Repair of gamma-ray-induced DNA base damage in xeroderma pigmentosum cells

The repair of DNA damage produced by 137Cs gamma irradiation was measured with a preparation from Micrococcus luteus containing DNA damage-specific endonucleases in combination with alkaline elution. The frequency of these endonuclease sensitive sites (ESS) was determined after 54 or 110 Gy of oxic irradiation in normal and xeroderma pigmentosum (XP) fibroblasts from complementation groups A, C, D, and G. Repair was rapid in all cell strains with greater than 50% repair after 1.5 h of repair incubation. At later repair times, 12-17 h, more ESS remained in XP than in normal cells. The frequency of excess ESS in XP cells was approximately 0.04 per 10(9) Da of DNA per Gy which was equivalent to 10% of the initial ESS produced. The removal of ESS was comparable in XP cells with normal radiosensitivity and XP3BR cells which have been reported to be moderately radiosensitive.     

Dense distribution of nuclear alkaline protease to nucleoli with increase in rapidly growing cells in rats

Neutral and alkaline proteases (chymotrypsin-like serine proteases) are tightly bound to chromatin in nuclei of various tissues of rats, and rapidly growing cells are abundant in the latter enzyme. Activities per mg DNA of these enzymes were measured with fractions of euchromatin, heterochromatin, nucleoli and extranucleoli from normal and regenerating livers, and Rhodamine sarcoma. With normal liver, both enzymes, respectively, were almost evenly distributed among the fractions, except that alkaline protease was significantly higher in nucleoli. The nucleolar alkaline protease increased by 30-60% with regenerating liver and by higher than 100% with the tumor.     

Ultrastructural studies of H-1 parvovirus replication : III. Intracellular localization of viral antigens with immunocytochrome c

Immunoelectron microscopy with cytochrome c conjugated anti-H-1 IgG was used to localize antigens of the parvovirus H-1 within synchronized human NB cells. Since glutaraldehyde destroyed H-1 antigenicity, a fixative containing formaldehyde was developed which preserved both cellular ultrastructure and antigenic function. The earliest H-1 specific staining occurred on the heterochromatin bordering the nuclear envelope at 8 h post infection (p.i.). At 10 h p.i., labeling was found on the chromatin associated with the nucleolar surface and tufts of heterochromatin distributed throughout the nucleoplasm. Except for this H-1 labeling, the chromatin appeared indistinguishable from that of uninfected cells. By 12 h p.i., however, coinciding with the abrupt rise in synthesis of H-1 hemagglutinin and infectious virus, H-1 labeled intranuclear chromatin had condensed and migrated toward the nuclear membrane. Also, trabeculae of intranuclear chromatin were tagged with anti-H-1 conjugate as contraction of nucleolar chromatin and disintegration of nucleolar ultrastructure began. Condensation of the nucleolar associated chromatin and nuclear heterochromatin appeared complete by 18–36 h p.i. when thick zones of this H-1 labeled material were observed at the nucleolar and nuclear periphery. Our results indicate that the binding of unassembled H-1 proteins to specific regions of chromatin is associated with their condensation and margination, resulting in early nucleolar destruction and subsequent nuclear damage during H-1 infection.     

Morphological and immunocytochemical characterization of snake-like chromatin cells

Snake-like chromatin (SLC) is a nuclear alteration occurring under various pathological conditions and in different tissues. The aim of this study was the morphological and immunocytochemical characterization of SLC-positive conjunctival epithelial cells from keratoconjunctivitis sicca (KCS) patients. Impression cytology specimens from the upper bulbar conjunctiva of 10 controls and 10 KCS patients with a high incidence of SLC cells were assessed, the morphology of SLC nuclei evaluated by light microscopy, and proliferation markers, nucleolar proteins, lamins and cytokeratin filaments detected immunocytochemically. In KCS patients, SLC cells with a normal nuclear shape, with nuclear membrane notching (2.3% of cells) and with binuclear dumb-bell structures (4.4% of cells) were observed. The most striking features of SLC cells were the absence of an A/C lamin signal, the redistribution of fibrillarin into two spots adjacent to SLC structures and cytokeratin 14 positivity in the strangulation belt of the dumb-bell structures. The deficiency of lamin A/C is the probable reason for the disintegration of chromatin from the nuclear lamina in SLC cells. The occurrence of SLC-positive cells, SLC-positive dumb-bell shaped nuclei and SLC-positive binucleated cells, together with the absence of mitotic markers, leads to the conclusion that the SLC phenomenon might be a form of nuclear segregation.     

