Four consensus sequences are conserved with the same linear arrangement in RNA-dependent DNA polymerases encoded by retroid elements and in RNA-dependent RNA polymerases encoded by plus-, minus- and double-strand RNA viruses. One of these motifs corresponds to the YGDD span previously described by Kamer and Argos (1984). These consensus sequences altogether lead to 4 strictly and 18 conservatively maintained amino acids embedded in a large domain of 120 to 210 amino acids. As judged from secondary structure predictions, each of the 4 motifs, which may cooperate to form a well-ordered domain, places one invariant amino acid in or proximal to turn structures that may be crucial for their correct positioning in a catalytic process. We suggest that this domain may constitute a prerequisite 'polymerase module' implicated in template seating and polymerase activity. At the evolutionary level, the sequence similarities, gap distribution and distances between each motif strongly suggest that the ancestral polymerase module was encoded by an individual genetic element which was most closely related to the plus-strand RNA viruses and the non-viral retroposons. This polymerase module gene may have subsequently propagated in the viral kingdom by distinct gene set recombination events leading to the wide viral variety observed today.

Identification of four conserved motifs among the RNA-dependent polymerase encoding elements.

Despite the rapid mutational change that is typical of positive-strand RNA viruses, enzymes mediating the replication and expression of virus genomes contain arrays of conserved sequence motifs. Proteins with such motifs include RNA-dependent RNA polymerase, putative RNA helicase, chymotrypsin-like and papain-like proteases, and methyltransferases. The genes for these proteins form partially conserved modules in large subsets of viruses. A concept of the virus genome as a relatively evolutionarily stable "core" of housekeeping genes accompanied by a much more flexible "shell" consisting mostly of genes coding for virion components and various accessory proteins is discussed. Shuffling of the "shell" genes including genome reorganization and recombination between remote groups of viruses is considered to be one of the major factors of virus evolution. Multiple alignments for the conserved viral proteins were constructed and used to generate the respective phylogenetic trees. Based primarily on the tentative phylogeny for the RNA-dependent RNA polymerase, which is the only universally conserved protein of positive-strand RNA viruses, three large classes of viruses, each consisting of distinct smaller divisions, were delineated. A strong correlation was observed between this grouping and the tentative phylogenies for the other conserved proteins as well as the arrangement of genes encoding these proteins in the virus genome. A comparable correlation with the polymerase phylogeny was not found for genes encoding virion components or for genome expression strategies. It is surmised that several types of arrangement of the "shell" genes as well as basic mechanisms of expression could have evolved independently in different evolutionary lineages. The grouping revealed by phylogenetic analysis may provide the basis for revision of virus classification, and phylogenetic taxonomy of positive-strand RNA viruses is outlined. Some of the phylogenetically derived divisions of positive-strand RNA viruses also include double-stranded RNA viruses, indicating that in certain cases the type of genome nucleic acid may not be a reliable taxonomic criterion for viruses. Hypothetical evolutionary scenarios for positive-strand RNA viruses are proposed. It is hypothesized that all positive-strand RNA viruses and some related double-stranded RNA viruses could have evolved from a common ancestor virus that contained genes for RNA-dependent RNA polymerase, a chymotrypsin-related protease that also functioned as the capsid protein, and possibly an RNA helicase.

Evolution and Taxonomy of Positive-Strand RNA Viruses: Implications of Comparative Analysis of Amino Acid Sequences

Kaposi's sarcoma (KS) is frequently associated with human immunodeficiency virus-1 (HIV-1) infection. Supernatants from HIV-1-infected T cells carrying the CD4 antigen promote the growth of cells derived from KS lesions of AIDS patients (AIDS-KS cells), and the HIV-1 tat gene, introduced into the germ line of mice, induces skin lesions closely resembling KS. Here we report that the tat gene product (Tat) is released from both HIV-1-acutely infected H9 cells and tat-transfected COS-1 cells. These Tat-containing supernatants specifically promote growth of AIDS-KS cells which are inhibited by anti-Tat antibodies; recombinant Tat has the same growth-promoting properties. Therefore a viral regulatory gene product can be released as a biologically active protein and directly act as a growth stimulator. These and previous data indicate that extracellular Tat could be involved in the development or progression, or both, of KS in HIV-1-infected individuals.

Tat protein of HIV-1 stimulates growth of cells derived from Kaposi's sarcoma lesions of AIDS patients

When the human immunodeficiency virus transactivating gene under the control of the viral regulatory region is introduced into the germline of mice, skin lesions are induced that resemble Kaposi's sarcoma seen in AIDS. Our findings indicate that HIV could play a direct part in causing cancer.

The HIV tat gene induces dermal lesions resembling Kaposi's sarcoma in transgenic mice.

Publisher Summary This chapter discusses the evidence that deregulation of c-myc is a critical component of the pathogenesis of Burkitt's lymphoma, and consideration is given to the potential pathogenetic role of Epstein–Barr virus (EBV). The clinical and epidemiological observations pertinent to the pathogenesis of this tumor are also presented. A consideration of the structural changes in c-myc provides strong, though indirect evidence that a major element in the pathogenesis of Burkitt's lymphoma is the deregulation of c-myc. The possibility of the structural changes in c-myc can be considered, if the c-myc allele involved in the translocation is the expressed allele in Burkitt's lymphoma. Evidence for this comes from two sources. First, in tumor cell lines in which there is a truncated c-myc gene that is, one in which the breakpoint on chromosome 8 is within the first intron, the first exon sequences cannot be detected by Northern blot in the messenger RNA. Second, somatic cell hybrid experiments have been performed in which the Burkitt's lymphoma cell lines were fused to mouse plasmacytoma cells and clones which contained either the derivative chromosomes or their normal alleles were isolated. In the case of hybrids in which one of the parents was a cell line bearing an 8;14 translocation four types of clone, each containing only one of the chromosomes 8, 14, derivative 8, or derivative 14, were isolated. Human c-myc expression was detectable only in clones bearing the derivative chromosome 14.

