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Results for “Ribosome Profiling”

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PubMed1990

Conformational comparisons among homologous bony fish 5S ribosomal RNAs.

The reversible thermal melting of homologous 5S ribosomal RNAs of three bony fishes, rainbow trout (Salmo gairdneri), carp (Cyprinus carpio), and dog salmon (Onchorhynchus keta), were compared in aqueous salts (sodium chloride and/or divalent metal chlorides) by means of differential optical absorption at 260 and 280 nm, and c.d. at 265 nm. The melting parameters of the multiphasic melting process were conserved among the homologous RNAs, and it was shown that they did not depend on the methods of measurement but on concentrations of [Na+] and of [metal2+]. Other physicochemical methods, such as FT-IR absorption, gave the same melting profiles as the u.v. methods. The c.d. spectrum was homologous among the ribosomal RNAs. The melting is explained by changes in local structures in the 'consensus secondary structure model'. Also, stabilization of the secondary and tertiary structures by binding of Mg2+ and Ca2+, and the catalysed hydrolysis of the main chains by various divalent ions are discussed.

Animals↗

PubMed2006

Assessment of biases associated with profiling simple, model communities using terminal-restriction fragment length polymorphism-based analyses.

Community profiles based on terminal-restriction fragment length polymorphism (T-RFLP) analyses of amplified ribosomal RNA genes are used to monitor changes in microbial community structure and are sometimes employed for semi-quantitative estimates of species richness and abundance in environmental samples. To assess the accuracy of T-RFLP community profiles representing the relative abundance of bacteria in a sample, five species of ruminal bacteria were used to construct simple "communities". Template DNA for PCR amplification was generated either by mixing equal quantities of genomic DNA from pure cultures or by mixing equal numbers of cells prior to DNA extraction. Pairwise mixtures of Fibrobacter succinogenes S85 with Ruminococcus albus 8, Ruminococcus flavefaciens FD-1, Butyrivibrio fibrisolvens 49 and Streptococcus bovis JB1 were created and a 5-member community was constructed. With genomic DNA mixes, relative abundance calculations based on T-RFLP patterns did not reflect input ratios. These discrepancies could not be accounted for by differences in genome size and rRNA operon copy number. In cell mixing experiments, easily lysed cells were overrepresented. To determine if a numerical correction factor could be used to compensate for observed discrepancies, we attempted to quantify biases attributed to DNA extraction and PCR amplification. Biases attributable to these factors led to deviations from expected PCR product ratios by 6% to 38%. We found that interactions were so complex that a suitable factor could not be derived. The unsystematic dependence of T-RFLP peak ratios on variability of DNA extraction and PCR amplification prevents accurate quantification of the relative abundance of microorganisms designed to represent simplified natural populations.

Bacteria↗

PubMed2006

Improved biplex quantitative real-time polymerase chain reaction with modified primers for gene expression analysis.

Stabilizing modified bases incorporated in primers allows the reduction of housekeeping gene primer concentration not possible with regular primers without sacrificing amplification efficiency. Low primer concentration allows coamplification of the most abundant housekeeping genes with very rare templates without mutual inhibition. Real-time polymerase chain reaction (PCR) coamplification of 18S ribosomal RNA with several genes of interest was used in this study with MGB Eclipse (Nanogen, San Diego, CA) hybridization probes. The results may be useful for high throughput gene expression studies as they simplify validation experiments.

Animals↗

PubMed1979

Cation-induced regulatory mechanism of GTPase activity dependent on polypeptide initiation factor 2.

Initiation factor IF-2 ribosome dependent GTP hydrolysis (uncoupled GTPase) presents a bell-shaped pH profile which is shifted by changes in ionic strength. At low ionic strength (I = 25 mM) the maximal hydrolytic activity occurs at pH 7.5; when the ionic strength is increased the pH optimum of the reaction is shifted toward more acidic values. Such behavior can be satisfactorily explained as the effect of an electrostatic potential developed by a neighboring polyanion, presumably RNA, on the catalytic site. The addition of fMet-tRNAfMet or AcPhe-tRNAPhe and messenger RNA (coupled GTPase) changes the ionic strength--pH characteristics of the reaction. Thus there is an effect, direct or indirect, of components located at the ribosomal P site. Investigation of the effect of neighboring polyanions on the catalytic activity of the factor-dependent ribosomal GTPases can be seen to provide information about their functional significance that is complementary to that gained from direct structural studies.