Karyometry of breast epithelial cells acquired by random periareolar fine needle aspiration in women at high risk for breast cancer

OBJECTIVE: To establish whether karyometry was likely to detect change in the proportion of abnormal cells in random periareolar fine needle aspiration (RPFNA) specimens from high-risk women in a 6-month prevention trial with an aromatase inhibitor. STUDY DESIGN: Papanicolaou-stained ThinPrep slides of RPFNA samples from 11 of 42 women were digitally recorded at high resolution, with 200 cells measured per slide, at baseline (BL) and at the end of study (ES) after 6 months. The nuclear chromatin pattern characteristics were assessed by multivariate analytic techniques; determination of nuclear abnormalities was performed and cells that showed expression of abnormality were identified. RESULTS: The BL FNA samples contain approximately 90% cells with a chromatin pattern as expected in a normal cell population. A small subpopulation of cells had deviations from normal. At ES the proportion of these cells was reduced, to a statistically significant degree,from < 10% to 2-5%. CONCLUSION: Nuclear karyometry is a promising technique for characterizing the proportion of cells deviating from normal in cytologic specimens and should be explored further as an intermediate endpoint in prevention trials.     

Nuclear matrix, hnRNA, and snRNA in friend erythroleukemia nuclei depleted of chromatin by low ionic strength EDTA

A novel and rapid procedure is described for the preparation of chromatin-depleted nuclei (CDN) from Friend erythroleukemia cells under conditions that avoid the use of high salt concentrations. By this procedure isolated nuclei that had previously been incubated with DNase I to partially digest DNA were washed twice in 2 mM EDTA to extract the chromatin. The resulting structures contained 1% of DNA, 65% of total RNA, 60-80% of hnRNA, 74% of snRNA, 29% of protein, and 2% of histones contained in isolated nuclei. Electron microscopy revealed intact, spherical structures similar in diameter to isolated nuclei and consisting of dense networks of fibrils 50-100 A thick surrounded by distinct nuclear laminae. Although no morphological evidence was found for residual nucleoli, C23, a nucleolus-specific phospho-protein, remained centrally localized in CDN, while Sm antigen specific for snRNPs was diffusely localized but absent from central regions. Addition of 2 mM MgCl2 to CDN resulted in the reformation of morphologically distinguishable residual nucleoli. These studies suggest that nucleolar morphology is, in part, dependent upon divalent cations and components unique to the nucleolar matrix and demonstrate that little randomization of nuclear and nucleolar matrix fibrils occurs during CDN isolation in the absence of divalent cations.     

Defect in UV-induced unscheduled DNA synthesis in cultured epidermal keratinocytes from xeroderma pigmentosum

DNA repair synthesis in 8 explant-outgrowth cultures of epidermal cells isolated from variant and complementation groups A and E of xeroderma pigmentosum (XP) was examined by measuring unscheduled DNA synthesis (UDS) on autoradiographs. The extents of UDS in XP epidermal cells were compared with those in normal epidermal cells obtained from 26 subjects. In both normal and XP epidermal cells, UDS was induced dose-dependently by radiation at doses of 5-20 J/m2. XP epidermal cells showed various extents of defect in DNA repair depending on the type of XP. In XP-A, the extent of UDS in epidermal cells was very low, being seen in only 3-10% of the normal epidermal cells. But epidermal cells isolated from XP-E and XP-variants exhibited relatively high levels of residual DNA repair; i.e., 69-84% of the control in XP-E and 67-85% in XP-variant. The extents of UDS in XP epidermal cells were almost the same as those in fibroblastic cells isolated from the same specimens.     

