The nucleotide sequence of the 4.5S and 5S rRNA genes and flanking regions from Spirodela oligorhiza chloroplasts

The base sequence of Spirodela oligorhiza chloroplast DNA coding for 4.5S and 5S ribosomal RNA, the flanking regions and the spacer between these two genes has been determined. We have compared these sequences with the corresponding ones in other higher plants. Besides a high degree of homology, some interesting differences are found.     

A physical map of the ribosomal DNA repeat unit of Aspergillus nidulans

The ribosomal DNA repeat unit of Aspergillus nidulans has been cloned in pBR322 and a restriction map constructed. The genes coding for the 17S, 5.8S and 25S rRNAs are found in blocks separated by a 1.7 kb spacer region, with the 5.8S RNA gene lying between the genes for the two larger RNAs. The total length of the repeat unit is 7.7 kb. The 5S rRNA is not present in the repeat unit.     

5S ribosomal RNA genes of the newt Notophthalmus viridescens.

The genes which code for the 5S ribosomal RNA in the newt, Notophthalmus viridescens have been cloned and analyzed. Two types of repeating unit were detected: a major type consisting of a 120 bp coding region with a 111 bp spacer, and a minor type composed of a coding region, a pseudogene, and a 113 bp spacer. The pseudogene is a 36 bp segment which corresponds to the 3' terminal third of the 5S RNA gene, and is situated immediately 3' to the gene, being separated from it by 2 bp. Two recombinant plasmids were obtained in which the major and minor units were arranged in an interspersed pattern.     

tRNA genes are found between 16S and 23S rRNA genes in Bacillus subtilis.

There are at least nine, and probably ten, ribosomal RNA gene sets in the genome of Bacillus subtilis. Each gene set contains sequences complementary to 16S, 23S and 5S rRNAs. We have determined the nucleotide sequences of two DNA fragments which each contain 165 base pairs of the 16S rRNA gene, 191 base pairs of the 23S rRNA gene, and the spacer region between them. The smaller space region is 164 base pairs in length and the larger one includes an additional 180 base pairs. The extra nucleotides could be transcribed in tRNAIIe and tRNA Ala sequences. Evidence is also presented for the existence of a second spacer region which also contains tRNAIIe and tRNA Ala sequences. No other tRNAs appear to be encoded in the spacer regions between the 16S and 23S rRNA genes. Whereas the nucleotide sequences corresponding to the 16S rRNA, 23S rRNA and the spacer tRNAs are very similar to those of E. coli, the sequences between these structural genes are very different.     

Escherichia coli 4.5S RNA gene function can be complemented by heterologous bacterial RNA genes.

The essential 4.5S RNA gene of Escherichia coli can be complemented by 4.5S RNA-like genes from three other eubacteria, including both gram-positive and gram-negative organisms. Two of the genes encode RNAs similar in size to the E. coli species; the third, from Bacillus subtilis, specifies an RNA more than twice as large. The heterologous genes are expressed efficiently in E. coli, and the product RNAs resemble those produced by cognate cells. We conclude that the heterologous RNAs can replace E. coli 4.5S RNA and that the essential function of 4.5S RNA is evolutionarily conserved. A consensus structure is presented for the functionally related 4.5S RNA homologs.     

Unique arrangement of coding sequences for 5 S, 5.8 S, 18 S and 25 S ribosomal RNA in Saccharomyces cerevisiae as determined by R-loop and hybridization analysis.

The arrangement of the coding sequences for the 5 S, 5.8 S, 18 S and 25 S ribosomal RNA from Saccharomyces cerevisiae was analyzed in λ-yeast hybrids containing repeating units of the ribosomal DNA. After mapping of restriction sites, the positions of the coding sequences were determined by hybridization of purified rRNAs to restriction fragments, by R-loop analysis in the electron microscope, and by electrophoresis of S₁ nuclease-treated rRNA/rDNA hybrids in alkaline agarose gels. The R-loop method was improved with respect to the length calibration of RNA/DNA duplexes and to the spreading conditions resulting in fully extended 18 S and 25 S rRNA R-loops. The qualitative results are: (1) the 5 S rRNA genes, unlike those in higher eukaryotes, alternate with the genes of the precursor for the 5.8 S, 18 S and 25 S rRNA; (2) the coding sequence for 5.8 S rRNA maps, as in higher eukaryotes, between the 18 S and 25 S rRNA coding sequences. The quantitative results are: (1) the tandemly repeating rDNA units have a constant length of 9060 ± 100 nucleotide pairs with one SstI, two HindIII and, dependent on the strain, six or seven EcoRI sites; (2) the 18 S and 25 S rRNA coding regions consist of 1710 ± 80 and 3360 ± 80 nucleotide pairs, respectively; (3) an 18 S rRNA coding region is separated by a 780 ± 70 nucleotide pairs transcribed spacer from a 25 S rRNA coding region. This is then followed by a 3210 ± 100 nucleotide pairs mainly non-transcribed spacer which contains a 5 S rRNA gene.     

Cloning and characterization of 4.5S and 5S RNA genes in tobacco chloroplasts

Tobacco chloroplast 4.5S and 5S RNAs were shown to hybridize with a 0.9 . 10(6) dalton EcoRI fragment of tobacco chloroplast DNA. Recombinant plasmids were constructed from fragments produced by partial digestion of the chloroplast DNA with EcoRI and the pMB9 plasmid as a vector. Five recombinants containing the 4.5S and 5S genes were selected by the colony hybridization technique. One of these plasmids contained also the 16S and 23S RNA genes and was mapped using several restriction endonucleases as well as DNA-RNA hybridization. The order of rRNA genes is 16S-23S-4.5S-5S and the four rRNA genes are coded for by the same DNA strand.     