Escherichia coli↗

PubMed1993

Ultrastructural neuropathology in murine trisomy 16 hippocampal grafts.

Trisomy 21, Down syndrome individuals demonstrate Alzheimer's disease-associated neuropathology at post mortem. The amyloid precursor protein, one of the pathological proteins is encoded for on chromosome 21. Mouse chromosome 16 is syntenic to human chromosome 21 for the region spanning the amyloid gene. We have previously reported the appearance of Alzheimer's disease-associated neuropathological proteins in cortical grafts derived from Trisomy 16 mice at 4-6 months survival. This paper reports the ultrastructural observations of large and multiple deposits of intracellular lipofuscin within these trisomic grafts at 12 months survival along with increased numbers of free ribosomes, extended Golgi apparatus and endoplasmic reticulum, membrane degeneration and abnormal axonal profiles. Similar but less severe neuro-degeneration is occasionally observed within 18-month-old normal, mouse hippocampal tissue and rarely observed in age-matched control grafts.

Aging↗

PubMed2002

Discrimination and taxonomy of geographically diverse strains of nitrile-metabolizing actinomycetes using chemometric and molecular sequencing techniques.

Mycolic acid-containing actinomycetes capable of metabolizing nitriles were recovered from deep-sea sediments and terrestrial soils by enrichment culture on acetonitrile, benzonitrile, succinonitrile or bromoxynil. A total of 43 nitrile-degrading strains were isolated and, together with previously recovered nitrile-degrading rhodococci, were identified by a polyphasic taxonomic approach, which included mycolic acid profiles, pyrolysis mass spectrometry (PyMS), genomic fingerprinting based on sequence variability of the 16S ribosomal RNA gene using polymerase chain reaction-restriction fragment length polymorphism-single-strand conformational polymorphism, and 16S rRNA gene sequence comparison. Isolates phylogenetically related to Rhodococcus erythropolis dominated the culturable microorganisms from most marine and terrestrial samples. These isolates clustered together in a major pyrogroup that showed high congruence with PRS profiles of the 16S rRNA gene. Such high congruence also was obtained for other recovered isolates that were assigned to species of Rhodococcus and Gordonia. Sequencing data validated the results obtained by PRS analysis and enabled phylogenetic relationships to be established. Some of the recovered bacteria probably represent novel microbial species. The fact that nitrile-metabolizing microorganisms were recovered from a wide range of habitat types suggests that nitrile transforming enzymatic activity is geographically widely distributed in nature.

Actinomycetales↗

PubMed1999

Synthetic lethality with conditional dbp6 alleles identifies rsa1p, a nucleoplasmic protein involved in the assembly of 60S ribosomal subunits.

Dbp6p is an essential putative ATP-dependent RNA helicase that is required for 60S-ribosomal-subunit assembly in the yeast Saccharomyces cerevisiae (D. Kressler, J. de la Cruz, M. Rojo, and P. Linder, Mol. Cell. Biol. 18:1855-1865, 1998). To identify factors that are functionally interacting with Dbp6p, we have performed a synthetic lethal screen with conditional dbp6 mutants. Here, we describe the cloning and the phenotypic analysis of the previously uncharacterized open reading frame YPL193W, which we renamed RSA1 (ribosome assembly 1). Rsa1p is not essential for cell viability; however, rsa1 null mutant strains display a slow-growth phenotype, which is exacerbated at elevated temperatures. The rsa1 null allele synthetically enhances the mild growth defect of weak dbp6 alleles and confers synthetic lethality when combined with stronger dbp6 alleles. Polysome profile analysis shows that the absence of Rsa1p results in the accumulation of half-mer polysomes. However, the pool of free 60S ribosomal subunits is only moderately decreased; this is reminiscent of polysome profiles from mutants defective in 60S-to-40S subunit joining. Pulse-chase labeling of pre-rRNA in the rsa1 null mutant strain indicates that formation of the mature 25S rRNA is decreased at the nonpermissive temperature. Interestingly, free 60S ribosomal subunits of a rsa1 null mutant strain that was grown for two generations at 37 degrees C are practically devoid of the 60S-ribosomal-subunit protein Qsr1p/Rpl10p, which is required for joining of 60S and 40S subunits (D. P. Eisinger, F. A. Dick, and B. L. Trumpower, Mol. Cell. Biol. 17:5136-5145, 1997). Moreover, the combination of the Deltarsa1 and qsr1-1 mutations leads to a strong synthetic growth inhibition. Finally, a hemagglutinin epitope-tagged Rsa1p localizes predominantly to the nucleoplasm. Together, these results point towards a function for Rsa1p in a late nucleoplasmic step of 60S-ribosomal-subunit assembly.

