Ty1 and delta elements occur adjacent to several tRNA genes in yeast

A comparative analysis of a number of yeast DNA-pBR322 recombinant plasmids carrying repetitive sequence elements has revealed that Ty1 or delta elements occur in the vicinity of several tRNA genes. Four examples have been characterized in detail: three glutamate tRNA genes and a serine tRNA gene. The tRNAGlu3 genes occupy different chromosomal locations; two of these genes are found adjacent to Ty1 elements, and the third is found adjacent to an independent delta element. A delta unit is also found adjacent to a tRNASer2 gene. Next to one of the tRNAGlu3 genes, the delta element is joined to a truncated sigma element. Junctions between different delta units were characterized by the sequence analysis of two DNA segments that carry no tRNA genes.     

tRNA genes and retroelements in the yeast genome.

A survey of tRNA genes and retroelements (Ty) in the genome of the yeast Saccharomyces cerevisiae is presented. Aspects of genomic organization and evolution of these genetic entities and their interplay are discussed. Attention is also given to the relationship between tRNA gene multiplicity and codon selection in yeast and the role of Ty elements.     

The histidine tRNA genes of yeast

Yeast has at least seven nuclear histidine tRNA genes although there is a single tRNAHis. We have sequenced three of the histidine tRNA genes. The genes have identical coding sequences and the DNA anti-codon sequence GTG corresponds to the GUG anti-codon in tRNAHis. None of the three yeast histidine tRNA genes has an intervening sequence. Two of the three genes contain repeated DNA elements in the region adjacent to the 5' end of the histidine tRNA gene. One of the elements, sigma, is 18 base pairs (bp) from the 5' end of each of these genes, sigma elements are highly conserved and flanked by 5-bp repeats. The other element, delta, is at variable distances from the tRNA gene; one is 439 bp from a histidine tRNA gene and the other is 52 bp from a histidine tRNA gene. These solo delta elements are quite divergent when compared with delta s associated with transposon yeast elements and are not flanked by 5-bp repeats.     

Nucleotide sequences of yeast genes for tRNA(2), tRNA(2) and tRNA(1): homology blocks occur in the vicinity of different tRNA genes

Three members of a collection of pBR322-yeast DNA recombinant plasmids containing yeast tRNA genes have been analyzed and sequenced. Each plasmid carries a single tRNA gene: pY44, tRNA^Ser₂; pY41, tRNA^Arg₂; pY7, tRNA^Val₁. All three genes are intronless and terminate in a cluster of Ts in the non-coding strand. The sequence information here and previously determined sequences allow an extensive comparison of the regions flanking several yeast tRNA genes. This analysis has revealed novel features in tRNA gene arrangement. Blocks of homology in the flanking regions were found between the tRNA genes of an isoacceptor family but, more interestingly, also between genes coding for tRNAs of different amino-acid specificities. Particularly, three examples are discussed in which sequence elements in the neighborhood of different tRNA genes have been conserved to a high degree and over long distances.     

Structural comparison of two yeast tRNA Glu 3 genes.

DNA sequences in a 1.7 kb Pst fragment from yeast have been determined. This fragment is part of a yeast 7.4 kb Hind III segment cloned ino pBR322 (pY 5). The fragment carries a single gene for a glutamate tRNA. The coding portion of this gene is identical in sequence to that of the tRNA Glu 3 gene from pY 20 [1]. The flanking regions differ in their sequences, but possible secondary structures within the 5'-flanking regions bear similar features. Sequence homologies between pY 5 and pY 20 were detected far outside the tRNA genes. More surprisingly, extended sequence homologies were seen between the flanking regions of the pY 20 tRNA Glu 3 gene and a tRNA Ser gene [2,3]. We have also checked the known tRNA genes for structural similarities. Hybridization studies indicate that portions of the Pst fragment are repeated within the yeast genome.     

The methionine initiator tRNA genes of yeast

We have isolated three distinct tRNAimet genes from a yeast DNA clone bank. The complete sequence of two shows that these genes are colinear with the mature tRNAimet and supports the RNA sequence of tRNAimet. Southern analysis of yeast genomic DNA indicates the presence of four copies of tRNAimet gene per haploid genome.     

Transcription and processing of intervening sequences in yeast tRNA genes.

