scholarly journals Unusual pattern of ribonucleic acid components in the ribosome of Crithidia fasciculata, a trypanosomatid protozoan.

1981 ◽  
Vol 1 (4) ◽  
pp. 347-357 ◽  
Author(s):  
M W Gray
Keyword(s):  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Large Subunit ◽  
Ribosomal Subunit ◽  
Ribosomal Rnas ◽  
High Potassium ◽  
Intact Cells ◽  
Crithidia Fasciculata ◽  
Low Magnesium ◽  
Chain Lengths

In a previous study from this laboratory, presumptive ribosomal ribonucleic acid (RNA) species were identified in the total cellular RNA directly extracted from intact cells of the trypanosomatid protozoan Crithidia fasciculata (M. W. Gray, Can. J. Biochem. 57:914-926, 1979). The results suggested that the C. fasciculata ribosome might be unusual in containing three novel, low-molecular-weight ribosomal RNA components, designated e, f, and g (apparent chain lengths 240, 195, and 135 nucleotides, respectively), in addition to analogs of eucaryotic 5S (species h) and 5.8S (species i) ribosomal RNAs. In the present study, all of the presumptive ribosomal RNAs were indeed found to be associated with purified C. fasciculata ribosomes, and their localization was investigated in subunits produced under different conditions of ribosome dissociation. When ribosomes were dissociated in a high-potassium (880 mM K+, 12.5 mM Mg2+) medium, species e to i were all found in the large ribosomal subunit, which also contained an additional, transfer RNA-sized component (species j). However, when subunits were prepared in a low-magnesium (60 mM K+, 0.1 mM Mg2+) medium, two of the novel species (e and g) did not remain with the large subunit, but were released, apparently as free RNAs. Control experiments have eliminated the possibility that the small RNAs are generated by quantitative and highly specific (albeit artifactual) ribonuclease cleavage of large ribosomal RNAs during isolation. In terms of RNA composition and dissociation properties, therefore, the ribosome of C. fasciculata is the most "atypical" eucaryotic ribosome yet described. These observations raise interesting questions about the function and evolutionary origin of C. fasciculata ribosomes and about the organization and expression of ribosomal RNA genes in this organism.

Unusual pattern of ribonucleic acid components in the ribosome of Crithidia fasciculata, a trypanosomatid protozoan

1981 ◽  
Vol 1 (4) ◽  
pp. 347-357
Author(s):  
M W Gray
Keyword(s):  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Large Subunit ◽  
Ribosomal Subunit ◽  
Ribosomal Rnas ◽  
High Potassium ◽  
Intact Cells ◽  
Crithidia Fasciculata ◽  
Low Magnesium ◽  
Chain Lengths

In a previous study from this laboratory, presumptive ribosomal ribonucleic acid (RNA) species were identified in the total cellular RNA directly extracted from intact cells of the trypanosomatid protozoan Crithidia fasciculata (M. W. Gray, Can. J. Biochem. 57:914-926, 1979). The results suggested that the C. fasciculata ribosome might be unusual in containing three novel, low-molecular-weight ribosomal RNA components, designated e, f, and g (apparent chain lengths 240, 195, and 135 nucleotides, respectively), in addition to analogs of eucaryotic 5S (species h) and 5.8S (species i) ribosomal RNAs. In the present study, all of the presumptive ribosomal RNAs were indeed found to be associated with purified C. fasciculata ribosomes, and their localization was investigated in subunits produced under different conditions of ribosome dissociation. When ribosomes were dissociated in a high-potassium (880 mM K+, 12.5 mM Mg2+) medium, species e to i were all found in the large ribosomal subunit, which also contained an additional, transfer RNA-sized component (species j). However, when subunits were prepared in a low-magnesium (60 mM K+, 0.1 mM Mg2+) medium, two of the novel species (e and g) did not remain with the large subunit, but were released, apparently as free RNAs. Control experiments have eliminated the possibility that the small RNAs are generated by quantitative and highly specific (albeit artifactual) ribonuclease cleavage of large ribosomal RNAs during isolation. In terms of RNA composition and dissociation properties, therefore, the ribosome of C. fasciculata is the most "atypical" eucaryotic ribosome yet described. These observations raise interesting questions about the function and evolutionary origin of C. fasciculata ribosomes and about the organization and expression of ribosomal RNA genes in this organism.

scholarly journals Heat-sensitive mutant strain of Neurospora crassa, 4M(t), conditionally defective in 25S ribosomal ribonucleic acid production.