Alleles↗

PubMed2004

Epigenetic and gene expression changes related to transgenerational carcinogenesis.

Transgenerational carcinogenesis refers to transmission of cancer risk to the untreated progeny of parents exposed to carcinogens before mating. Accumulated evidence suggests that the mechanism of this process is epigenetic, and might involve hormonal and gene expression changes in offspring. To begin to test this hypothesis, we utilized a mouse model (NIH Swiss) in which exposure of fathers to Cr(III) chloride 2 wk before mating can alter incidence of neoplastic and nonneoplastic changes in offspring tissues. Utilizing a MS-RDA approach, we found that the sperm of these fathers had a significantly higher percentage of undermethylated copies of the 45S ribosomal RNA gene (rRNA); this finding was confirmed by bisulfite sequencing. Because gene methylation is a known mechanism of expression control in germ cells, and ribosomal RNA levels have been linked to cancer, these findings are consistent with the hypothesis. Secondly, we observed that offspring of Cr(III)-treated fathers were significantly heavier than controls, and had higher levels of serum T3. Possible effects of T3 levels on gene expression in the offspring were examined by microarray analysis of cDNAs from liver. A total of 58 genes, including 25 named genes, had expression ratios that correlated significantly with serum T3 ratios at P </= 0.001. Some of these genes have potential roles in growth and/or tumor suppression. These results also support the hypothesis of an epigenetic and/or gene expression-based mechanism for transgenerational carcinogenesis. Published 2004 Wiley-Liss, Inc.

Animals↗

PubMed1975

Resuscitation of the monkey brain after one hour complete ischemia. III. Indications of metabolic recovery.

Adult rhesus monkeys were subjected to complete cerebral ischemia for one hour and subsequent recirculation for up to 24 h. Animals with signs of functional recovery (e.g. spontaneous EEG activity) exhibited a partial replenishment of cellular energy sources (ATP, phosphocreatine) and a progressive normalization of cerebral lactate levels. Glucose and pyruvate concentrations showed a transient increase over control values during the early stages of postischemic recirculation. Monkeys without functional recovery lacked a significant resynthesis of energy-rich compounds; adenine nucleotides continued to decrease and lactate concentrations were higher than in animals subjected to ischemia without recirculation. Cerebral polysome profiles remained unaltered during the ischemic period but in all animals a marked disaggregation of polyribosomes with a concomitant increase in ribosomal subunits occurred after the onset of recirculation. In monkeys with indications of functional recovery these changes were reversible but a normal polysome profile was only observed after 24 h of recirculation. The results obtained indicate a postischemic depression of protein synthesis due to an inhibition of peptide chain initiation. After recirculation of the brain for 3-6 h there was evidence for an induction of enzymes involved in polyamine synthesis (ornithine decarboxylase and S-adenosylmethionine decarboxylase). No changes in the activity of these enzymes were observed at the end of the ischemic period, indicating that during complete cerebral ischemia not only the synthesis but also the catabolism of proteins is inhibited.

Adenosine Diphosphate↗

PubMed2006

Medulloblastoma outcome is adversely associated with overexpression of EEF1D, RPL30, and RPS20 on the long arm of chromosome 8.

BACKGROUND: Medulloblastoma is the most common malignant brain tumor of childhood. Improvements in clinical outcome require a better understanding of the genetic alterations to identify clinically significant biological factors and to stratify patients accordingly. In the present study, we applied cytogenetic characterization to guide the identification of biologically significant genes from gene expression microarray profiles of medulloblastoma. METHODS: We analyzed 71 primary medulloblastomas for chromosomal copy number aberrations (CNAs) using comparative genomic hybridization (CGH). Among 64 tumors that we previously analyzed by gene expression microarrays, 27 were included in our CGH series. We analyzed clinical outcome with respect to CNAs and microarray results. We filtered microarray data using specific CNAs to detect differentially expressed candidate genes associated with survival. RESULTS: The most frequent lesions detected in our series involved chromosome 17; loss of 16q, 10q, or 8p; and gain of 7q or 2p. Recurrent amplifications at 2p23-p24, 2q14, 7q34, and 12p13 were also observed. Gain of 8q is associated with worse overall survival (p = 0.0141), which is not entirely attributable to MYC amplification or overexpression. By applying CGH results to gene expression analysis of medulloblastoma, we identified three 8q-mapped genes that are associated with overall survival in the larger group of 64 patients (p < 0.05): eukaryotic translation elongation factor 1D (EEF1D), ribosomal protein L30 (RPL30), and ribosomal protein S20 (RPS20). CONCLUSION: The complementary use of CGH and expression profiles can facilitate the identification of clinically significant candidate genes involved in medulloblastoma growth. We demonstrate that gain of 8q and expression levels of three 8q-mapped candidate genes (EEF1D, RPL30, RPS20) are associated with adverse outcome in medulloblastoma.