Composite structure of the chloroplast 23 S ribosomal RNA genes of Chlamydomonas reinhardii: Evolutionary and functional implications

The chloroplast ribosomal unit of Chlamydomonas reinhardii displays two features which are not shared by other chloroplast ribosomal units. These include the presence of an intron in the 23 S ribosomal RNA gene and of two small genes coding for 3 S and 7 S rRNA in the spacer between the 16 S and 23 S rRNA genes (Rochaix & Malnoë, 1978). Sequencing of the 7 S and 3 S rRNAs as well as their genes and neighbouring regions has shown that: (1) the 7 S and 3 S rRNA genes are 282 and 47 base-pairs long, respectively, and are separated by a 23 base-pair A + T-rich spacer. (2) A sequence microheterogeneity exists within the 3 S RNA genes. (3) The sequences of the 7 S and 3 S rRNAs are homologous to the 5′ termini of prokaryotic and other chloroplast 23 S rRNAs, indicating that the C. reinhardii counterparts of 23 S rRNA have a composite structure. (4) The sequences of the 7 S and 3 S rRNAs are related to that of cytoplasmic 5.8 S rRNA, suggesting that these RNAs may perform similar functions in the ribosome. (5) Partial nucleotide sequence complementarity is observed between the 5′ ends of the 7 S and 3 S RNAs on one hand and the 23 S rRNA sequences which flank the ribosomal intron on the other. These data are compatible with the idea that these small rRNAs may play a role in the processing of the 23 S rRNA precursor.     

Oocyte and somatic 5S ribosomal RNA and 5S RNA encoding genes in Xenopus tropicalis.

We have investigated the structure of oocyte and somatic 5S ribosomal RNA and of 5S RNA encoding genes in Xenopus tropicalis. The sequences of the two 5S RNA families differ in four positions, but only one of these substitutions, a C to U transition in position 79 within the internal control region of the corresponding 5S RNA encoding genes, is a distinguishing characteristic of all Xenopus somatic and oocyte 5S RNAs characterized to date, including those from Xenopus laevis and Xenopus borealis. 5S RNA genes in Xenopus tropicalis are organized in clusters of multiple repeats of a 264 base pair unit; the structural and functional organization of the Xenopus tropicalis oocyte 5S gene is similar to the somatic but distinct from the oocyte 5S DNA in Xenopus laevis and Xenopus borealis. A comparative sequence analysis reveals the presence of a strictly conserved pentamer motif AAAGT in the 5'-flanking region of Xenopus 5S genes which we demonstrate in a separate communication to serve as a binding signal for an upstream stimulatory factor.     

Isolation of Euglena gracilis chloroplast 5S ribosomal RNA and mapping the 5S rRNA gene on chloroplast DNA.

Ribosomal RNA (5S) from Euglena gracilis chloroplasts was isolated by preparative electrophoresis, labeled in vitro with 125I, and hybridized to restriction nuclease fragments from chloroplast DNA or cloned chloroplast DNA segments. Euglena chloroplast 5S rRNA is encoded in the chloroplast genome. The coding region of 5S rRNA has been positioned within the 5.6 kilobase pair (kbp) repeat which also codes for 16S and 23S rRNA. There are three 5S rRNA genes on the 130-kbp genome. The order of RNAs within a single repeat is 16S-23S-5S. The organization and size of the Euglena chloroplast ribosomal repeat is very similar to the ribosomal RNA operons of Escherichia coli.     

The 5S ribosomal genes in the Drosophila melanogaster species subgroup. Nucleotide sequence of a 5S unit from Drosophila simulans and Drosophila teissieri

The 5S genes of the eight species of the D. melanogaster subgroup have been mapped. The spacers, in contrast with coding regions, differ markedly between most species. One 5S gene unit has been sequenced for both D. simulans and D. teissieri. The mature 5S RNA region in these two species is identical to the corresponding region of D. melanogaster. Only 5 nucleotide variations occur between the D. melanogaster and D. simulans 5S gene spacers. The spacer in D. teissieri is very different. Only two segments, located one at each side of the coding region, are clearly homologous to corresponding sequences of D. melanogaster and D. simulans.     

Ribosomal Gene Structure, Variation and Inheritance in Maize and Its Ancestors

We have examined the structure of nuclear genes coding for ribosomal RNAs in maize and its wild relatives, the teosintes and Tripsacum. Digestion of the rDNA (genes coding for 18S, 5.8S and 26S RNAs) with 15 restriction endonucleases (with six base pair recognition sites) yields essentially a single map for the approximately 10,000 repeat units within an individual plant or species. Both length and site variation were detected among species and were concentrated in the intergenic spacer region of the rDNA repeat unit. This result is in agreement with patterns of rDNA change observed among wheat and its relatives (Triticeae), and among vertebrate species. Digestion of these nuclear DNAs with BamHI and subsequent hybridization with a 5S RNA gene-specific probe allowed determination of the size of the 5S gene repeat unit in maize, teosintes, and Tripsacum. Groupings in the genus Zea were characterized by distinct repeat unit types five Tripsacum species examined shared a 260 base pair major repeat unit type. Additionally, several other restriction endonuclease cleavage patterns differentiated among the 5S DNAs within the genus Zea. The rDNA and 5S DNA restriction site variation among the species can be interpreted phylogenetically and agrees with biochemical, karyotypic, and morphological evidence that places maize closest to the Mexican teosintes. For both gene arrays, contributions from each parental genome can be detected by restriction enzyme analysis of progeny from crosses between maize and two distantly related teosintes, Zea luxurians or Zea diploperennis, but certain teosinte arrays were underrepresented in some of the hybrids.