1981 ◽  
Vol 1 (3) ◽  
pp. 199-207 ◽  
Author(s):  
M W Loo ◽  
N S Schricker ◽  
P J Russell
Keyword(s):  
Neurospora Crassa ◽  
Mutant Strain ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Ribosomal Subunit ◽  
Macromolecular Synthesis ◽  
Sensitive Mutant ◽  
Ribosomal Subunits ◽  
Sequence Of Events ◽  
Ribosomal Rna Processing

A heat-sensitive mutant strain of Neurospora crassa, 4M(t), was studied in an attempt to define its molecular lesion. The mutant strain is inhibited in conidial germination and mycelial extension at the nonpermissive temperature (37 degrees C). Macromolecular synthesis studies showed that both ribonucleic acid (RNA) and protein syntheses are inhibited when 4-h cultures are shifted from 20 to 37 degrees C. Density gradient analysis of ribosomal subunits made at 37 degrees C indicated that strain 4M(t) is deficient in the accumulation of 60S ribosomal subunits in that the ratio of 60S/37S subunits was 0.29:1 compared with 1.6:1 for the parental strain. This phenotype was shown to be the result of a slow rate of processing of, and a deficiency in the amount of, the immediate precursor to 25S ribosomal RNA (the large RNA of the 60S subunit) in the sequence of events constituting the production of mature ribosomal RNAs from the primary transcript of the ribosomal deoxyribonucleic acid, the precursor ribosomal RNA molecule. Analysis of polysomes suggested that the heat-sensitive gene product might function in both the assembly and the function of the 60S ribosomal subunit, since there was a smaller proportion of newly made 60S subunits synthesized at 37 degrees C in the polysome region of the gradients than in the monosome-plus-subunit region. The ribosomal RNA processing defect is apparently responsible for the observed defects in germination and macromolecular synthesis at 37 degrees C, but the precise molecular lesion is not known. On the basis of these results, the heat-sensitive mutant allele in the 4M(t) strain is considered to define the rip1 (ribosome production) gene locus.

scholarly journals Multiple spacer sequences in the nuclear large subunit ribosomal RNA gene of Crithidia fasciculata

1987 ◽  
Vol 6 (4) ◽  
pp. 1063-1071 ◽  
Author(s):  
David F. Spencer ◽  
James C. Collings ◽  
Murray N. Schnare ◽  
Michael W. Gray
Keyword(s):  
Ribosomal Rna ◽  
Large Subunit ◽  
Crithidia Fasciculata ◽  
Ribosomal Rna Gene ◽  
Large Subunit Ribosomal Rna

scholarly journals The universally conserved GTPase HflX is an RNA helicase that restores heat-damaged Escherichia coli ribosomes

2018 ◽  
Vol 217 (7) ◽  
pp. 2519-2529 ◽  
Author(s):  
Sandip Dey ◽  
Chiranjit Biswas ◽  
Jayati Sengupta
Keyword(s):  
Escherichia Coli ◽  
Heat Stress ◽  
Ribosomal Rna ◽  
Large Subunit ◽  
Ribosomal Subunit ◽  
Rna Helicase ◽  
E Coli ◽  
Cryo Electron Microscopy ◽  
Guanosine Triphosphatase

The ribosome-associated GTPase HflX acts as an antiassociation factor upon binding to the 50S ribosomal subunit during heat stress in Escherichia coli. Although HflX is recognized as a guanosine triphosphatase, several studies have shown that the N-terminal domain 1 of HflX is capable of hydrolyzing adenosine triphosphate (ATP), but the functional role of its adenosine triphosphatase (ATPase) activity remains unknown. We demonstrate that E. coli HflX possesses ATP-dependent RNA helicase activity and is capable of unwinding large subunit ribosomal RNA. A cryo–electron microscopy structure of the 50S–HflX complex in the presence of nonhydrolyzable analogues of ATP and guanosine triphosphate hints at a mode of action for the RNA helicase and suggests the linker helical domain may have a determinant role in RNA unwinding. Heat stress results in inactivation of the ribosome, and we show that HflX can restore heat-damaged ribosomes and improve cell survival.