The nucleoli matrix proteins with molecular masses 40 and 27 kDa are transported as peripheral chromosomal material

Using immunofluoresence method, sera M-311 and K-30 obtained from patients with autoimmune disease were shown to stain interphase nuclei and the periphery of chromosomes. Western blotting revealed a polypeptide with mol. mass 27 kDa in serum K-30. Both proteins were localized in the karyoplasm. One of them (27 kDa) has a diffuse form and contains small granules, while the other (40 kDa) is in the form of small clearly outlined granules. Both proteins are also revealed around the nucleolar periphery, making a continental ring, while the main part of the nucleolus remains unstained. During pro- and metaphase, these proteins were associated with the chromosomal periphery: 27 kDa protein formed separate groups, and 40 kDa protein was seen over the whole chromosomal periphery. After nuclear and chromosomal decondensation, induced by hypotonic treatment (15% of culture medium solution), both antibodies stain diffusively interphase nuclei, but in mitotic cells they stained the surface of the swollen chromosomes. After chromatin recondensation in isotonic medium these proteins were localized similarly as in normal cells. Thus, both proteins maintained their association with the periphery of chromosomes. To reveal the nuclear protein matrix, cells were treated with 2M NaCl, DNAase and RNAase A. After this procedure, the antibodies stained only the nucleolar periphery, and no fluorescence in the karyoplasm was seen. It shows that of all the components of the nuclear protein matrix (lamina, internuclear network, residual nucleoli) only 27 and 40 kDa proteins are contained in the nucleolar rim. The data allow to suggest that the nucleolar matrix proteins may be transported to new cell nuclei as part of the peripheral chromosomal material likely as other nucleolar (fibrillarin, B-23, and others) or some non-nuclear components of the nuclear protein matrix are transported.     

Quantification of expression of linked cloned genes in a simian virus 40-transformed xeroderma pigmentosum cell line.

We wished to determine whether simian virus 40 (SV40)-transformed xeroderma pigmentosum cells, despite their defective DNA repair, were suitable for DNA-mediated gene transfer experiments with linked genes. Expression of a nonselectable gene (cat, coding for chloramphenicol acetyltransferase [CAT]) linked to a selectable gene (gpt, coding for xanthine-guanine phosphoribosyltransferase [XPRT]) in the plasmid pSV2catSVgpt was quantified after transfection of SV40-transformed xeroderma pigmentosum [XP20s(SV40)] and normal human [GM0637(SV40)] fibroblast cell lines. A novel autoradiographic assay with [3H]xanthine incorporation showed 0.5 to 0.7% phenotypic expression of XPRT in both cell lines. Without selection, transient CAT activity was 20 times greater in the GM0637(SV40) than in the XP20s(SV40) cells, and transient XPRT activity was 5 times greater. Both of these transient activities were increased and equalized in both cell lines by transfection with pRSVcat or pRSVgpt. Genotypic transformation to gpt+ occurred at a frequency of 2 X 10(-4) to 4 X 10(-4) in both cell lines with pSV2catSVgpt. After 2 to 3 months in selective medium, stable expression of the (nonselected) cat gene was found in 11 (92%) of 12 gpt-containing clones derived from GM0637(SV40) cells and in 13 (81%) of 16 gpt-containing clones from XP20s(SV40) cells. However, the levels of CAT activity did not correlate with those of XPRT activity, and both of these activities varied more than 100-fold among different clones. Copies (1 to 4) of the gpt gene were integrated in four clones of the GM0637(SV40) cells having an XPRT activity of 1 to 5 nmol/min per mg, but 5 to 80 copies were integrated in four XP20s(SV40) clones with an XPRT activity of 0.8 to 1.8 nmol/min per mg. This study shows that XP20s(SV40) is as suitable for gene transfer experiments as the normal human line GM0637(SV40).     

Repair of bleomycin-damaged DNA by human fibroblasts

The ability of human fibroblasts to repair bleomycin-damaged DNA was examined in vivo. Repair of the specific lesions caused by bleomycin (BLM) was investigated in normal cell strains as well as those isolated from patients with apparent DNA repair defects. The diseases ataxia telangiectasia (AT), Bloom syndrome (BS), Cockayne syndrome (CS), Fanconi anemia (FA), and xeroderma pigmentosum (XP) were those selected for study. The method used for studying the repair of DNA after BLM exposure was alkaline sucrose gradient centrifugation. After exposure to BLM, a fall in the molecular weight of DNA was observed, and after drug removal the DNA reformed rapidly to high molecular weight. The fall in molecular weight upon exposure to BLM was observed in all cells examined with the exception of some XP strains. Prelabeled cells from some XP complementation groups were found to have a higher percentage of low molecular weight DNA on alkaline gradients than did normal cells. This prelabeled low molecular weight DNA disappeared upon exposure to BLM.     

