The nucleotide sequence of the transforming HpaI-E fragment of adenovirus type 5 DNA

The primary structure of the HpaI-E fragment of adenovirus type 5 (Ad5) DNA has been determined, mainly by the method of Maxam and Gilbert (1977). This fragment comprises the leftmost 4.5% of the Ad5 genome, and has been shown to be the shortest DNA fragment capable of transforming cells. The identification of potential initiation and termination codons in the determined sequence indicates that two small polypeptides consisting of 186, and 81 amino acids, respectively, could be synthesized. Taking into account recent data on RNA splicing, a possibility is considered that this DNA may code also for larger polypeptides.     

The nucleotide sequence of the right-hand terminal SmaI-K fragment of adenovirus type 2 DNA

The primary structure of the SmaI-K fragment of adenovirus type 2 (Ad2) DNA has been determined. This region includes one of the origins of DNA replication (Winnacker, 1978; Sussenbach and Kuijk, 1978). A leader sequence for an early mRNA in region 4 (Berk and Sharp, 1977; 1978) has also been mapped in this region. The comparison of the primary structure of this region in Ad2 DNA with the corresponding region in Ad5 DNA shows a remarkable homology which may be significant in view of the fact that Ad2 and Ad5 DNAs can interchangeably function in the in vitro replication system of Challberg and Kelly (1979).     

The nucleotide sequence of the transforming early region E1 of adenovirus type 5 DNA

The sequence of the leftmost 11.3% of the non-oncogenic human adenovirus type 5 (Ad5) DNA has been determined. This segment contains the entire early region E1 of the Ad5 genome which has been shown to be involved in in vitro transformation of non-permissive rodent cells (Van der Eb et al., 1980). From the DNA sequence, and from the mRNA sequence data obtained by Perricaudet et al, (1979, 1980) for the E1 mRNAs from the closely related adenovirus type 2 (Ad2), it is possible to predict the primary structure of the polypeptides encoded by this region. The function of these proteins in cell transformation is discussed. From the positions of mapped restriction endonuclease sites and termini of RNA segments in the nucleotide sequence the length of the Ad5 DNA is estimated to be 36.6 kb.     

The nucleotide sequence at the termini of adenovirus type 5 DNA.

The sequences of the first 194 base pairs at both termini of adenovirus type 5 (Ad5) DNA have been determined, using the chemical degradation technique developed by Maxam and Gilbert (Proc. Nat. Acad. Sci. USA 74 (1977), pp. 560-564). The nucleotide sequences 1-75 were confirmed by analysis of labeled RNA transcribed from the terminal HhaI fragments in vitro. The sequence data show that Ad5 DNA has a perfect inverted terminal repetition of 103 base pairs long.     

Gene organization of the transforming region of adenovirus type 7 DNA

The sequence of the leftmost 11% of the weakly oncogenic human adenovirus type 7 (Ad7) DNA has been determined. This part of the Ad7 viral genome encompasses early region E1 which has been shown to be involved in the process of cell transformation in vitro (Dijkema et al., 1979). From the nucleotide sequence and determined coordinates of the E1 mRNAs, we are able to predict the primary structure of the polypeptides encoded by the transforming region of Ad7. The organization of the E1 region of Ad7 and of other adenovirus serotypes (Bos et al. 1981) leads to the proposal of a novel mechanism for gene regulation at the translational level in which protein synthesis can initiate at either the first or the second AUG triplet available in mRNA. The differences between the large E1b-specific tumor antigens of adenovirus types 12, 7 and 5 may explain the differences in oncogenicity of these viruses.     

The nucleotide sequence of the transforming HindIII-G fragment of adenovirus type 5 DNA. The region between map positions 4.5 (HpaI site) and 8.0 (HindIII site).

