THE IN VIVO PROTEIN SYNTHETIC ACTIVITIES OF FREE VERSUS MEMBRANE-BOUND RIBONUCLEOPROTEIN IN A PLASMA-CELL TUMOR OF THE MOUSE

Cytoplasmic extracts of the transplantable RPC-20 plasma-cell tumor were fractionated by sucrose density gradient centrifugation. Four major fractions were distinguished: (a) microsomes and mitochondria; (b) membrane-free polyribosomes; (c) free monomeric ribosomes; and (d) soluble fraction. The fractions were analyzed for RNA and lipid phosphorus, and their particulate components were characterized by electron microscopy. Particular attention was paid to the problem of membrane contamination of the free polyribosome fraction. It was shown that this contamination was small in relation with the total content of ribosomes in the fraction, and that it consisted primarily of smooth-surfaced membranes which were not physically associated with the polyribosomes themselves. In vivo incorporation studies were carried out by injecting tumor-bearing animals intravenously with leucine-C¹⁴, removing the tumors at various times thereafter, and determining the distribution of protein radioactivity among the gradient-separated cytoplasmic fractions. The free polyribosome and the microsome-mitochondria fractions constituted active centers for protein synthesis. It was shown that nascent protein of the free polyribosome fractions was not associated significantly with the contaminating membranes. The kinetics of labeling during incorporation times up to 11 min suggested that protein synthesized on the free polyribosomes was rapidly transferred in vivo to the soluble fraction of the cell, while protein synthesized by the microsomes and mitochondria remained localized within these elements. It was estimated that the free polyribosome fraction and the microsome-mitochondria fraction accounted for approximately equal proportions of the total cytoplasmic protein synthesis in vivo.     

TRANSPORT OF VIRAL RNA IN KB CELLS INFECTED WITH ADENOVIRUS TYPE 2

Messenger RNA transport was studied in KB cells infected with the nuclear DNA virus adenovirus type 2. Addition of 0.04 µg/ml of actinomycin completes the inhibition of ribosome synthesis normally observed late after infection and apparently does not alter the pattern of viral RNA synthesis: Hybridization-inhibition experiments indicate that similar viral RNA sequences are transcribed in cells treated or untreated with actinomycin. The polysomal RNA synthesized during a 2 hr labeling period in the presence of actinomycin is at least 60% viral specific. Viral messenger RNA transport can occur in the absence of ribosome synthesis. When uridine-³H is added to a late-infected culture pretreated with actinomycin, viral RNA appears in the cytoplasm at 10 min, but the polysomes do not receive viral RNA-³H until 30 min have elapsed. Only 25% of the cytoplasmic viral RNA is in polyribosomes even when infected cells have been labeled for 150 min. The nonpolysomal viral RNA in cytoplasmic extracts sediments as a broad distribution from 10S to 80S and does not include a peak cosedimenting with 45S ribosome subunits. The newly formed messenger RNA that is ribosome associated is not equally distributed among the ribosomes; by comparison to polyribosomes, 74S ribosomes are deficient at least fivefold in receipt of new messenger RNA molecules.     

Polyribosome metabolism, growth and water status in the growing tissues of osmotically stressed plant seedlings

Relationships between growth of osmotically stressed intact seedlings and polyribosome levels and water status of growing tissues were examined. Sudden exposure of barley (Hordeum vulgare L. cv. Arivat) roots to a solution of ?0.8 MPa polyethylene glycol caused leaf growth to stop almost immedately, but growth resumed at a much lower rate after 0.5–1 h. In the growing region of leaves, the polyribosome: total ribosome ratio of free (non-membrane-bound) ribosomes was significantly reduced after 15 min stress, but a decrease in the large polyribosome:total polyribosome ratio occurred only after 1–2 h. Membrane-bound and free polyribosome levels both decreased to 70% of unstressed control values after 4 h stress. Recovery of total polyribosomes occurred within 1 h after relief of 4 h stress, but required 3 h after relief of 24 h stress. Stress detectably reduced the water potential and osmotic potential of growing tissue within 0.5–1.0 h, and osmotic adjustment continued for up to 10 h. Recovery of water status was incomplete after 1 h relief of a 4 h stress. In contrast, expanded blade tissues of stressed plants underwent minor changes in water status and slow decreases in polyribosomes levels. These results confirm that growing tissues of barley leaves are selectively responsive to stress, and suggest that changes in growth, water status and polyribosome levels may be initiated by the same signal. Measurements of seedling growth, polyribosome levels and water status of growing tissues of barley and wheat (Triticum aestivum L. cv. Zaragoza) leaves, etiolated pea (Pisum sativum L. cv. Alaska) epicotyl and etiolated squash (Cucurbita pepo L. cv. Elite) hypocotyl stressed with polyethylene glycol solutions of ?0.3 to ?0.8 MPa for 12 h or more showed that polyribosome levels were highly correlated with seedling growth rate as well as with tissue water and osmotic potentials, while turgor remained unchanged. These results suggest that long-term growth of osmotically stressed plants may be limited by a reduced capacity for protein synthesis in growing tissues and is not dictated by turgor loss.     