Heat-sensitive mutant strain of Neurospora crassa, 4M(t), conditionally defective in 25S ribosomal ribonucleic acid production

1981 ◽  
Vol 1 (3) ◽  
pp. 199-207
Author(s):  
M W Loo ◽  
N S Schricker ◽  
P J Russell
Keyword(s):  
Neurospora Crassa ◽  
Mutant Strain ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Ribosomal Subunit ◽  
Macromolecular Synthesis ◽  
Sensitive Mutant ◽  
Ribosomal Subunits ◽  
Sequence Of Events ◽  
Ribosomal Rna Processing

A heat-sensitive mutant strain of Neurospora crassa, 4M(t), was studied in an attempt to define its molecular lesion. The mutant strain is inhibited in conidial germination and mycelial extension at the nonpermissive temperature (37 degrees C). Macromolecular synthesis studies showed that both ribonucleic acid (RNA) and protein syntheses are inhibited when 4-h cultures are shifted from 20 to 37 degrees C. Density gradient analysis of ribosomal subunits made at 37 degrees C indicated that strain 4M(t) is deficient in the accumulation of 60S ribosomal subunits in that the ratio of 60S/37S subunits was 0.29:1 compared with 1.6:1 for the parental strain. This phenotype was shown to be the result of a slow rate of processing of, and a deficiency in the amount of, the immediate precursor to 25S ribosomal RNA (the large RNA of the 60S subunit) in the sequence of events constituting the production of mature ribosomal RNAs from the primary transcript of the ribosomal deoxyribonucleic acid, the precursor ribosomal RNA molecule. Analysis of polysomes suggested that the heat-sensitive gene product might function in both the assembly and the function of the 60S ribosomal subunit, since there was a smaller proportion of newly made 60S subunits synthesized at 37 degrees C in the polysome region of the gradients than in the monosome-plus-subunit region. The ribosomal RNA processing defect is apparently responsible for the observed defects in germination and macromolecular synthesis at 37 degrees C, but the precise molecular lesion is not known. On the basis of these results, the heat-sensitive mutant allele in the 4M(t) strain is considered to define the rip1 (ribosome production) gene locus.

scholarly journals Identical 3′-terminal octanucleotide sequence in 18S ribosomal ribonucleic acid from different eukaryotes. A proposed role for this sequence in the recognition of terminator codons

1974 ◽  
Vol 141 (3) ◽  
pp. 609-615 ◽  
Author(s):  
John Shine ◽  
Lynn Dalgarno
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Synthesis ◽  
Drosophila Melanogaster ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
18S Rrna ◽  
Base Sequence ◽  
Terminal Sequence ◽  
Ribosomal Ribonucleic Acid ◽  
Rrna Species

The 3′-terminal sequence of 18S ribosomal RNA from Drosophila melanogaster and Saccharomyces cerevisiae was determined by stepwise degradation from the 3′-terminus and labelling with [3H]isoniazid. The sequence G-A-U-C-A-U-U-AOH was found at the 3′-terminus of both 18S rRNA species. Less extensive data for 18S RNA from a number of other eukaryotes are consistent with the same 3′-terminal sequence, and an identical sequence has previously been reported for the 3′-end of rabbit reticulocyte 18S rRNA (Hunt, 1970). These results suggest that the base sequence in this region is strongly conserved and may be identical in all eukaryotes. As the 3′-terminal hexanucleotide is complementary to eukaryotic terminator codons we discuss the possibility that the 3′-end of 18S rRNA may have a direct base-pairing role in the termination of protein synthesis.

scholarly journals Characterization of the nuclear export adaptor protein Nmd3 in association with the 60S ribosomal subunit

2010 ◽  
Vol 189 (7) ◽  
pp. 1079-1086 ◽  
Author(s):  
Jayati Sengupta ◽  
Cyril Bussiere ◽  
Jesper Pallesen ◽  
Matthew West ◽  
Arlen W. Johnson ◽  
...  
Keyword(s):  
Binding Site ◽  
Nuclear Export ◽  
Large Subunit ◽  
Ribosomal Subunit ◽  
Adaptor Protein ◽  
Structural Data ◽  
Release Mechanism ◽  
Nuclear Pore ◽  
Subunit Joining

The nucleocytoplasmic shuttling protein Nmd3 is an adaptor for export of the 60S ribosomal subunit from the nucleus. Nmd3 binds to nascent 60S subunits in the nucleus and recruits the export receptor Crm1 to facilitate passage through the nuclear pore complex. In this study, we present a cryoelectron microscopy (cryo-EM) reconstruction of the 60S subunit in complex with Nmd3 from Saccharomyces cerevisiae. The density corresponding to Nmd3 is directly visible in the cryo-EM map and is attached to the regions around helices 38, 69, and 95 of the 25S ribosomal RNA (rRNA), the helix 95 region being adjacent to the protein Rpl10. We identify the intersubunit side of the large subunit as the binding site for Nmd3. rRNA protection experiments corroborate the structural data. Furthermore, Nmd3 binding to 60S subunits is blocked in 80S ribosomes, which is consistent with the assigned binding site on the subunit joining face. This cryo-EM map is a first step toward a molecular understanding of the functional role and release mechanism of Nmd3.

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