The nucleotide sequence of the region between map positions 4.5 (HpaI-site) and 8.0 (HindIII-site) of adenovirus type 5 (Ad5) DNA has been determined. This stretch of DNA is part of the transforming HindIII-G fragment, which is 2809 nucleotides long. The sequenced segment was found to have a long open reading frame for protein biosynthesis, starting 23 nucleotides from the HpaI site and extending all the way to the HindIII-G site, which could code for a protein of at least 44 000 daltons. The possible correlation beteen the coding capacity of the HindIII-G fragment and the "transforming" proteins specified by it will be discussed in the light of the recent data on the splicing of early mRNAs.     

Unique species of mRNA from adenovirus type 7 early region 1 in cells transformed by adenovirus type 7 DNA fragment.

Adenovirus type 7 (Ad7) early region 1 mRNA species transcribed in rat cell lines transformed by the HindIII-I . J fragment (the left 7.8% of the viral genome) and in human KB cells infected with Ad7 were mapped on the viral genome, using S1 nuclease gel and diazobenzyloxymethyl paper hybridization techniques. At the early stage of productive infection, two mRNA's (950 and 840 nucleotides long) with the common 5' and 3' ends but different internal splicings were mapped from region 1A (map units 1.4 to 4.3), and one mRNA (2,310 nucleotides long, with the internal splicing between map units 9.9 to 10.1) was mapped from region 1B (map units 4.6 to 11.4). At the late stage, these early spliced mRNA's were also found and at least three additional Ad7 mRNA's were identified: 700-nucleotide-long mRNA in region 1A; and 1,100- and nucleotide-long mRNA's in region 1B. In transformed rat cell lines, two early region 1A mRNA's (950 and 840 nucleotides long) were also transcribed. Surprisingly, in addition, several unique Ad7 mRNA's, not found in productivity infected cells, were identified in all of the transformed cell lines. Their molecular sizes and coding sequences varied in individual cell lines. However, these mRNA's had the 5' end-proximal portion in region 1B and the 3' end-proximal portion in region 1A, these portions being transcribed by extending from region 1B to 1A on viral DNA fragments joined in a tandem array in transformed cells.     

The nucleotide sequence of the genes encoded in early region 2b of human adenovirus type 7

The nucleotide sequence of a cloned DNA segment encoding the early region 2b from the group B human adenovirus Ad7 has been determined. When compared to Ad2, a group C adenovirus, these sequences were found to be approx. 80% homologous within the l-strand gene-coding regions. Most changes are transitions or transversions, although several deletions/insertions also occur within the N-terminal domain of one of the coding regions. The substantial nucleotide homology results in a high degree of amino acid conservation in the predicted polypeptides encoded by the early region 2b genes. Two major open reading frames, corresponding to the Mr 87000 and Mr 140000 polypeptides of Ad2, are found in the l strand of Ad7 between genome coordinates 28.5 to 23.1 and 13.8, respectively. The r strand of the DNA in this region encodes the three leader segments joined to the 5' end of the most late viral mRNAs, and also encodes the i-leader segment found between the second and third leaders on some mRNAs. The positions of the donor and acceptor splice sites of the three leaders are conserved and can be identified by homology to Ad2. Only two of the unidentified open reading frames (URF) in Ad2 (Gingeras et al., J. Biol. Chem., in press) can be found in Ad7. URF1, encoding an Mr 13500 polypeptide at genome coordinate 17, is predominantly conserved in nucleotide and amino acid sequence, but contains one half as many arginine amino acids as does URF1 of Ad2. URF2, encoding an Mr 13600 protein which lies within the i-leader region, is not well conserved in either nucleotide or amino acid sequence.     

The nucleotide sequence of the right-hand terminus of adenovirus type 5 DNA: implications for the mechanism of DNA replication.

The nucleotide sequence of the right-hand terminal 3% of adenovirus type 5 (Ad5) DNA has been determined, using the chemical degradation technique developed by Maxam and Gilbert (1977). This region of the genome comprises the 1003 basepair long HindIII-I fragment and the first 75 nucleotides of the adjacent HindIII-F fragment, extending from the right-hand terminus to the sequences from which the main body of the mRNA of early region 4 is transcribed. One of the origins of adenovirus DNA replication is located within this part of the genome. The sequencing results are discussed in relation to several models proposed for the mechanism of replication of linear DNA molecules, which invariably depend on the presence of specific arrangements of nucleotides at the termini of those linear DNAs.     

