Metabolic stability of nonhistone chromosomal proteins in two different classes of developing rat brain nuclei.

The metabolic behaviour of NHCP was studied in rat brain nuclei from fully differetiated cells and in nuclei from still differentiating cells. Eight-day-old rats were injected intracisternally with [14C]thymidine and [3H]tryptophan and the 3H/14C ratio of the chromatin was followed for a period of 33 days. It was found that in the fraction of differentiated cells this ratio remained unchanged, showing the presence of metabolically stable NHCP. At the same time in the fraction containing nondifferentiated cells a substantial part of the NHCP was turned over, which was indicated by a sharp decrease in their 3H/14C ratio with time.     

Follicle-stimulating hormone and human chorionic gonadotropin induced changes in granulosa cell glycosyl-phosphatidylinositol concentration

In the present investigation, a hCG sensitive glycosyl-phosphatidylinositol (GPI) was isolated from cultured rat granulosa cells obtained from the ovaries of diethylstilbestrol (DES) implanted immature rats. The inositol-phosphoglycan (IPG) moiety of the GPI-lipid contains galactose, glucosamine, and myoinositol as demonstrated by metabolic labelling of granulosa cells for different time periods (5–96 h) with [³H]galactose, [³H]glucosamine, or [³H]myoinositol and treatment of the purified [³H]GPI with phosphatidylinositol-specific phospholipase C. Labelling equilibrium of the GPI-lipid was achieved after 24 h ([³H]galactose and [³H]myoinositol) or 72 h ([³H]glucosamine) incubation, whereas incorporation of other labelled carbohydrates tested ([³H]galactosamine, [³H]mannose, and [³H]sorbitol) was negligible throughout the time period studied. The glucosamine C-1 appears to be linked through a glycosidic bond to the myoinositol molecule of the IPG moiety as revealed by the generation of phosphatidylinositol (PtdIns) after nitrous acid deamination of dual labelled ([³H]glucosamine/[¹⁴C]palmitate or [³H]glucosamine/[¹⁴C]myristate) glycosyl-phosphatidylinositol. To investigate the fatty acid composition of the diacylglycerol (DAG) backbone of the GPI, granulosa cells were also labelled (5–72 hr) with [¹⁴C]linoleate, [³H]myristate, [³H]-oleate, [³H]palmitate, or [³H]stearate and the radioactivity associated with the purified glycosyl-phosphatidylinositol determined. Incorporation of [³H]palmitate and [³H]myristate into the GPI-lipid peaked after 8 h and 24 h of labelling, respectively, and both fatty acids were partially released after PLA₂ treatment of the dual labelled ([³H]glucosamine/[¹⁴C]palmitate or [³H]glucosamine/[¹⁴C]myristate) GPI. In parallel experiments no significant incorporation of labelled stearate, oleate, or linoleic acid into the DAG backbone of the glycosylphosphatidylinositol could be detected. Granulosa cells were also labelled with [³H]glucosamine in the presence of FSH (30 ng/ml), cholera toxin (1 μg/ml), or the membrane permeable cAMP analog (but)₂ cAMP (1 mM). Time related increases in GPI-labelling were apparent after 48 h and reached a maximum level (3-, 5-, and 7-fold for FSH, CT, and (but)₂ cAMP, respectively) after 72 h in culture. In another set of experiments, granulosa cells were labelled for 72 h with [³H]glucosamine in the presence of (but)₂cAMP (1 mM), TPA (10^?7 M), or combination thereof. The effect of treatment with the membrane permeable cAMP analog on GPI labelling was prevented in the presence of TPA, whereas no differences in [³H]GPI content could be observed in untreated granulosa cells or cells cultured in the presence of the protein kinase C-activating phorbol ester alone. In cells differentiated with FSH (30 ng/ml for 3 days) to induce LH receptors, treatment with hCG (100 ng/ml) induced a rapid (60 sec) and transient (5 min) decrease in the GPI content, whereas no efect of the hormone on undifferentiated granulosa cells could be observed. The rapid effect elicited by hCG on GPI content and turnover may be an early transduction mechanism involved in the biological effects of LH/hCG in differentiated granulosa cells. © 1993 Wiley-Liss, Inc.     