Hormonal control of polyribosome formation in barley aleurone layers

The addition of abscisic acid to barley (Hordeum vulgare L. cv. Himalaya) aleurone layers at the same time as gibberellic acid completely prevents the gibberellin-induced increases in the percentage of polysomes, the formation of polyribosomes, and the synthesis of α-amylase, even when the molar concentration of gibberellic acid is four times greater than the concentration of abscisic acid. The addition of abscisic acid to aleurone cells producing α-amylase (midcourse addition) inhibits the further synthesis of α-amylase and decreases the percentage of polysomes but does not change the number of ribosomes per cell.     

Polyamine metabolism and its relation to response of the aleurone layers of barley seeds to gibberellic Acid

Polyamine metabolism and its relation to the induction of α-amylase formation in the aleurone layers of barley seeds (Hordeum vulgare cv Himalaya) in response to gibberellic acid (GA₃) has been investigated. A high-performance liquid chromatographic system has been employed for qualitative and quantitative analyses of putrescine (Put), cadaverine (Cad), spermidine (Spd), spermine (Spm), and agmatine (Agm).

Active polyamine metabolism occurs in the aleurone cells of deembryonate barley half seeds during imbibition. The aleurone layers isolated from fully imbibed half seeds contain about 880 nanomoles of Put, 920 nanomoles of Spd, and 610 nanomoles of Spm as free form per gram tissue dry weight while the levels of Cad and Agm are relatively low. The polyamine levels do not change significantly in the aleurone layers in response to added GA₃ (1.5 micromolar) during the 8-hour lag period of the growth substance-induced formation of α-amylase. Also, the polyamine levels are not altered by the presence of abscisic acid (3 micromolar) which inhibits the enzyme induction by GA₃. Kinetic studies show that both applied [U-¹⁴C]ornithine and [U-¹⁴C]arginine are primarily incorporated into Put during 2 hours of incubation, but the incorporation is not significantly affected by added GA₃. Additionally, added GA₃ does not affect the uptake and turnover of [1,4-¹⁴C]Put, nor does it affect the conversion of Put → Spd or Spd → Spm. Treatment of the aleurone layers with GA₃ for 2 hours results in no significant changes in the total activities or the specific activities of ornithine decarboxylase and arginine decarboxylase.

Experiments with polyamine synthesis inhibitors demonstrate that the level of Spd in the aleurone layers could be substantially reduced by the presence of methylglyoxal-bis(guanylhydrazone) (MGBG) during imbibition. MGBG treatment does not affect in vivo incorporation of [8-¹⁴C] adenosine into ATP. The lower the level of Spd the less α-amylase formation is induced by added GA₃. The reduction of GA₃-induced α-amylase formation by MGBG treatment can be either completely or partially overcome by added Spd, depending upon the concentration of MGBG used in the imbibition medium. The results indicate that the early action of GA₃, with respect to induction of α-amylase formation in barley aleurone layers, appears to be not on polyamine metabolism. However, polyamines, particularly Spd, may be involved in regulation of the growth substance-dependent enzyme induction.

     

Hormonal control of enzyme synthesis: on the mode of action of gibberellic Acid and abscisin in aleurone layers of barley