The nucleotide sequence of the polypeptide IX gene of human adenovirus type 3

The nucleotide sequence of the DNA segment encompassing the polypeptide IX gene of class B human adeno-virus serotype 3 (Ad3) has been determined using cloned restriction fragments. There is only a single, open translational reading frame capable of specifying a protein of 138 amino acids, comparable to the M_r 12000–13000 of protein IX detected in virions (Wadell, 1980). The corresponding region of a closely related class B virus, Ad7, is virtually identical (Dijkema et al., 1981), but the comparable segments of class C viruses Ad2 or Ad5 are much less homologous (Aleström et al., 1980; Maat et al., 1980). There are 150 single bp changes and 19 deletion-insertions, at least one frameshift, together affecting 210 nucleotides within the 455 bp comparison positions of the protein-coding regions of Ad2 (423 bp) and Ad3 (417 bp). Each of the 19 deletion-insertions involves an integral multiple of 3 bp in phase with the open translation frame. There is no “TATA” promoter box in Ad3 DNA at the position comparable to that of Ad2. The deduced protein sequences near the amino-terminus are extensively conserved between the two classes of viruses, but the carboxy-terminal portion and the nucleotide sequences flanking the gene are much more diverged. In both classes, these N- and C-terminal regions of the inferred proteins are linked by an alanine-rich chain, an arrangement suggestive of two functional domains.     

The nucleotide sequence of the inverted terminal repetition of the tree shrew adenovirus DNA

The termini of the tupaia (tree shrew) adenovirus (TAV) DNA have been sequenced. The inverted terminal repetitions (ITR) are 166 bp long containing the A + T-rich, highly conserved sequence present in all adenovirus DNAs so far analysed. An unusual feature within the TAV ITR is the presence of four sets of a conserved sequence TGACCG which occur at or near the ends of many adenovirus ITR.     

Determination of the nucleotide sequence for the penton-base gene of human adenovirus type 5

The major structural proteins of adenovirus (Ad), which form the external capsid, are hexon, penton base and fiber. The primary structure of the Ad5 penton base has been deduced from the nucleotide sequence of the corresponding gene. It has 98.6% homology with the sequence of the analogous protein from Ad2. This result is in contrast with the significantly lower homology found for the two other major structural proteins, the hexon and the fiber.     

The inverted terminal repetition of the DNA of weakly oncogenic adenovirus type 7

By use of the chemical modification technique of Maxam and Gilbert (1977), the first 180 base pairs at both termini of the human adenovirus 7 genome have been determined. The results show that adenovirus 7 DNA contains a perfect inverted terminal repetition of 136 base pairs.     

Polar encapsidation of adenovirus DNA: cloning and DNA sequence of the left end of adenovirus type 3

The left-end adenovirus type 3 DNA sequence is very similar to those of other subgroup B adenoviruses, especially in the area between the HinfI site (320 base pairs) and the early-region Ia gene. This segment of the genome has been implicated as necessary for the left-end polarity of adenovirus DNA encapsidation. This segment and the sequences flanking it are compared with the corresponding sequences of adenovirus type 5 and adenovirus type 12, and the extent and pattern of intersubgroup homologies are discussed.     

Unusual nucleotide sequence of a DNA fragment isolated from nuclear envelopes of mouse hepatocytes

Fragments of chromosomal DNA isolated from nuclear envelopes of mouse hepatocytes were previously cloned and partially characterized in our laboratory. One of the cloned fragments (EnvM4) had unusual characteristics: abundant representability in the clone library (about 50%) and homology with DNA of archebacteria. An analysis of nucleotide sequence of this fragment conducted in more detail allowed the detection of an evolutionary conserved region present in the genomes of various organisms (from bacteria to human). Other characteristics of fragment EnvM4 suggest a possible functional role of chromosomal DNA regions determining the attachment of chromosomes to the nuclear envelope.