Dynamics of structural changes of Paramecium caudatum and Bursaria truncatella macronuclear chromatin and nucleoli under hypotonic conditions

Dynamics of structural changes of nucleoli, complex nucleolar aggregates and chromatin bodies in macronuclei (Ma) of ciliates Paramecium candatum and Bursaria truncatella under hypotonic conditions was studied. It was shown that after a 3 min hypotonic treatment nuclei swelled and became highly vacuolated. 3D-reconstruction showed that such nucleoli were formed by nucleolonema-like threads about 100-200 nm in thickness. Intranucleolar chromatin bodies decompacted, but remained bound with fibrillar component of the nucleolus by thin fibres about 10 nm thick. After 6 min hypotonic treatment the nucleolar material loosened and had a "gauze", or network-like appearence. After 10 min hypotonic treatment nucleoli dissociated completely. It was shown that a transition of chromatin bodies from completely compact to partially and fully decompacted state occurred cooperatively in different regions of Ma. In particular, chromatin bodies in the central part of complex nucleolar aggregates decompacted much faster than those in the Ma karyoplasm. It evidences for a specific, well-ordered chromatin organization in Ma. Prolonged hypotonic treatment led to a complete dissociation of Ma components; fibres 6-10 nm thick were solely observed in such preparations. Such fibres may represent remnant structures of the nuclear matrix. Dynamics of Ma chromatin bodies decompaction correlates well with that of chromomeres in the nuclei of higher eukaryotes. Our data confirm that chromatin 100-200 nm bodies in the ciliate Ma are analogues of chromomeres--looped discrete chromatin domains, observed in the nuclei of higher eukaryotes.     

A monoclonal antibody to lateral element proteins in synaptonemal complexes of Lilium longiflorum

To identify synaptonemal complex (SC) proteins in Lilium longiflorum (lily), monoclonal antibodies were generated using mice immunized with isolated pachytene nuclei. While most of the resulting monoclonal antibodies recognized nucleolar or chromatin proteins, one monoclonal antibody (anti-LE) was found that binds to lateral elements. Anti-LE bound more to lateral elements of SCs digested with DNase than to lateral elements that had not been digested with DNase. The opposite pattern of labeling was observed using monoclonal antibodies to lily chromatin and nucleolar proteins. These results indicate that anti-LE is specifically recognizing lateral element proteins and not chromatin or nucleolar proteins surrounding the lateral elements. On immunoblots, anti-LE binds to three pachytene nuclear proteins (Mr 60000, 66000 and 70000), two tetrad (early microspore) nuclear proteins (Mr 60000 and 70000), and two root tip nuclear proteins (Mr 52000 and 60000). However, anti-LE does not bind to proteins from leaf nuclei. Of these four tissues, leaf is the only one that does not have actively dividing cells. This observation suggests that at least some SC proteins are related to nuclear proteins from mitotically active cells.     

Loss of Rna and Protein, And Changes in Dna During A 30-Hour Cold Perchloric Acid Extraction of Cultured Cells

KB cells derived from human carcinoma were fixed in acetic-alcohol (1:3) and extracted with 10% perchloric acid (PCA) at 4 C for 1, 3, 6, 9, 12, 24 and 30 hr. Cells were then washed in water and stained for nucleic acids, proteins, polysaccharides, and lipids. Control cells were kept in water for 30 hr prior to staining. Acridine orange (AO) fluorochroming revealed color changes in residual cytoplasmic and nucleolar RNA as well as DNA during extraction--interpreted as indicative of molecular alterations. All nucleic acid stains (AO, gallocyanin chromalum, and azure B bromide) demonstrated a differential extraction of RNA, with cytoplasmic RNA being removed in about 6 hr and nucleolar RNA requiring 6 more hours for complete extraction. Large granules appeared early in nuclei. These were positive for DNA by azure B, gallocyanin chromalum, Feulgen, and fluorescent-Feulgen. These same granules stained for protein by mercuric bromphenol blue and alkaline Biebrich scarlet. At 24 hr, there was visual and Feulgen-cytophotometric evidence for a slight loss of DNA, which may amount to 10-20%. There was a progressive loss of cytoplasmic and nuclear but not nucleolar protein during PCA treatment. Concurrently, large protein-positive granules appeared in the cytoplasm. Apparently, PCA treatment in combination with an aqueous wash was responsible for some protein loss. Glycogen was gradually lost (fluorescent PAS) and redistributed in cells. Lipids were unaffected (Sudan black B).     