Gibberellic acid (GA) enhances the synthesis of α-amylase and ribonuclease in isolated aleurone layers and this process is inhibited by abscisin. Removal of gibberellic acid in mid-course of α-amylase production results in a slowing down of α-amylase synthesis, suggesting a continued requirement of GA for enzyme synthesis. This is paralleled by a continuous requirement for RNA synthesis. Addition of 6-methylpurine or 8-azaguanine in mid-course results in an inhibition of α-amylase synthesis within 3 to 4 hours. However, actinomycin D added in mid-course is almost without effect. This is not due to its failure to enter the cells, because it does inhibit ¹⁴C-uridine incorporation at this stage. Addition of abscisin to aleurone layers which are synthesizing α-amylase results in an inhibition of this synthesis within 2 to 3 hours. Cycloheximide on the other hand inhibits enzyme synthesis immediately upon its addition. These data are consistent with the hypothesis that the expression of the GA effect requires the synthesis of enzyme-specific RNA molecules. The similarity in the kinetics of inhibition between abscisin on the one hand and 8-azaguanine or 6-methylpurine on the other suggests that abscisin may exert its action by inhibiting the synthesis of these enzyme-specific RNA molecules or by preventing their incorporation into an active enzyme-synthesising unit.     

Inhibition of Gibberellic Acid-induced alpha-Amylase Formation by Polyethylene Glycol and Mannitol

Both polyethylene glycol (PEG) and mannitol inhibit gibberellic acid-induced α-amylase production in barley aleurone layers. The effect of the osmotic solution is on enzyme synthesis rather than α-amylase secretion. The inhibition of α-amylase synthesis does not appear to be mediated via an indirect effect on respiration or protein synthesis. Rather it seems that the osmotic solutions reduce the extent of proteolysis of the stored aleurone grain protein thus making available less substrate for new protein synthesis.     

Response of barley aleurone layers to abscisic Acid

Cordycepin, an inhibitor of RNA synthesis in barley (Hordeum vulgare L.) aleurone cells, does not inhibit the gibberellic acid-enhanced α-amylase (EC 3.2.1.1.) synthesis in barley aleurone layers if it is added 12 hours or more after the addition of the hormone. However, the accumulation of α-amylase activity after 12 hours of gibberellic acid can be decreased by abscisic acid. The accumulation of α-amylase activity is sustained or quickly restored when cordycepin is added simultaneously or some time after abscisic acid, indicating that the response of aleurone layers to abscisic acid depends on the continuous synthesis of a short lived RNA. By analysis of the newly synthesized proteins by gel electrophoresis with sodium dodecylsulfate, we observed that the synthesis of α-amylase is decreased in the presence of abscisic acid while the synthesis of most of the other proteins remains unchanged. From the rate of resumption of α-amylase production in the presence of cordycepin and abscisic acid, it appears that abscisic acid does not have a measurable effect on the stability of α-amylase mRNA.     

Rates of synthesis of ribosomal protein and total ribonucleic acid through the cell cycle of the fission yeast Schizosaccharomyces pombe

The rates of synthesis of ribosomal proteins through the cell cycle of the fission yeast Schizosaccharomyces pombe have been examined by spec. act. estimations of isolated 80S ribosomes pulse-labelled with ³⁵S-sulphate. The spec. act. have minimum values at the beginning (0.0) and maxima between 0.6 and 0.9 of the cell cycle. This pattern in spec. act. is also shown by isolated 80S ribosomes pulse-labelled with ³H-uridine during synchronous cultures and is in marked contrast to the small, random variations in the spec. act. of isolated 80S ribosomes from control, asynchronous cultures pulse-labelled with ³⁵S-sulphate or ³H-uridine.A detailed examination of the rates of synthesis of total RNA through the cell cycle measured by the rates of incorporation of ³H-uridine and ³H-adenine shows a step in the rates of incorporation at the time of DNA synthesis. This step has further been shown to be independent both of the uridine concentration, over a range from 0.03 μM to 820 μM, and of pre-filling the adenine pool. This step thus appears to be independent of variations in rates of uptake of both purines and pyrimidines, or fluctuations in the pool size of the precursors and may be explained as a gene-dosage effect.The step in the pattern of synthesis of total RNA has been shown to yield a cyclic pattern in the spec. act. of the total RNA through the cell cycle. This pattern is similar to that of the spec, act. of RNA and of protein recovered from ribosomes. The variation exhibited by the ribosomal proteins is believed to be a consequence of the step in the pattern of RNA synthesis, with a concomitant fluctuation in the pool of ribosomal proteins synthesised continuously through the cell cycle.     

Nuclear protein synthesis in regenerating rat liver : Selective histone inhibition by α-amanitin

Nuclear protein synthesis has been studied in regenerating rat hepatocytes after partial hepatectomy and α-amanitin treatment. The toxin induced a marked and precocious inhibition of histone synthesis without affecting the acidic nuclear proteins. This inhibition preceded the inhibition of DNA synthesis. The modification of polyribosome profile and of [¹⁴C]lysine incorporation on synthesized polypeptides were consistent with a reduction of specific mRNAs.     