Adolescent↗

PubMed1978

The neurons and the synaptic endings in the primate basilar pontine gray.

Two types of neurons, projection and intrinsic, previously identified in Golgi preparations of the adult monkey (Macaca mulatta) basilar pontine gray (Cooper and Fox, '76) were observed electronmicroscopically in Macaca mulatta and the squirrel monkey Saimiri sciureus. The cell body of the projection neuron measures up to 37 micrometer and its cytoplasm is rich in organelles. The Goli apparatus, ribosomes, and mitochondria are disposed around the nucleus, while rough endoplasmic reticulum though abundant is usually confined to one half of the cell body. The cell body of the intrinsic neuron measures less than 20 micrometer and its cytoplasm displays prominent ribosomes, but a paucity of other organelles. Five types of synaptic profiles have been identified in the neuropil of the basilar pons; one measures up to 5 micrometer and the rest 2 micrometer or less. They are: (1) a large profile (MSV) containing medium size vesicles (500A) and a central core of mitochondria and neurofilaments; (2) a profile (SSV) containing small round vesicles (250-500 A) which is the most abundant and ubiquitous; (3) a profile (F) containing flattened or pleomorphic vesicles; (4) a profile (LSV) containing large oval egg shaped vesicles (750 A); and (5) a pale profile (PP) that contains oval and occasionally pleomorphic vesicles. MSV, SSV, and LSV terminals form asymmetrical contacts and F terminals form symmetrical contacts with both dendritic and vesicle-containing, pale profiles. The vesicle-containing, pale profile is both pre- and post-synaptic and participates in serial synapses. Following unilateral cortical ablations both dark and filamentous degeneration were observed in the ipsilateral basilar pontine gray.

Animals↗

PubMed1977

The fine structure of the interrenal cells of the duck (Anas platyrhynchos) with evidence for possible exocytotic release of steroids.

The duck interrenal cell possesses ultrastructural characteristics common to other steroid-secreting cells. Lipid droplets and mitochondria are abundant and lie principally at the apical end of the cell. Lipid droplets are not membrane-limited. Cisternae of smooth endoplasmic reticulum that are occasionally continuous with the less abundant rough endoplasmic reticulum are a prominent feature of the interrenal cell. Tubular profiles of rough endoplasmic reticulum often lie tangentially to mitochondria and ribosomes are either free, grouped in polyribosomal clusters, or bound to the endoplasmic reticulum. Mitochondria possess tubular cristae in the inner regions of the gland and frequently contain a paracrystalline array of small 10nm (o.d.) tubules and less frequently a hexagonal array of 40 nm trilaminar rings. Other cytoplasmic components include dense bodies, residual bodies, microtubules, microfilaments and specialized single membrane-bound vesicles. Gap junctions, intermediate junctions and interdigitating processes constitute the main intercellular associations. No tight junctions were identified. The single membrane-bound vesicles which are occasionally filled with a low electron-dense, lipid-like material form septate-like "junctions" with the plasma membrane. The septa bridge an intracellular gap of 15-17 nm. The vesicles are usually located near the subendothelial space at the basal and basilateral regions of the cell. Occasionally, vesicles fuse with the plasma membrane. It is suggested that these vesicles represent morphological evidence for the exocytotic release of steroid hormones.

Adrenal Glands↗

PubMed1998

Mutations in the cytosolic DnaJ homologue, YDJ1, delay and compromise the efficient translation of heterologous proteins in yeast.