Fine structural organization of a non-reticulate plant cell nucleus

We studied the fine structural organization of the meristematic nucleus in roots of Lycopesicon esculentum (tomato) using ultracytochemical and immunocytochemical approaches. The nucleus has a non-reticulate (i.e. low DNA content) structure whose supramolecular organization differs in some respects from that in reticulate nuclei, principally in the organization of the chromocentres associated with the nuclear envelope, with which centromeric structures appear to be associated. The main difference at the nucleolar level is found in the fibrillar centres, which have a low amount of DNA labelling and in which inclusions of condensed chromatin are present only very rarely. The distribution of nucleolar DNA amongst the nucleolar compartments is similar to that in reticulate nucleoli as demonstrated using an anti-DNA monoclonal antibody. Tomato nuclei have nucleolus-associated bodies or karyosomes, like other plant species with a low DNA content and non-reticulate nuclear organization. The nuclear ribonucleoprotein structures in the inter- and perichromatin regions, namely inter- and perichromatin fibrils and granules, show similar ultrastructural and cytochemical characteristics in both types of nuclei.Abbreviations NAC nucleolus associated chromatin - CES centromeric structures - NOR nucleolar organizing region - NAB nucleolus associated body - IG interchromatin granules - RNP ribonucleoprotein - Mab monoclonal antibody by M.F. Trendelenburg     

Discrimination of normal and transformed cells in vitro by cytologic and morphologic analysis

Malignant A-549 lung carcinoma and adenovirus-12 SV40 hybrid virus transformed non-tumorigenic human bronchial epithelial cells (BEAS-2B) were objectively discriminated from normal bronchial epithelial (BE) cells on the basis of Papanicolaou stained nuclear features (e.g. shape, chromatin texture, hyperchromasia) and nucleolar morphology (e.g. number per cell, irregular contours). Morphometric analysis indicated that significant differences in cellular morphology existed between BE, BEAS-2B, and A-549 cells. Similar analyses of transformed, tumorigenic cell lines demonstrated that nuclear features (i.e., chromatin texture, clearing of parachromatin, hyperchromasia, variation in thickness of the nuclear envelope, sharp indentations in the nuclear envelope), and nucleolar features (i.e., degree of roundness, presence of angular projections, number per cell) discriminated chemically and virally transformed cells from spontaneously transformed cells. Nuclear and nucleolar features were correlated with the growth rate of tumorigenic cell lines. These analytical approaches will be helpful in studies of the effects of various factors (e.g. vitamin A, phorbol ester, oncogene transfection) on cellular proliferation and/or differentiation.Contribution No. 2708 from the Pathobiology Laboratory, University of Maryland.     

Low-level DNA exchanges in normal human and xeroderma pigmentosum cells after UV irradiation

We have used a T4 endonuclease V assay method for UV-induced pryrimidine dimers in cellular DNA in vivo to obtain evidence for recombinational DNA exchanges after UV irradiation of normal human and Xeroderma pigmentosum (XP) cells. Our data indicate that the endonuclease-sensitive sites in excision-defective XP cells are removed very slowly from the irradiated parental strands and appear concomitantly in daughter strands newly synthesized during post-UV incubation. In the defective XP cells, the extent of appearance of sensitive sites in daughter strands synthesized during a period of 24 h after 10 J/m² appears to be small, probably less than 15% of the initial number of sensitive sites detected in cellular parental strands. Demonstration of such exchanges between normal-density parental and 5-bromodeoxyuridine-labeled daughter strands by alkaline CsCl isopycnic centrifugation was unsuccessful. Further, the extent is much lower in normal human cell because of their efficiet excision repair of the dimers before and after exchanges than in the defective XP cells.     

