounding them. Clusters of gold particles were frequently seen covering both of these structural components of the nucleolus at the same time. Moreover, the same technique, applied to transcriptionally arrested quiescent onion cells, showed the nucleolar DFC devoid of DNA. Also, in mitotic cells at telophase, the prenucleolar material, which has the same morphological and cytochemical features as the DFC, does not contain DNA. These data suggest the existence of at least two subcomponents of the DFC in the onion cell nucleolus, one associated with pre-rRNA synthesis, and the other, with further processing of transcripts, already released from the rDNA template. We conclude that the first subcomponent forms part of the “transition between FC and DFC”, which is the in situ structural counterpart of pre-rRNA synthesis. This transition is morphologicaly sizeable in onion cells, because of their high number of rRNA genes and the large size of the DFC mass; however, it would be barely detectable in situ in other cell systems, where the whole DFC comprises only a thin layer and the amount of rDNA is considerably reduced.

Variation in structure and expression of ribosomal DNA loci in wheat

Abstract: Ribosomal RNA genes are organised in tandem arrays at complex loci called nucleolus organisers. The structure of a locus and of a wheat rRNA gene is described in detail. Active or potentially activ...

Isolation and characterization of yeast ribosomal RNA precursors and preribosomes.

Abstract: Publisher Summary This chapter describes the identification and characterization of the precursor RNAs and ribonucleoproteins (RNPs) from the 35 S pre-rRNA operon. In addition to 35 S prerRNA, 20 and 27 S pre-rRNAs are the predominant intermediates. The 32 S pre-rRNA is found in variable yields depending on the genetic background of the particular strain and the conditions of growth and harvesting. The major part of the ribosome biogenesis pathway is located in the nucleolus. Shortly after, or possibly already during transcription, 35 S pre-rRNA is subjected to a large number of modifications, including methylation and pseudouridylation. High-molecular-weight pre-rRNAs (20-35 S) are fractionated on agarose gels after glyoxylation or on standard formamide–formaldehyde gels and then are blotted onto a solid support. Filters are hybridized with probes containing ETS, ITS1, or ITS2 sequences, but lacking sequences corresponding to the mature and S rRNAs. Spheroplasts have to be poured into a 10-fold excess of ice-cold 1 M sorbitol containing unlabeled uracil or methionine.

Conservation of nucleolar structure in polytene tissues of Ceratitis capitata (Diptera: Tephritidae)

Abstract: Nucleolar structure was studied in mitotic and three polytene tissues of the Mediterranean fruit fly, Ceratitis capitata using in situ hybridization with a tritium-labelled rDNA probe and silver staining. In mitotic metaphase chromosomes nucleolar organiser regions were localised in the short arms of both sex chromosomes. In polytene nuclei of trichogen cells, salivary glands and fat body rDNA was detected within nucleoli. Nucleoli in these tissues have a similar structure with rDNA labelling concentrated in a central core. Silver staining resulted in very heavy staining of polytene nucleoli and interphase nucleoli in diploid cells. Silver staining of nucleolar organisers in metaphase chromosomes is weak or absent although the X chromosome has numerous dark silver bands in other locations. The results suggest that nucleolar structure is conserved in polytene tissues contrasting with the variability of autosomal banding patterns and sex chromosome structure. They also indicate that silver staining is not necessarily specific for nucleolar regions.

Effects of Hyperthermia on Chromatin Condensation and Nucleoli Disintegration as Visualized by Induction of Premature Chromosome Condensation in Interphase Mammalian Cells

Abstract: The effects of hyperthermia on chromatin condensation and nucleoli disintegration, as visualized by induction of premature chromosome condensation in interphase mammalian cells, was studied in exponentially growing and plateau phase Chinese hamster ovary cells. Exposure to heat reduced the ability of interphase chromatin to condense and the ability of the nucleolar organizing region to disintegrate under the influence of factors provided by mitotic cells when fused to interphase cells. Based on these effects treated cells were classified in three categories. Category 1 contained cells able to condense their chromatin and disintegrate the nucleolar organizing region. Category 2 contained cells able to only partly condense their chromatin and unable to disintegrate the nucleolar organizing region. Category 3 contained cells unable to condense their chromatin and unable to disintegrate the nucleolar organizing region. The fraction of cells with nondisintegrated nucleoli increased with increasing exposure time at 45.5°C and reached a plateau at almost 100% after about 20 min. Exponentially growing and plateau phase cells showed similar response. Recovery from the effects of heat on chromatin condensation and disintegration of the nucleolar organizing region depended upon the duration of the heat treatment. For exposures up to 15 min at 45.5°C, a gradual reduction in the fraction of cells with nondisintegrated nucleoli was observed when cells were allowed for repair at 37°C. However, only a very limited amount of repair was observed after a 30-min exposure to 45.5°C. The repair times observed at the chromosome level were similar to those reported for the removal of excess protein accumulating in chromatin or the nuclear matrix, suggesting a causal relationship between the two phenomena. It is proposed that nuclear protein accumulation on chromatin or in the nuclear matrix reduces the accessibility of chromatin to enzymes responsible for the phosphorylation reactions necessary for chromatin condensation and disintegration of the nucleolus.