The Saccharomyces cerevisiae YDJ1 gene encodes a yeast homologue of DnaJ, an Escherichia coli molecular chaperone and regulator of Hsp70 function. We examined the function of Ydj1p in vivo by analyzing the activity and production of firefly luciferase (FFLux) and green fluorescent protein (GFP) after inducible expression in yeast strains containing a wild type or a mutant YDJ1 gene. Although FFLux and GFP mRNA levels were similar in the wild type and mutant strains, the FFLux protein was translated about half as efficiently in the ydj1-151 mutant compared to the wild type strain; the lower FFLux level was not the result of increased FFLux turnover in the mutant. In contrast, GFP translation was significantly delayed in the ydj1-151 mutant compared to the wild type strain. Surprisingly, we observed that FFLux and GFP mRNA bound efficiently to polysomes in the ydj1-151 mutant. Analysis of polysome profiles also revealed a modest increase in the amount of 60S ribosomal subunits in the ydj1-151 strain, consistent with a translation defect in the mutant, although the Ydj1 protein was not found to be associated with polysomes. To determine whether the inducible expression of an endogenous yeast protein was also less efficient in the ydj1-151 strain, we examined the inducible synthesis of the yeast TATA-binding protein (TBP) but observed no translation defect. Statistical analysis of the FFLux, GFP, and TBP encoding genes suggests that Ydj1p facilitates the expression of proteins that are poorly translated because both FFLux and GFP contain an abundance of codons that are rarely used in yeast.

Animals↗

PubMed1972

Requirement for extraction of polyribosomes from barley tissue.

The isolation of barley (Hordeum vulgare L.) polyribosomes, showing minimal degradation effects of endogenous RNase, required a buffer adjusted to pH 8.0 and containing 0.40 m KCl in addition to common extraction components. The extracted polyribosomes were characterized in sucrose gradients by their conversion to monosomes when incubated with pancreatic RNase and by their dependence on adequate amounts of Mg(2+) during extraction and analysis. Factors which contributed to polyribosome stability were evaluated by the relative sedimentation rates of aggregates in sucrose gradients. Tissue extraction at KCl concentrations less than 0.40 m and below pH 8.0 resulted in an appearance of larger amounts of ribosomes in the less dense region of the sucrose gradient after centrifugation. The addition of 10 mm dithiothreitol was partially effective in preventing the loss of higher polymerized states of polyribosomes at KCl concentrations below 0.40 m. Extractions conducted at KCl concentrations greater than 0.40 m and at pH 8.0 reduced the amount of ribosomes obtained from the tissue. The monosome portion of the polyribosomal profile was partially dissociated into subunits when the tissue was extracted in 0.60 m KCl. A similar effect on monosomes was obtained when polyribosomes were incubated with cycloheximide and 0.40 m KCl, a result not observed by use of a combination of 0.10 m KCl and the drug or 0.40 m KCl alone.

Journal Article↗

PubMed2000

Protein phosphorylation cascades associated with methamphetamine-induced glial activation.

Reactive gliosis is the most prominent response to diverse forms of central nervous system (CNS) injury. The signaling events that mediate this characteristic response to neural injury are under intense investigation. Several studies have demonstrated the activation of phosphoproteins within the mitogen-activated protein kinase (MAPK) and Janus kinase (JAK) pathways following neural insult. These signaling pathways may be involved or responsible for the glial response following injury, by virtue of their ability to phosphorylate and dynamically regulate the activity of various transcription factors. This study sought to delineate, in vivo, the relative contribution of MAPK- and JAK-signaling components to reactive gliosis as measured by induction of glial-fibrillary acidic protein (GFAP), following chemical-induced neural damage. At time points (6, 24, and 48 h) following methamphetamine (METH, 10 mg/kg x 4, s.c.) administration, female C57BL/6J mice were sacrificed by focused microwave irradiation, a technique that preserves steady-state phosphorylation. Striatal (target) and nontarget (hippocampus) homogenates were assayed for METH-induced changes in markers of dopamine (DA) neuron integrity as well as differences in the levels of activated phosphoproteins. GFAP upregulation occurred as early as 6 h, reaching a threefold induction 48 h following METH exposure. Neurotoxicant-induced reductions in striatal levels of DA and tyrosine hydroxylase (TH) paralleled the temporal profile of GFAP induction. Blots of striatal homogenates, probed with phosphorylation-state specific antibodies, demonstrated significant changes in activated forms of extracellular-regulated kinase 1/2 (ERK 1/2), c-jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK), MAPK/ERK kinase (MEK1/2), 70-kDa ribosomal S6 kinase (p70 S6), cAMP responsive element binding protein (CREB), and signal transducer and activator of transcription 3 (STAT3). MAPK-related phosphoproteins exhibited an activation profile that peaked at 6 h, remained significantly increased at 24, and fell to baseline levels 48 h following neurotoxicant treatment. The ribosomal S6 kinase was enhanced over 60% for all time points examined. Immunoreactivity profiles for the transcription factors CREB and STAT3 indicated maximal increases in phosphorylation occurring at 24 h, and measuring greater than 2- or 17-fold, respectively. Specific signaling events were found to occur with a time course suggestive of their involvement in the gliotic response. The toxicant-induced activation of these growth-associated signaling cascades suggests that these pathways could be obligatory for the triggering and/or persistence of reactive gliosis and may therefore serve as potential targets for modulation of glial response to neural damage.

