The genes of Saccharomyces cerevisiae coding for the mitochondrial threonine and tryptophan tRNA synthetases and for a putative mitochondrial ribosomal protein have been cloned. These, and the previously cloned gene for a mitochondrial elongation factor, were used to disrupt or partially delete the wild‐type chromosomal copies of the genes in the respiratory‐competent strain W303. In each case, inactivation of a gene whose product is required for mitochondrial protein synthesis causes an instability in mitochondrial DNA. Although intact mitochondrial genomes are rapidly and quantitatively eliminated in the protein synthesis defective strains, specific rho‐ genomes can be maintained stably over many generations. These results indicate that mitochondrial protein synthesis is required for the propagation of wild‐type mitochondrial DNA in yeast.
About 85% of the maize genome consists of highly repetitive sequences that are interspersed by low-copy, gene-coding sequences. The maize community has dealt with this genomic complexity by the construction of an integrated genetic and physical map (iMap), but this resource alone was not sufficient for ensuring the quality of the current sequence build. For this purpose, we constructed a genome-wide, high-resolution optical map of the maize inbred line B73 genome containing >91,000 restriction sites (averaging 1 site/∼23 kb) accrued from mapping genomic DNA molecules. Our optical map comprises 66 contigs, averaging 31.88 Mb in size and spanning 91.5% (2,103.93 Mb/∼2,300 Mb) of the maize genome. A new algorithm was created that considered both optical map and unfinished BAC sequence data for placing 60/66 (2,032.42 Mb) optical map contigs onto the maize iMap. The alignment of optical maps against numerous data sources yielded comprehensive results that proved revealing and productive. For example, gaps were uncovered and characterized within the iMap, the FPC (fingerprinted contigs) map, and the chromosome-wide pseudomolecules. Such alignments also suggested amended placements of FPC contigs on the maize genetic map and proactively guided the assembly of chromosome-wide pseudomolecules, especially within complex genomic regions. Lastly, we think that the full integration of B73 optical maps with the maize iMap would greatly facilitate maize sequence finishing efforts that would make it a valuable reference for comparative studies among cereals, or other maize inbred lines and cultivars.
The intergenic spacer of the mouse ribosomal genes contains repetitive 140-base-pair (bp) elements which we show are enhancers for RNA polymerase I transcription analogous to the 60/81-bp repetitive enhancers (enhancers containing a 60-bp and an 81-bp element) previously characterized from Xenopus laevis. In rodent cell transfection assays, the 140-bp repeats stimulated an adjacent mouse polymerase I promoter when located in cis and competed with it when located in trans. Remarkably, in frog oocyte injection assays, the 140-bp repeats enhanced a frog ribosomal gene promoter as strongly as did the homologous 60/81-bp repeats. Mouse 140-bp repeats also competed against frog promoters in trans. The 140-bp repeats bound UBF, a DNA-binding protein we have purified from mouse extracts that is the mouse homolog of polymerase I transcription factors previously isolated from frogs and humans. The DNA-binding properties of UBF are conserved from the mouse to the frog. The same regulatory elements (terminators, gene and spacer promoters, and enhancers) have now been identified in both a mammalian and an amphibian spacer, and they are found in the same relative order. Therefore, this arrangement of elements probably is consequences.widespread in nature and has important functionalThe genes coding for the large rRNAs of most eucaryotes are organized in a similar fashion. From yeast cells to humans, these genes are arranged in multiple tandem copies with precursor-coding regions separated from each other by intergenic spacers (reviewed in references 42 and 54). Recent work from a number of laboratories has suggested that, at least among the multicellular eucaryotes, there is also a broadly conserved arrangement of transcriptional regulatory elements in the spacer (reviewed in reference 49a). The ribosomal genes of the frog, Xenopus laevis, may be considered a paradigm for this type of organization since all of the known regulatory elements have been identified in this organism.A typical intergenic spacer from an X. laevis ribosomal gene is shown in the top line of Fig. 1, with the spacer from a mouse ribosomal gene shown below for comparison. In X. laevis, the intergenic spacer is bounded on the left by a site for 3'-end formation of the precursor (31) and on the right by the gene promoter that directs initiation of the precursor transcript. Between these points are located one or more spacer promoters (4, 41, 53) (the only other known promoters that are recognized by polymerase I), and downstream of the spacer promoters are repetitive 60-and 81-base-pair (bp) elements (60/81-bp elements) that act as enhancers for polymerase I transcription and are additive in effect (11,30,44,48). Between the enhancers and the gene promoter is a termination site (31, 40). The 60/81-bp enhancers bind a Xenopus transcription factor, xUBF, which also binds to the gene promoter (47) and is the frog homolog of human UBF (hUBF; 2) and rat UBF (rUBF; 48a), factors which also stimulate transcription from the gene promoters of these species. The frog...