Impact of oxidative stress on the function,abundance, and turnover of the Arabidopsis 80S cytosolic ribosome

Abiotic stress in plants causes accumulation of reactive oxygen species (ROS) leading to the need for new protein synthesis to defend against ROS and to replace existing proteins that are damaged by oxidation. Functional plant ribosomes are critical for these activities, however we know little about the impact of oxidative stress on plant ribosome abundance, turnover, and function. Using Arabidopsis cell culture as a model system, we induced oxidative stress using 1 µm of H₂O₂ or 5 µm menadione to more than halve cell growth rate and limit total protein content. We show that ribosome content on a total cell protein basis decreased in oxidatively stressed cells. However, overall protein synthesis rates on a ribosome abundance basis showed the resident ribosomes retained their function in oxidatively stressed cells. ¹⁵N progressive labelling was used to calculate the rate of ribosome synthesis and degradation to track the fate of 62 r‐proteins. The degradation rates and the synthesis rates of most r‐proteins slowed following oxidative stress leading to an ageing population of ribosomes in stressed cells. However, there were exceptions to this trend; r‐protein RPS14C doubled its degradation rate in both oxidative treatments. Overall, we show that ribosome abundance decreases and their age increases with oxidative stress in line with loss of cell growth rate and total cellular protein amount, but ribosome function of the ageing ribosomes appeared to be maintained concomittently with differences in the turnover rate and abundance of specific ribosomal proteins. Data are available via ProteomeXchange with identifier PXD012840.     

Water Stress and Protein Synthesis: II. Interaction between Water Stress, Hydrostatic Pressure, and Abscisic Acid on the Pattern of Protein Synthesis in Avena Coleoptiles

Water stress causes a reduction in hydrostatic pressure and can cause an increase in abscisic acid in plant tissues. To assess the possible role of abscisic acid and hydrostatic pressure in water stress effects, we have compared the effects of water stress, abscisic acid, and an imposed hydrostatic pressure on the rate and pattern of protein synthesis in Avena coleoptiles. Water stress reduces the rate and changes the pattern of protein synthesis as judged by a double labeling ratio technique, Abscisic acid reduces the rate but does not alter the pattern of protein synthesis. Gibberellic acid reverses the abscisic acid-induced but not the stress-induced inhibition of protein synthesis. The effect of hydrostatic pressure depends on the gas used. With a 19: 1 N₂-air mixture, the rate of protein synthesis is increased in stressed but not in turgid tissues. An imposed hydrostatic pressure alters the pattern of synthesis in stressed tissues, but does not restore the pattern to that found in turgid tissues. Because of the differences in response, we conclude that water stress does not affect protein synthesis via abscisic acid or reduced hydrostatic pressure.     

The effect of cycloheximide upon polyribosome stability in two yeast mutants defective respectively in the initiation of polypeptide chains and in messenger RNA synthesis

Summary The effect of cycloheximide upon protein synthesis, RNA metabolism, and polyribosome stability was investigated in the parent and in two temperature-sensitive mutant yeast strains defective respectively in the initiation of polypeptide chains and in messenger RNA synthesis. Cycloheximide at high concentrations (100 g/ml) severely inhibits but does not completely stop protein synthesis (Fig. 1); the incorporation of ¹⁴C-amino acids into polyribosome-associated nascent polypeptide chains continues at a slow but measurable rate (Figs. 2 and 3). Polyribosome structures are stable in the parent strain at 36° whether or not cycloheximide is present (Fig. 5). However, in Mutant ts^- 136, a mutant defective in messenger as well as in stable RNA production, polyribosomes decay at the restrictive temperature (36° C) at the same rate whether or not cycloheximide is present (Fig. 5). Thus the maintenance of polyribosome structures is dependent upon the continued synthesis of messenger RNA even under conditions of extremely slow polypeptide chain elongation. In mutant ts^- 187, a mutant defective in the initiation of polypeptide chains, all of the polyribosomes decay to monoribosomes within 2 minutes after a shift to the restrictive temperature; cycloheximide completely prevents this decay demonstrating that this mutant is capable of continued messenger RNA synthesis at 36° C. Consistent with these observations is the fact that a newly synthesized heterogeneously sedimenting RNA fraction continues to enter polyribosomes in the presence of cycloheximide whereas the entrance of newly synthesized ribosomal RNA is severely inhibited (Figs. 7, 8, 9). The decay or lack of decay of polyribosomes at the restrictive temperature is, therefore, a rapid and discriminating test for the analysis of mutants defective in macromolecule synthesis. Mutants which exhibit a decay of polyribosomes in the presence of cycloheximide are likely to be defective directly or indirectly in the synthesis of messenger RNA whereas mutants in which decay is prevented or slowed by cycloheximide are likely to be defective in some factor required for the association of ribosomes and messenger RNA.     