Inhibition by 1-beta-D-arabinofuranosylcytosine of the incorporation of N-acetylneuraminic acid into glycolipids and glycoproteins of hamster embryo cells

1-β-D-Arabinofuranosylcytosine which interferes with DNA synthesis in bacteria and mammalian cells and brings about transformation of hamster embryo fibroblasts, has been found to inhibit the incorporation of N-Acetylneuraminic acid into glycolipids and glycoproteins of both normal and transformed hamster embryo cells in tissue culture. Three hours after commencement of treatment (10^?3M ara-C), incorporation of [¹⁴C] thymidine into DNA was inhibited by 95 per cent, while incorporation of [³H] D-glycosamine (precursor of sialic acid) into glycolipids and glycoproteins was inhibited by 85 per cent. At 24 hours, the inhibition of incorporation of the two labelled components was 83 and 80 per cent respectively. In homogenates of both cell types, incorporation of [¹⁴C] N-acetylneuraminic acid was competitively inhibited by ara-CMP. Ara-C was found to have no effect on the incorporation of [¹⁴C] choline into phospholipids of cells grown in tissue culture. These results suggest that interference with DNA synthesis by ara-C may not be the only factor involved in cell transformation by this substance.     

Preferential incorporation of [3H]thymidine into polypyrimidine-containing regions in DNA following ultraviolet irradiation of human cells

Ultraviolet irradiation of cells causes damage to DNA, principally to pyrimidine bases. Regions of DNA which are very rich in pyrimidines are therefore likely to be more susceptible to damage by this agent. Polypyrimidines (pyrimidine tracts > 25 nucleotides in length) are known to occur in DNA from many higher organisms. We investigated whether these tracts are particularly sensitive to ultraviolet light by irradiating human diploid flbroblasts and preparing DNA which was labelled with [³H]thymidine during post-irradiation repair replication. Polypyrimidine-containing regions of the DNA were isolated and their content of [³H]thymidine (principally arising from repair synthesis) was compared to their content of [¹⁴C]thymidine (representing bulk DNA). It was found that polypyrimidine-containing regions were enriched (approximately twofold) in ³H compared to ¹⁴C, probably because of greater susceptibility of pyrimidine-rich regions in DNA to ultraviolet light.     

Intracellular degradation of glycoproteins in Acer pseudoplatanus L. cells

The degradation of endogenously labelled glycoproteins was studied in Acer pseudoplatanus L. cell suspension cultures in experiments using a dual-label with [¹⁴C]mannose and [³H]leucine.After harvesting the cells, protoplasts were prepared and vacuoles isolated. More than 30% of both total newly synthesized proteins (³H radioactivity) and glycoproteins (¹⁴C radioactivity) were recovered inside the vacuoles, the lytic compartment of plant cells. Half of these proteins were degraded when isolated vacuoles were incubated for 6 h at 20°C. So, the vacuolar compartment appears to be a major site of glycoprotein degradation in the cell.The glycoproteins were degraded at the same rate as the total newly synthesized proteins. However, some vacuolar hydrolytic enzymes were found to be glycoproteins and resistant to proteolytic attack. The biochemical explanation for such a resistance is not clear at this time, but in Acer cells the presence of covalently bound carbohydrates in proteins does not seem to be involved in the selectivity of protein turnover.     

An investigation of acid-soluble nuclear proteins of human leucocytes in relation to fraction RP2-L, a component of neoplastic cells

1. The claim that tumour cells contain a specific nuclear protein was investigated. The presence of this component was confirmed in Walker tumour cells by the chromatography on CM-cellulose of nuclear proteins labelled with [¹⁴C]lysine. This protein was studied further in a number of human leucocyte cells. 2. The labelling of leucocyte nuclear proteins with [¹⁴C]lysine was attempted during incubation and culture in vitro. Incorporation of the label into acid-soluble nuclear proteins was highest in normal lymphocytes cultured with phytohaemagglutinin, followed by chronic-myeloid-leukaemic leucocytes and mixed samples of normal leucocytes incubated in plasma. Little incorporation was seen in similar extracts of chronic-lymphatic or normal leucocytes. 3. Lymphocytes were the only cells that gave nuclear extracts with amino acid analysis similar to that of unfractionated histones. 4. Little of the [¹⁴C]lysine in nuclear extracts of incubated leucocytes proved to be of chromosomal origin. No evidence was found of an RP2-L component in the highly labelled nuclear extracts of phytohaemagglutinin-treated lymphocytes until after 6 days of culture with [¹⁴C]lysine. This component was soluble in saline. 5. Evidence is presented that fraction RP2-L is a non-histone protein constituent of cell nuclei whose labelling with [¹⁴C]lysine may be dependent on the metabolic state of the cell. Thus this component is not specific to the neoplastic state.     