Animals↗

PubMedDate not supplied

Ultrastructural effects of DDT on cells grown in vitro.

In vitro administration of DDT at 0.5 ppm, 10 ppm, and 50 ppm (0.5, 10 and 50 micrograms/ml culture medium) to HeLa S, human amnion (HA-FL), canine venereal tumor (CVT), canine kidney (MD-CK) and rat nephroma (RN) cells resulted in alterations of the ultrastructure of cells. Responses to the cytotoxic chemical compound were qualitatively and quantitatively similar among all mammalian cell lines tested. The tested cells exhibited dose-specific responses. Al low dose (0.5 ppm) rough endoplasmic reticulum and free ribosomes increased as did the number and size of smooth endoplasmic reticulum profiles. At higher doses (10 ppm and 50 ppm) there was indication of cellular degradation including mitochondrial disorganization, nuclear heterochromatic condensation and peripheralization, and increases in numbers of lysosomes, autophagic vacuoles and multivesicular bodies. This suggests that low level exposure to cytotoxic elements such as DDT initiates a repair and regeneration response from the cells while degenerative changes at higher exposure levels suggest less success with detoxification and possible irreversible cellular damage.

Animals↗

PubMedDate not supplied

Chronic administration of pentagastrin: effect on gastric mucosal protein synthesis in rats.

Groups of adult rats were injected with either saline or pentagastrin (500 microgramg/kg) for 14 days. The capacity of the gastric mucosal ribosomes to synthesize endogenous and exogenous mRNA-directed protein in a cell-free system was then investigated. Polyribosomal profiles were also analysed on sucrose gradients. The endogenous mRNA-directed incorporation of 14C-labelled amino acids into protein by the gastric mucosal ribosomes from the pentagrastrin-injected rats was found to be considerably greater than that of the control. In the presence of exogenous mRNA (poly-U) the polyribosomes as well as the endogenous mRNA-free ribosomes (run-off ribosomes) from the pentagastrin-treated group showed 86% and 136% increment in [14C]-phenylalanie incorporation as compared to the corresponding control preparation. The ribosomal ribonuclease activity between the groups was found to be the same. The results of the present investigation indicate that an enhancement in the translational capacity of the ribosomes is in part responsible for stimulation of gastric mucosal protein synthesis after chronic administration of pentagastrin.

Animals↗

PubMed1976

The site of ribosome degradation in starved Escherichia coli cells.

An attempt was made, in starved Escherichia coli cells, to locate the site at which the process of ribosome degradation is initiated. Supernatant and rapidly sedimenting pellet fractions from exponentially growing and from carbon-starved cells were prepared, and the ribonucleic acids from these fractions were seperated by polyacrylamide electrophoresis and quantitated. The data indicated that 23 S, 16 S, and 5 S RNA are lost only from the pellet; and also that the low molecular weight RNA degradation products are confined to this fraction. Ribosomes from supernatant and pellet fractions were seperated on sucrose density gradients. The sedimentation profiles obtained indicated that pellet fractions of starved cells contain for the most part 50 S and 30 S subunits, whereas 70 S monosomes were most abundant in the supernatant fraction. In vitro measurements on RNA degradation in supernatant and rapidly sedimenting pellet fractions confirmed the in vivo data on the exclusive degradation of ribosomal RNA in the pellet. Based on these data and on previous observations, we suggest that the endoribonucleolytic attack which triggers ribosome degradation, occurs in free subunits attached to the rapidly sedimenting membrane fraction. Subsequently, the ribosome falls apart, and the small RNA pieces generated remain attached to the pellet fraction until their final degradation by the exonuclease.

Carbon↗