Binding of spectinomycin by ribosomes fromEscherichia coli

The binding of the aminocyclitol antibiotic spectinomycin to 70S ribosomes and to 30S subunits fromEscherichia coli has been investigated. The association was influenced by the presence of messenger RNA. The K_d for [³H]-4 OH-spectinomycin binding to 70S ribosomes was 2×10^–7 M without mRNA (polyinosinic acid), and 1×10^–6 M with polyinosinic acid. Dissociation of the antibiotic from the ribosomes was significantly affected by the presence of a bound messenger RNA, which reduced the rate of dissociation by a factor of 5.7. The presence of mRNA did not influence the association of spectinomycin with the 30S subunit. The dissociation rate from the small subunit was comparable to the rate of dissociation from the 70S ribosome and was not affected by the presence of mRNA.     

Protein synthesis by 80S ribosomes during plant development

Protein synthesis by ribosomes from the meristematic region of pea roots (0–0·3 cm) and 2-day-old corn shoots (young tissues) relative to ribosomes from matured regions of pea roots (2·0–2·5 cm) and 10-day-old corn leaves (aged tissues) was compared in the poly U-phenylalanine system. With normal polyribosome preparations, ribosomes from young tissues required approx. 16 mM Mg²⁺ while ribosomes from aged tissues required 20–22 mM Mg²⁺ for optimal activity. With monomeric ribosome preparations induced by anaerobic treatment of the seedlings, the Mg²⁺ optimum was 20–22 mM for ribosomes from both young and aged tissues. A higher level of peptidyl-tRNA in ribosomes from young tissues accounts, at least in part, for the differences in Mg²⁺ optima between ribosomes from young and aged tissues. Monomeric ribosomes were used for assaying the activity of ribosomes per se. Ribosomes from young pea root tips and ribosomes from 2-day-old corn shoots were 25–30% and 100–150% more active, respectively, than the corresponding ribosomes from aged tissues. Differences in ribosomal proteins revealed by gel electrophoresis correlated with the change in ribosomal activity. Reduced activity in the aged ribosomes was not due to RNase activity or inhibitors.     

Cytokinin-Mediated Translational Control of Protein Synthesis in Cultured Cells of Glycine max

Polyribosome formation was stimulated by cytokinin treatmentof cultured cells of Glycine max cv. Funk Delicious. When suspensioncultures were given 0·5 µM zeatin after 24 h inculture in medium lacking a cytokinin, a nearly 2-fold increasein the polyribosome/monoribosome ratio occurred over the subsequent3 h. The effect of actinomycin D and of 5-fluorouridine on RNAsynthesis and on the polyribosome/monoribosome ratios of thesecells was examined. Actinomycin D at 5 and 20 µg/ml^–1inhibitedtotal RNA synthesis by 39 and 60%, respectively, as measuredby [³H]uridine incorporation into acid-precipitable material.The degree of inhibition of precursor incorporation into polyribosomalRNA was similar. At 0·1 mM, 5-fluorouridine inhibited[³H]uridine incorporation by 76%, and [³H]guanosine incorporationby 66% into polyribosomal RNA after 3 h of treatment. Fractionationof the polyribosomal RNA by oligo(dT)-cellulose chromatographydemonstrated that low concentrations of both actinomycin D (5µg ml^–1) and 5-fluorouridine (0·1 mM) inhibitedthe synthesis of ribosomal RNA to a greater extent than thepoly(A)-containing fraction of the messenger RNA. Synthesisof the poly(A)-containing RNA was inhibited by 24% with 5µgml^–1 actinomycin D and by 30% with 0·1 mM 5-fluorouridine.At the above concentrations, these two inhibitors reduced thepolyribosome/monoribosome ratio of the cytokinin-deprived cellsover a 3 h period, but they did not prevent cytokinin-inducedpolyribosome formation. These results provide further evidencethat cytokinin regulates polyribosome levels through an effecton protein synthesis at the translational level     

