Long terminal repeat of murine retroviral DNAs: sequence analysis, host-proviral junctions, and preintegration site.

The nucleotide sequence of the long terminal repeat (LTR) of three murine retroviral DNAs has been determined. The data indicate that the U5 region (sequences originating from the 5' end of the genome) of various LTRs is more conserved than the U3 region (sequences from the 3' end of the genome). The location and sequence of the control elements such as the 5' cap, "TATA-like" sequences, "CCAAT-box," and presumptive polyadenylic acid addition signal AATAAA in the various LTRs are nearly identical. Some murine retroviral DNAs contain a duplication of sequences within the LTR ranging in size from 58 to 100 base pairs. A variant of molecularly cloned Moloney murine sarcoma virus DNA in which one of the two LTRs integrated into the viral DNA was also analyzed. A 4-base-pair duplication was generated at the site of integration of LTR in the viral DNA. The host-viral junction of two molecularly cloned AKR-murine leukemia virus DNAs (clones 623 and 614) was determined. In the case of AKR-623 DNA, a 3- or 4-base-pair direct repeat of cellular sequences flanking the viral DNA was observed. However, AKR-614 DNA contained a 5-base-pair repeat of cellular sequences. The nucleotide sequence of the preintegration site of AKR-623 DNA revealed that the cellular sequences duplicated during integration are present only once. Finally, a striking homology between the sequences flanking the preintegration site and viral LTRs was observed.     

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.     

Nucleotide sequence of the haptoglobin and haptoglobin-related gene pair. The haptoglobin-related gene contains a retrovirus-like element

Thirty-three kilobase pairs (kb) of human DNA containing the haptoglobin (Hp) and haptoglobin-related (Hpr) gene pair were cloned, and the nucleotide sequence of 21-kb DNA was determined. The two genes are closely linked, with Hpr being 2.2 kb downstream of Hp. Six hundred nucleotides of DNA occur between the two genes that are not found either 5' to the Hp gene or 3' to the Hpr gene. After the duplication event, the first intron of the Hpr gene acquired a 9-kb insert consisting mainly of a retrovirus-like element with a potential primer-binding site homologous to a mouse isoleucine tRNA. The element forms a repeated family in the human genome that I name RTVL-I (retrovirus-like element-isoleucine). In the coding region of the Hpr gene, there are no frameshift or nonsense mutations and its exon-intron splicing sites, 5' flanking and 3' flanking sequences do not show any obvious defects. There are 28 amino acid differences between the decoded amino acid sequences of the Hpr and Hp genes. Sixteen of these differences occur in the hpr beta chain, and all appear to be located on the surface of the molecule in places not thought to be involved in the hemoglobin binding function of haptoglobin. The structure of the Hpr gene suggests that the gene may be expressed and give rise to a functional product.     

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.     

The nucleotide sequence of an untranslated but conserved domain at the 3' end of the avian sarcoma virus genome.

The genomes of numerous avian retroviruses contain at their 3' termini a conserved domain denoted "c". The precise boundaries and function of "c" have been enigmas. In an effort to resolve these issues, we determined the sequence of over 900 nucleotides at the 3' end of the genome of the Schmidt-Ruppin subgroup A strain of avian sarcoma virus (ASV). We obtained the sequence from a suitable fragment of ASV DNA that had cloned into the single-stranded DNA phage M13mp2. Computer-assisted analysis of the sequence revealed the following structural features: i) the length of "c" - 473 nucleotides; ii) the 3' terminal domain of src, ending in an amber codon at the 5'boundary of "c"; iii) terminator codons that preclude continuous translation from "c"; iv) suitably located sequences that may serve as signals for the initiation of viral RNA synthesis and for the processing and/or polyadenylation of viral mRNA; v) a repeated sequence that flanks src and that could facilitate deletion of this gene; vi) repeated sequences within "c"; and vii) unexplained homologies between sequences in "c" and sequences in several other nucleic acids, including the 5' terminal domain of the ASV genome, tRNATrp and its inversion, the complement of tRNATrp and its inversion, and the 18S RNA of eukaryotic ribosomes. We conclude that "c" probably does not encode a protein, but its sequence may nevertheless serve several essential functions in viral replication.     