Utilization of the label from bacterial [3H] DNA by isolated barley roots and embryos under different conditions

[³H] DNA fromEscherichia coli and [³H] thymidine were applied, in sterile conditions, on isolated barley embryos and on roots excised from these embryos, both cultivated in the liquid medium and on halves of barley seeds, through the endosperm bridge. In embryos and roots, the labelled compounds were applied in 1.5% sucrose + 0.2 SSC alone, or together with either unlabelled thymidine or DEAE-dextran. Similar labelling indices were found after [³H] thymidine and [³H] DNA treatment which shows that the activity of [³H] DNA is utilized during the S phase. After application of [³H] thymidine, only cell nuclei in S phase were labelled. After the application of [³H] DNA an extranuclear label, in addition to the labelling of nuclei in the S phase, was observed in some experimental variants. The density of label above labelled nuclei after [³H] DNA treatment sharply decreased when unlabelled thymidine or DEAE-dextran was added, while the density of label above nuclei labelled by [³H] thymidine decreased when unlabelled thymidine but not DEAE-dextran was added. The labelling of nuclei with the label from [³H] DNA is the result of degradation of exogenous DNA reutilization of low molecular weight products. Extranuclear labelling is most probably due to the polymerous or partly degraded DNA.     

Cytological detection of poly (ADP-ribose) polymerase

An attempt was made to demonstrate poly (ADP-ribose) polymerase cytologically. In vitro incorporation from the nucleotide, [³H]NAD was detected in frozen sections of onion embryo and meristematic tissue by autoradiography. In meristematic tissue, there was a correlation between the number of cells displaying intensein vitro incorporation from [³H]NAD and cytological DNA polymerase activity. Performed enzymes effecting a distinct incorporation from [³H]NAd were localized in the nuclei of all tissues of the ungerminated seed except the endosperm. Evidence for poly (ADP-ribose) polymerase has been obtained for the first time from higher plant cells and localized cytologically.     

Chromosomal proteins of conifers

During imbibition and germination of jack pine seeds, the composition of the total extractable chromatin varied. Relative to DNA, the histone levels decreased as the nonhistone chromosomal proteins (NHCP) increased. New chromosomal proteins were synthesized after 2 days of imbibition as judged by recovery of ¹⁴C-amino acids from the major protein fractions. Phosphorylation of histones from ³²P-phosphoric acid was detected before the incorporation of ¹⁴C-amino acids. In the seed the synthesis and relative changes of chromatin coincided with a fall in total soluble protein and free arginine N. By contrast, adenylate energy charge, free glutamine N and in vitro template activity of chromatin increased during chromatin protein synthesis. When seeds had germinated for 4 days after the start of imbibition more radioactivity, derived from free ¹⁴C-amino acids, was recovered from the NHCP than from the histones. The percentage amino acid composition of most histone fractions remained stable, whereas the composition of NHCP changed more with time. The phosphorylation of NHCP was 8- to 41-fold greater than that of the histones. Phosphorylation of histone H4 was not detected at any stage of germination. Correlations between recovery of radioactivity (³²P and ¹⁴C) from chromosomal proteins and higher adenylate energy charge were positive.     