Structural studies of a membrane-bound acetylcholine receptor from Torpedo californica

We investigated the differential repair of DNA lesions induced by bifunctional mitomycin C, monofunctional decarbamoyl mitomycin C and ultraviolet irradiation in normal human, Xeroderma pigmentosum and Fanconi's anemia cells using assays for the survival of clone-forming ability, alkaline sucrose sedimentation and hydroxyapatite chromatography of DNA. Four FA cell lines exhibited about 5 to 15 times higher sensitivity to MC killing, despite normal resistance to u.v. and DMC, than did normal human cells. The XP cells, however, were highly sensitive to u.v. and DMC killings due to their deficiency in excision repair, but the cells unexpectedly had an almost normal capacity for surviving MC and repairing the MC interstrand cross-links.In experiments to determine the sedimentation velocity of the DNA in alkaline sucrose gradients, normal and XP cells showed evidence for single-strand cutting following MC treatment. The sedimentation velocity of the DNA covalently cross-linked by MC in an FA strain was 2.5 times faster than that of the untreated control, and remained unaltered during post-incubation due to the lack of half-excision⁴ of cross-links. However, FA cells, but not XP cells, had the normal ability to incise DNA with the DMC monoadducts. Hydroxyapatite chromatography revealed the reversibly bihelical property of MC cross-linked DNA after denaturation. Normal and XP cells lost such reversibility during post-MC incubation as the result of cross-link removal with first-order kinetics (half-life = 2 h). The three FA lines studied exhibited two- to eightfold reduced rates of cross-link removal than normal and XP cells, indicating a difference in the repair deficiency of the FA strain. Thus we have been led to conclude that FA cells may have different levels of deficiency in half-excision repair of interstrand cross-links induced by MC, despite having normal mechanisms for repair of u.v.-induced pyrimidine dimers and DMC monoadducts, and vice versa in XP cells.     

Nucleolar changes in bovine nucleotransferred embryos.

This study focused on nucleolar changes in bovine embryos reconstructed from enucleated mature oocytes fused with blastomeres of morulae or with cultured, serum unstarved bovine fetal skin fibroblasts (embryonic vs. somatic cloning). The nucleotransferred (NT) embryos were collected and fixed at time intervals of 1-2 h (early 1-cell stage), 10-15 h (late 1-cell stage), 22-24 h (2-cell stage), 37-38 h (4-cell stage), 40-41 h (early 8-cell stage), 47-48 h (late 8-cell stage), and 55 h (16-cell stage) after fusion. Immunocytochemistry by light and electron microscopy was used for structure-function characterization of nucleolar components. Antibodies against RNA, protein B23, protein C23, and fibrillarin were applied. In addition, DNA was localized by the terminal deoxynucleotidyl transferase (TdT) technique, and the functional organization of chromatin was determined with the nick-translation immunogold approach. The results show that fully reticulated (active) nucleoli observed in donor cells immediately before fusion as well as in the early 1-cell stage after fusion were progressively transformed into nucleolar bodies displaying decreasing numbers of vacuoles from the 2- to 4-cell stage in both types of reconstructed embryos. At the late 8-cell stage, morphological signs of resuming nucleolar activity were detected. Numerous new small vacuoles appeared, and chromatin blocks reassociated with the nucleolar body. During this period, nick-translation technique revealed numerous active DNA sites in the periphery of chromatin blocks associated with the nucleolar body. Fully reticulated nucleoli were again observed as early as the 16-cell stage of embryonic cloned embryos. In comparison, the embryos obtained by fetal cloning displayed a lower tendency to develop, mainly during the first cell cycle and during the period of presumed reactivation. Correlatively, the changes in nucleolar morphology (desegregation and rebuilding) were at least delayed in many somatic NT embryos in comparison with the embryonic NT group. It is concluded that complete reprogramming of rRNA gene expression is part of the general nuclear reprogramming necessary for development after NT.     

Phospholipids in plant and animal chromatin

Isolated hepatic nuclei and hepatic chromatin have been analysed for their DNA, RNA, protein and phospholipid content. The protein/DNA ratio is 3 for nuclei and 1.95 for chromatin extracted from Triton X-100 treated nuclei. The phospholipids, (2.36 +/- 0.91 (S.D.) per cent of the total nuclear material), are lost during the chromatin preparation mainly during the Triton X-100 washings of the nuclei. Nevertheless, 10 per cent of the total nuclear phospholipids remain bound to the chromatin. The comparative analysis of both nuclei and chromatin shows a difference in phospholipids and fatty acid composition. Thus, the chromatin-associated phospholipid cannot be attributed simply to contaminating nuclear membrane. This is supported by the autoradiographic study of semi-thin sections of interphase nuclei from root apices of Vicia faba in which [3H] ethanolamine is clearly localized in the chromatin and nucleolar regions of the nuclei.