Structural organization of the bovine thyroglobulin gene and of its 5'-flanking region

The structural organization of the bovine thyroglobulin gene has been investigated by a combination of Southern genomic blotting and direct analysis of cloned gene fragments isolated from a chromosomal DNA library. The entire locus is spread over more than 200,000 base pairs which makes it one of the largest eukaryotic genes studies to date. The coding information is scattered into at least 42 exons, 34 of which have been precisely identified. A different evolutionary origin of the 5' and 3' regions of the gene is supported by the highly different proportion of exonic material they contain (12% and 3%, respectively) and by the existence of sequence homology between the 3' region of thyroglobulin and acetylcholinesterase. Detailed sequence analysis of the 5' region of the gene and its flanking segment demonstrated that a significant homology exists between bovine and human thyroglobulin sequences, except for the presence within the ruminant promoter region of a 220-base-pair sequence belonging to the bovine monomer repeated family.     

A repetitive and apparently transposable DNA sequence in Dictyostelium discoideum associated with developmentally regulated RNAs.

Dictyostelium discoideum AX3 genomic DNA contains about 40 copies of a 4.5 Kb sequence. Each has the same restriction map, suggesting that they are all very similar. The sequences are scattered throughout the genome, and each of the six representative copies we cloned have different flanking sequences. Partial fragments of the 4.5 Kb sequence also are scattered throughout the genome. The DNA sequences flanking the 4.5 Kb repetitive sequences are different in different strains of Dictyostelium suggesting that the 4.5 Kb sequence is a transposable element. Each sub-region of this 4.5 Kb sequence is associated with a large number of mRNAs, all of which are developmentally regulated, but whose function is not known.     

Cloned embryonic DNA sequences flanking the mouse immunoglobulin C gamma 3 and C gamma 1 genes

To investigate the DNA surrounding genes for immunoglobulin heavy chain constant (CH) regions, we have isolated two clones bearing a C gamma 3 gene and two bearing a C gamma 1 gene from a library of mouse embryo DNA fragments. The C gamma 3 clones span 8.6 kilobase pairs (kb) on the 5' side of the gene and 6.7 kb on its 3' side, while the C gamma 1 clones together span 13 kb of 5' flanking sequence and 2.5 kb of 3' flanking sequence. Restriction mapping of the C gamma 3 gene indicates that intervening sequences divide the gene into segments of domain size, as in other CH genes. Hybridization of clone fragments to restriction digests of mouse DNA indicates that both the C gamma 1 and C gamma 3 genes probably occur as single copies in the genome. Moreover, the entire cloned sequences on the 5' side of both genes appear to be unique in the genome, indicating that no large common sequences flank CH genes. Restriction data suggest that the C gamma 3 gene is 37-40 kb 5' to the C gamma 1 gene.     

Conserved sequences in both coding and 5'' flanking regions of mammalian opal suppressor tRNA genes.

The rabbit genome encodes an opal suppressor tRNA gene. The coding region is strictly conserved between the rabbit gene and the corresponding gene in the human genome. The rabbit opal suppressor gene contains the consensus sequence in the 3' internal control region but like the human and chicken genes, the rabbit 5' internal control region contains two additional nucleotides. The 5' flanking sequences of the rabbit and the human opal suppressor genes contain extensive regions of homology. A subset of these homologies is also present 5' to the chicken opal suppressor gene. Both the rabbit and the human genomes also encode a pseudogene. That of the rabbit lacks the 3' half of the coding region. Neither pseudogene has homologous regions to the 5' flanking regions of the genes. The presence of 5' homologies flanking only the transcribed genes and not the pseudogenes suggests that these regions may be regulatory control elements specifically involved in the expression of the eukaryotic opal suppressor gene. Moreover the strict conservation of coding sequences indicates functional importance for the opal suppressor tRNA genes.