Background: Rice feeds much of the world, and possesses the simplest genome analyzed to date within the grass family, making it an economically relevant model system for other cereal crops. Although the rice genome is sequenced, validation and gap closing efforts require purely independent means for accurate finishing of sequence build data.
A processing site has been identified within the 5' external transcribed spacer (ETS) ofXenopus laevis and X. borealis pre-RNAs, and this in vivo processing can be reproduced in vitro. It involves a stable and specific association of the pre-rRNA with factors in the cell extract, including at least four RNA-contacting polypeptides, yielding a distinct complex that sediments at 20S. Processing also requires the U3 small nuclear RNA. This processing, at residue +105 of the 713-nucleotideX. klevis 5' ETS, is highly reminiscent of the initial processing cleavage of mouse pre-rRNA within its 3.5-kb 5' ETS, previously thought to be mammal specific. The frog and mouse processing signals share a short essential sequence motif, and mouse factors can faithfully process the frog pre-rRNA. This conservation suggests that this 5' ETS processing site serves an evolutionarily selective function.The 18S, 5.8S, and 28S RNAs of the ribosome are initially transcribed as a single, large precursor molecule that is then processed to yield the mature species. Major processing sites of metazoan pre-rRNA, originally identified by mapping relatively abundant rRNA processing intermediates in frogs, mice, and humans (8,35,36), were concluded to be at the ends of the mature rRNA regions, with the order of cleavage frequently but not obligatorily progressing 5' to 3' along the pre-rRNA (arrows in Fig. 1) (reviewed in reference 30). The transcribed spacer regions that separate the mature rRNA segments evidently are degraded rapidly following their excision. Although it was initially thought that the 5' external transcribed spacer (ETS) was removed from the pre-rRNA in one step, the first processing cleavage in the maturation of mouse pre-rRNA was then found to be at position +650 within the 5' ETS, -3 kb upstream from the 18S region (arrowhead in Fig. 1) (11,20). The mouse 5' ETS processing region specifically associates with a number of polypeptides, directing the assembly of an -20S complex (14), and processing also requires the U3 small nuclear ribonucleoprotein (snRNP) (15). The 200-nucleotide (nt) segment just downstream from the processing site is -85% conserved in sequence in various mammalian species. Processing at analogous positions (400 to 800 nts beyond the initiation site) also occurs in humans, rats, and Chinese hamsters (13,31,33). The mouse 5' ETS processing signal is in the proximal 120 nts of this conserved region, the first -11 nts being the residues most critical for processing (6, 7).The 5' ETSs of mammals are 3 to 4 kb in length, but the 5' ETSs of most other eukaryotes are much shorter. In various Xenopus species, the 5' ETS is 600 to 750 nts in length; therefore, the upstream processing site of the 18S rRNA region in frogs is the same distance from the 5' end of the transcript as is the 5' ETS processing site in mammals (Fig. 1). This fact raises the question of whether mammalian-type 5' ETS processing involving the formation of a large com-* Corresponding author. (27).In this article, we demonstrate that X. laevis p...
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