Gibberellic Acid-enhanced Release of beta-1,3-Glucanase from Barley Aleurone Cells

A β-1, 3-glucanase of barley (Hordeum vulgare) aleurone cells accumulates when half-seeds are imbibed on water, and accumulation continues when the aleurone layers are incubated in buffer solution. The release of the enzyme is a gibberellic acid-dependent process, however. Although gibberellic acid stimulates glucanase release, it does not markedly affect the total amount of glucanase obtained from these cells when compared with water controls. β-1, 3-Glucanase release from aleurone cells is a function of gibberellic acid concentration and commences after a 4-hour lag period. Processes occurring during this lag period are also dependent upon gibberellic acid concentration. Removal of gibberellic acid from the incubation medium at the end of the lag period, however, does not affect subsequent release of glucanase. The release of glucanase from aleurone cells is an active process with a Q₁₀ greater than 3. Inhibitors of respiration and protein and RNA synthesis effectively inhibit the formation and release of glucanase. It is concluded that gibberellic acid functions primarily to enhance glucanase release rather than its formation.     

Changes in polyribosome populations during follicle cell maturation in the ovary of the bug, Oncopeltus fasciatus.

An examination of polyribosome profiles has revealed a distinction between the messenger RNA populations present during the delta, gamma-beta, and alpha₁ stages of the follicular epithelium. The delta and gamma-beta stages are both characterized by polysome profiles with no clearly prominent peaks. There is an increase both in the overall activity of the polysome region and in the amount of heavier polysomes in relation to the lighter ones during the transition from the delta to the gamma-beta condition. The alpha₁ oöcytes possess polysome profiles which are characterized by several extremely prominent peaks and by a reduction in the amount of single ribosomes in the cell. This is taken to indicate that the alpha₁ cells are engaged in the synthesis of large quantities of several different proteins, possibly the chorion precursors.     

ROLE OF RIBONUCLEASE ACTION IN PHENYLALANINE-INDUCED DISAGGREGATION OF RAT CEREBRAL POLYRIBOSOMES

—l -phenylalanine (1 mg/g body wt) or physiological saline (0.9% NaCl) was given intraperitoneally to infant (7-day old), immature (14-day old), and adult (42-day old) rats. The state of ribosomal aggregation was determined in the cerebral postmitochondrial supernatant and purified polyribosome fractions prepared in the presence of rat liver ribonuclease inhibitor. Polyribosomes isolated from cerebral cortices of infant and immature rats 30 or 60 min after administration of phenylalanine were partially disaggregated, whereas the state of aggregation of polyribosomes from mature cerebrum was unchanged. In contrast, little or no evidence of phenylalanine-induced polyribosome disruption was noted in the postmitochondrial supernatant fractions, from which the cerebral polyribosomes were prepared, in any of the animals. Omission of the ribonuclease inhibitor resulted in polyribosome disaggregation in the postmitochondrial supernatant fractions prepared from saline-treated as well as phenylalanine-treated infant rats, but the disruption was more profound in the latter group. Ribonuclease activities in cerebral postmitochondrial supernatant preparations from infant and immature rats were higher than the corresponding values in preparations from adult animals. In addition, the administration of phenylalanine resulted in increases in ribonuclease activities in cerebral postmitochondrial supernatant preparations from the younger animals, but had no effect on these activities in adult animals. These results suggest that alterations in structure and function of polyribosomes from the infant rat cerebrum following a loading dose of phenylalanine were related to exposure of the polyribosomes during isolation to elevated activities of cerebral ribonucleases resulting from this treatment. This hypothesis was supported by the finding that phenylalanine treatment had no effect on the incorporation in vivo of intracisternally-administered radioactive lysine into total, soluble or ribosomal protein of infant cerebrum. However, when cerebral ribosomal RNA was differentially labelled in phenylalanine-treated and saline-treated infant rats by the intracisternal administration of [³H] or [¹⁴C]uridine, and polyribosome fractions were then prepared from the pooled cerebral cortices of both groups, radioactive ribosomes derived from saline-treated rats were more highly aggregated than those derived from phenylalanine-treated animals. It is concluded that gross alterations in cerebral polyribosome structure and function do not occur in vivo in young rats given a large amount of phenylalanine intraperitoneally. However, this treatment, in addition to increasing ribonuclease activity in cerebral cell-free preparations, also sensitizes cerebral polyribosomes to subsequent breakdown upon exposure to ribonucleases during isolation.