THE INFLUENCE OF PORTOCAVAL ANASTOMOSIS ON THE METABOLISM OF LABELLED OCTANOATE, BUTYRATE AND LEUCINE IN RAT BRAIN

Rats were given a portocaval anastomosis and 3 weeks later, when the only ultrastructural change in the CNS is watery swelling of astrocytes, several aspects of brain metabolism were studied. The uptake of leucine by the brain, its incorporation into protein and its oxidation were followed after the simultaneous injection of a mixture of L-[1¹⁴C]leucine and L-[4,5-³H]leucine. The concentration of leucine in blood was lowered in the operated animals whereas in brain it was increased. The specific radioactivity of leucine in the brain was comparable to values in control animals and there was no evidence of a decrease in incorporation of [1-¹⁴C]leucine into brain proteins over the short experimental time period studied. The only difference from the controls in the oxidation of [4,5-³H]leucine was a greater accumulation in glutamine. The amount of glutamine in the brains of the operated animals had increased 4-fold at the time of the metabolic studies. From dual-labelled experiments in which a mixture containing [1-¹⁴C]butyrate and L-[4,5-³H]leucine was injected intravenously, it was shown that, in both control and operated animals, the pools of brain glutamate and glutamine labelled from butyrate were metabolically distinct from those labelled from leucine. The total radioactivity appearing in brain from [1-¹⁴C]butyrate was markedly reduced in operated animals, but the radioactivity from L-[4,5-³H]leucine was not. The metabolism of [1-¹⁴C]octanoate was compared with that of [1-¹⁴C]butyrate. In control animals the labelling of metabolites was almost identical with either precursor. In operated animals there was no reduction in the uptake of [1-¹⁴C]octanoate into the brain. There was evidence that the size of the glutamine pool labelled, relative to glutamate, was increased but that it had a slower fractional turnover coefficient. A link between astroglial changes and an impairment to the carrier mechanism for transport of short chain monocarboxylic acids across the blood-brain barrier is suggested.     

CATECHOLAMINE METABOLISM IN THE DOG: COMPARISON OF INTRAVENOUSLY AND INTRAVENTRICULARLY ADMINISTERED [14C]DOPAMINE AND [3H]NOREPINEPHRINE

—The urinary excretion of labelled metabolites was measured in dogs which had been injected intravenously or intraventricularly with [³H]norepinephrine or [¹⁴C]dopamine. [³H]Norepinephrine injected by either route produced more labelled 3-methoxy-4-hydroxy-phenylglycol than 3-methoxy-4-hydroxymandelic acid, as did [¹⁴C]dopamine after intravenous administration. In contrast, following the intraventricular injection of [¹⁴C]dopamine, more [¹⁴C]3-methoxy-4-hydroxymandelic acid was formed than [¹⁴C]3-methoxy-4-hydroxyphenylglycol. These observations suggest that the metabolism of exogenously-administered and endogenously-formed norepinephrine may proceed through different routes and that the predominant metabolite of norepinephrine in canine brain may be 3-methoxy-4-hydroxymandelic acid rather than 3-methoxy-4-hydroxyphenylglycol.     

Incorporation of various amino acids into non-histone chromatin protein fractions of spleen cells of mice immunized with IgG

Summary Incorporation of three various amino acids ([³H]-tryptophan, [³H]-methionine or [³H]-leucine) into the non-histone chromatin proteins, synthesized in spleen cells of mice after immunization with IgG, is described. Two new fractions of non-histone chromatin proteins (I-mol. wt. below 3 000 and B₁-mol. wt. about 120 000), appearing during the immune reaction were labelled with [³H]-tryptophan and [³H]-methionine but not with [³H]-leucine. Synthesis of these fractions was observed only at the time of maximal RNA synthesis. A suggestion about the role of tryptophan and methionine in non-histone chromatin proteins in the regulatory processes of gene activation is discussed on the basis of their selective binding to DNA.     

Effect of pyridoxine deficiency on nucleic acid metabolism in the rat

1. The nucleic acid metabolism in the pyridoxine-deficient rat has been investigated through studies on the incorporation of radioactivity from various isotopically labelled compounds into liver and spleen DNA and RNA. 2. In pyridoxine deficiency, the incorporation of radioactivity from sodium [¹⁴C]formate was apparently increased. The magnitude of this effect on incorporation into liver RNA and DNA and spleen RNA was approximately the same. The incorporation into spleen DNA was enhanced to a much greater degree. Administration of pyridoxine 24hr. before the rats were killed reversed the changes in incorporation of radioactivity from [¹⁴C]formate. 3. In pyridoxine deficiency, the incorporation of radioactivity from dl-[3-¹⁴C]serine, [8-¹⁴C]adenine, [Me-³H]thymidine and [2-¹⁴C]deoxyuridine was decreased. The incorporation of radioactivity from l-[Me-¹⁴C]methionine was not affected. No noteworthy differences in the effect of pyridoxine deficiency on the incorporation of radioactivity from dl-[3-¹⁴C]serine into DNA and RNA were observed, whereas the effect of the deficiency on the incorporation of radioactivity from [8-¹⁴C]adenine into spleen DNA was somewhat greater than that into spleen RNA. Administration of pyridoxine 24hr. before the rats were killed reversed the changes in incorporation of radioactivity from [3-¹⁴C]serine and [8-¹⁴C]adenine. 4. The adverse effects of pyridoxine deficiency on the biosynthesis of nucleic acids and cell multiplication are discussed in relation to the role of pyridoxal phosphate in the production of C₁ units via the serine-hydroxymethylase reaction.     