Genes for yeast tRNATyr and tRNAPhe have been sequenced (Goodman, Olson and Hall, 1977; Valenzuela et al., 1978) which contain additional nucleotides (intervening sequences) within the middle of the gene that are not present in the mature tRNA. We have isolated precursors to rRNATyr and tRNAPhe from a yeast temperature-sensitive mutant (at the rna1 locus) which accumulates only certain precursor tRNAs at the nonpermissive temperature. The tRNATyr and tRNAPhe precursors were analyzed by oligonucleotide mapping; they each contain the intervening sequence and fully matured 5' and 3' termini. Furthermore, these precursors were used as substrates to search for an enzymatic activity which can remove the intervening sequences and religate the ends. We have shown that wild-type yeast contains such an activity, and that this activity specifically removes the intervening sequences to produce mature-sized RNAs.     

Sequence analysis of two yeast mitochondrial DNA fragments containing the genes for tRNA Ser UCR and tRNA Phe UUY.

Two restriction enzyme fragments containing yeast mitochondrial tRNA genes have been characterized by DNA sequence analysis. One of these fragments is 320 base pairs long and contains a tRNA Ser gene. The corresponding tRNA SER was isolated from yeast mitochondria and its nucleotide sequence also was determined. This mitochondrial tRNA is 90 nucleotides in length, has a G + C content of 38%, and has UGA as the anticodon. A portion of a 680-base-pair DNA fragment containing a tRNA Phe gene was also sequenced. The portion of this gene which codes for the mature tRNA is 75 base pairs in length, has a G + C content of 33%, and contains the anticodon GAA. Neither gene contains an intervening sequence or codes for the 3' CCA terminus. Both are surrounded by regions of more than 90% A + T. The significance of these sequences is discussed.     

Assembly of the mitochondrial membrane system. Sequences of yeast mitochondrial tRNA genes

Two cytoplasmic "petite" (rho-) clones of Saccharomyces cerevisiae have been selected for the retention of the aspartic acid tRNA gene. The two clones, designated DS200/A102 and DS200/A5, have tandemly repeated segments of mitochondrial DNA (mtDNA) with unit lengths of 1,000 and 6,400 base pairs, respectively. The DS200/A102 genome has a single tRNA gene with a 3'-CUG-5' anticodon capable of recognizing the 5'-GAC-3' and 5'-GAU-3' codons for aspartic acid. The mtDNA segment of DS200/A102 has been determined to represent the wild type sequence from 5.3 to 6.8 map units. The genome of DS200/A5 is more complex encompassing the region of wild type mtDNA from 3.5 to 12.7 units. A continuous sequence has been obtained from 3.5 to 8.6 units. In addition to the aspartic acid tRNA, this region codes for the tRNAUGCAla,tRNAUCUArg, tRNAACGArg, tRNAGCUSer,tRNAUCCGly and tRNAUUULys. The DNA sequence of the DS200/A5 genome has allowed us to deduce the secondary structures of the seven tRNAs and to assign precise map positions for their genes. All the tRNAs except tRNA GUCAsp exhibit most of the invariant features of prokaryotic and eukaryotic tRNAs. The aspartic acid tRNA has unusual D and T psi C loops. The structure of this tRNA is similar to the mitochondrial initiator tRNA of Neurospora crassa (Heckman, J.E., Hecker, L.I., Shwartzbach, S.D., Barnett, W.E., Baumstark, B., and RajBhandary, U.L. Cell 13, 83-95).     

Viral sequences are associated with many histocompatibility genes

A C57BL/6By 5.5 kb Pvu II polymorphic restriction fragment which hybridizes with a spleen focus-forming env probe and maps in the H-30 region has been cloned, and a 358 by subfragment subcloned. Hybridization and sequencing studies show that the 358 by fragment is encoded by the region of the pol gene of murine retrovirus which codes for an endonuclease critical for viral integration. Hybridizations of digested murine genomic DNAs with the 358 by probe generate 31 restriction fragment length polymorphisms (RFLPs); 16 of these can be placed near the following 15 minor histocompatibility (H) loci: H-3, H-4, H-7, H-13, H-15, H-16, H-17, H-19, H-22, H-24, H-27, H-30, H-34, H-36, and H-38. We suggest that the proximity of viral sequences to H loci is probably evolutionarily and functionally significant and that the closeness of viral sequences and minor H loci can probably be utilized to facilitate the cloning of minor H genes. During the course of these studies, it has become possible to tentatively assign H-17, H-34, and H-38 to chromosome 12. In addition, it was observed that several H-2 congenic strains retain portions of chromosome 12 from the parental donor strains used in their derivation.     

Some bacterial tRNA genes are transcribed by eukaryotic RNA polymerase III.

E. coli genes for tRNAasp and tRNAtrp are transcribed by eukaryotic RNA polymerase III upon injection into nuclei of Xenopus oocytes. This indicates that some bacterial tRNA genes contain RNA polymerase III promoter sequences. We discuss evolutionary aspects of this finding.     

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.