DIFFERENT LABELLING PATTERNS IN MOUSE LYMPHOID TISSUES WITH [3H]DEOXYCYTIDINE AND [3H]THYMIDINE

The percentages of labelled lymphocytes in smear preparations of mouse thymus were higher than those in similar preparations of mesenteric lymph nodes with either generally labelled tritiated deoxycytidine, [³H]CdR, or tritiated thymidine, [³H]TdR. Lymphocytes in the thymus cortex and in germinal centres of mesenteric lymph nodes were intensely labelled with [³H]CdR, whereas with [³H]TdR lymphocytes in the peripheral region of thymus and medullary cords of mesenteric lymph nodes were heavily labelled. The majority of lymphocytes in thymic cortex and germinal centres of mesenteric lymph nodes were labelled weakly with [³H]TdR. Thus, labelling patterns with [³H]CdR differed from those with [³H]TdR in lymphoid tissues of the mouse. Mouse lymphocytes can utilize [³H]CdR as a precursor molecule for cytosine and thymine in DNA. The ratio of radioactivity of thymine to that of cytosine was measured biochemically in DNA extracted from lymphocytes labelled with [³H]CdR. This radioactivity ratio in thymus was higher than that in mesenteric lymph nodes. These results suggest that the metabolic activities of utilizing CdR for DNA synthesis differ within lymphocyte populations in various lymphoid tissues in the mouse.     

Studies on the turnover of plasma membranes in cultured mammalian cells.: II. Rates of synthesis and degradation of plasma membrane proteins and carbohydrates

The rate of turnover of membrane proteins and membrane-bound carbohydrates in exponentially growing and in confluent contact-inhibited cultures of strain MK-2 cells was investigated. Cells were labelled with [¹⁴C]leucine and [³H]glucosamine, incubated in isotope-free medium and, at various times thereafter, the cells were harvested and membranes isolated from them. The rate of decay of the protein and carbohydrate components was determined from specific activity dilution of the labeled components in the isolated membranes.Although the rate of membrane synthesis is different in exponential and contact-inhibited cells, the rate of degradation (turnover) of membrane proteins and carbohydrates was found to be the same (25%, per generation (42 h) or 0.6%/h).     

Specific protein synthesis in isolated epithelium of guinea-pig seminal vesicle. General features

Four intrinsic soluble proteins are synthesized and secreted by sexually mature guinea-pig seminal-vesicle mucosa, which comprises a monolayer of a homogeneous columnar epithelial cell. All four proteins can be extracted readily in 154mm-NaCl from the organ's luminal constituents in which they are present in high concentration. They are referred to as proteins 1, 2, 3 and 4 in order of their elution during DEAE-cellulose column chromatography. Specific primary antibodies were harvested from goats that had been inoculated with the purified vesicular proteins; secondary antibodies were obtained from a donkey inoculated with goat γ-globulins. Double-antibody-immunoprecipitation techniques were developed to precipitate the vesicular proteins. Thus proteins newly synthesized from ¹⁴C-labelled amino acids could be precipitated and the incorporated radioactivity assessed. Isolated seminal-vesicle mucosa, incubated in only a buffered salt solution containing glucose, readily synthesized the soluble secreted proteins from added [¹⁴C]lysine plus [¹⁴C]glycine, [¹⁴C]histidine plus [¹⁴C]glutamate, [¹⁴C]glutamine alone and [¹⁴C]arginine alone. The rates of incorporation (d.p.m./mg of total soluble protein) of labelled lysine and glycine and of labelled arginine were linear with time over 180min. With the other labelled precursors, rates diminished between 60 and 180min. Labelled protein could be detected after only 10–15min of incubation. Only 4–9% of the newly synthesized protein remained associated with the mucosa; the remainder was found in the cell-free incubation medium. The isolated seminal-vesicle mucosal preparation will provide a unique opportunity to study the synthesis and secretion of abundant cell-specific proteins by this androgen-dependent tissue.     

The interaction of carbon-14-labelled alkyl carbamates, labelled in the alkyl and carbonyl positions, with DNA in vivo

The binding of ethyl carbamate labelled with carbon-14 in the alkyl or carbonyl group, and of methyl, n-butyl and n-propyl carbamates labelled in the alkyl group, to the DNA of mouse liver, lung and kidney has been studied in male Crackenbush mice. Only ethyl carbamate bound to liver and kidney DNA to any significant extent.The binding of ethyl carbamate labelled with carbon-14 in the C₁, C₂ or the carbonyl position was examined and compared. The levels of binding of [1-¹⁴C]- and [2-¹⁴C]ethyl carbamate to liver DNA were not significantly different (328 ± 34 and 267 ± 24 dpm/mg DNA, respectively), but there was very little binding of the [carbonyl-¹⁴C]ethyl carbamate (26 ± 3 dpm/mg DNA). Furthermore, only 18% of the radioactivity was removed from the DNA labelled with the alkyl-labelled carbamates, whereas 65% of the radioactivity was removed from the DNA labelled with carbonyl-labelled ethyl carbamate on continuous ether extraction. It was concluded that the bound molecule does not contain the carbonyl carbon and is probably an ethyl group.     

Discrepancies between flow cytometric analysis and [3H]thymidine incorporation in stimulated lymphocytes

The DNA synthesis system of freshly isolated tonsillar lymphocytes and those stimulated by phytohaemagglutinin were compared by different methods. Both cell populations had high DNA polymerase α and thymidine kinase activities, as well as a high rate of incorporation of [³H]thymidine into DNA. However, the two cell populations differed when their DNA distributions were compared by flow cytometry. Freshly isolated cells contained many less (6%) cells in S phase than were found in phytohaemagglutinin-stimulated lymphocytes (18%) as detected by flow cytometry. The labelling of different subpopulations of lymphocytes was studied by sorting them electrically with a fluorescence-activated cell sorter. Analysis of the radioactivity of [³H]thymidine pulse-labelled cells, sorted according to their DNA content, showed that cells in the G₁ peak of DNA distribution had a significant amount of incorporated [³H]thymidine. Sorting of cells according to their size (i.e., by light scattering) revealed that only large cells were labelled with [³H]thymidine.     

Measurement of protein turnover in rat brain

Abstract— Degredation rates of rat brain proteins were measured by following the decay in specific radioactivity of carboxyl labelled aspartate and glutamate over a 17-day period. Initial labelling of these amino acids was achieved by a single intraperitoneal injection 0f NaH¹⁴CO₃. The non-linear decay curve for total brain proteins could be approximated by assuming that the mixture contained two classes of proteins with half-lives of 3.3 and 8.7 days, respectively. Half-lives of 2.5 and 7.7 days were estimated for such protein classes in the microsomal fraction. The half-lives of soluble proteins, synaptic membranes, cell body and synaptic mitochondria were 3.1, 5.8, 5.6 and 8.4 days, respectively. Identical results were obtained if the change in specific activity of intact protein labeled by NaH¹⁴CO₃ was followed. Two-fold slower decay rates were obtained when brain proteins were labeled with a pulse of [4,5-³H]leucine or [l-¹⁴C]leucine. Half-lives calculated for the two classes of proteins in whole brain were 8.4 and 16.5 days, respectively with [4,5-³H]leucine and 8.9 and 14.2 days, respectively with [1-¹⁴C]leucine. These results indicate the very significant reutilization of this amino acid in brain. Sodium [¹⁴C]bicarbonate is a more satisfactory isotopic precursor for accurate assessment of rates of protein turnover in brain.     

Cell cycle-related transfer of proteins between the nucleus and cytoplasm of Physarum polycephalum

The proteins of wild-type and polyploid plasmodia of P. polycephalum were prelabelled with [³H]leucine and [¹⁴C]leucine. The two types of plasmodia were then fused for 2 h. Following fusion the nuclei were isolated and the smaller wild-type cell nuclei separated from the larger polyploid cell nuclei. The proteins were isolated from the recipient cell nuclei and the recipient nuclear proteins extracted. Ratios of ³H/¹⁴C in the various nuclear protein fractions show that during fusion differential transfer of labelled preformed proteins from the donor cell into the recipient cell nucleus occurs. The quantity of proteins transferred varies among the different fractions and with the phase of the cell cycle. Isotopic dilution experiments indicate that these differences in protein transfer are, in part, due to a high rate of synthesis and turnover of the nuclear proteins.