The Pathogenesis of Burkitt's Lymphoma

AIDS (acquired immune deficiency syndrome) and ARC (AIDS-related complex) are associated with a spectrum of lymphoproliferative disorders ranging from lymphadenopathy syndrome (LAS), an apparently benign polyclonal lymphoid hyperplasia, to B cell non-Hodgkin's lymphoma (B-NHL), i.e., malignant, presumably monoclonal B cell proliferations. To gain insight into the process of lymphomagenesis in AIDS and to investigate a possible pathogenetic relationship between LAS and NHL, we investigated the clonality of the B or T lymphoid populations by Ig or T beta gene rearrangement analysis, the presence of rearrangements involving the c-myc oncogene locus, and the presence of human immunodeficiency virus (HIV) sequences in both LAS and B-NHL biopsies. Our data indicate that multiple clonal B cell expansions are present in a significant percentage of LAS (approximately 20%) and B-NHL (60%) biopsies. c-myc rearrangements/translocations are detectable in 9 of our 10 NHLs, but not in any of the LAS cases. However, only one of the B cell clones, identified by Ig gene rearrangements carries a c-myc gene rearrangement, suggesting that only one clone carries the genetic abnormality associated with malignant B cell lymphoma. Furthermore, the frequency of detection of c-myc rearrangements in AIDS-associated NHLs of both Burkitt and non-Burkitt type suggest that the biological alterations present in AIDS favor the development of lymphomas carrying activated c-myc oncogenes. Finally, our data show that HIV DNA sequences are not detectable in LAS nor in NHL B cell clones, suggesting that HIV does not play a direct role in NHL development. Taken together, these observations suggest a model of multistep lymphomagenesis in AIDS in which LAS would represent a predisposing condition to NHL. Immunosuppression and EBV infection present in LAS can favor the expansion of B cell clones, which in turn may increase the probability of occurrence of c-myc rearrangements leading to malignant transformation.

https://rupress.org/jem/article-pdf/164/6/2049/1096548/2049.pdf

Multiple monoclonal B cell expansions and c-myc oncogene rearrangements in acquired immune deficiency syndrome-related lymphoproliferative disorders. Implications for lymphomagenesis

The RNA genome of human hepatitis A virus (HAV) was molecularly cloned. Recombinant DNA clones representing the entire HAV RNA were used to determine the primary structure of the viral genome. The length of the viral genome is 7478 nucleotides. An open reading frame starting at nucleotide 734 and terminating at nucleotide 7415 encodes a polyprotein of Mr 251,940. Comparison of the HAV nucleotide sequence with that of other picornaviruses has failed to reveal detectable areas of homology. However, a computer analysis of the putative amino acid sequence of HAV and poliovirus demonstrated the existence of short areas of homology in virion protein 3 (VP3) and throughout the carboxyl-terminal portion of the polyproteins. In addition, extensive protein structural homologies with poliovirus were detected.

https://www.pnas.org/content/pnas/82/9/2627.full.pdf

Primary structure and gene organization of human hepatitis A virus.

The frequent occurrence of Kaposi's sarcoma (KS) in association with the acquired immune deficiency syndrome (AIDS) could be due to the fact that the etiological agent of this tumor is the same retrovirus causing AIDS, to another oncogenic virus frequently found in AIDS patients, or to the unmasking of the tumorigenic potential of KS cells by immunosuppression We have therefore investigated the presence of DNA sequences homologous to the AIDS retrovirus, cytomegalovirus (CMV), and hepatitis B virus in 13 KS necropsies and biopsies from AIDS patients All KS DNA samples were negative for AIDS retrovirus or hepatitis B DNA sequences Two DNAs from necropsies contained CMV DNA, but the data suggested the presence of replicating CMV DNA due to generalized infection We have also studied cell cultures derived from KS skin biopsies of AIDS patients These cultures had a short lifetime in vitro and expressed some markers of endothelial cells The cells were not tumorigenic in nude mice but contained a number of chromosomal rearrangements which were often monoclonal within the same culture However, these abnormalities were different from culture to culture and even in cultures from the same biopsy The presence of these chromosomal abnormalities seemed to correlate with the cell positivity for endothelial markers Taken together these results indicate that neither the AIDS retrovirus, CMV, or hepatitis B virus is directly responsible for the altered growth of KS cells, that KS may be polyclonal even within the same lesion, and that KS cells have a tendency to karyotypic rearrangements

https://cancerres.aacrjournals.org/content/canres/46/12_Part_1/6333.full.pdf

Presence of Chromosomal Abnormalities and Lack of AIDS Retrovirus DNA Sequences in AIDS-associated Kaposi's Sarcoma

The gene for glycoprotein gB1 of herpes simplex virus type 1 strain Patton was expressed in stable Chinese hamster ovary cell lines. Expression vectors containing the dihydrofolate reductase (dhfr) cDNA plus the complete gB1 gene or a truncated gene lacking the 194 carboxyl-terminal amino acids of gB1 were transfected into CHO DHFR-deficient cells. Radioimmunoprecipitation demonstrated that the complete gB1 protein expressed in CHO cell lines was cell associated, whereas the truncated protein was secreted from the cells due to deletion of the transmembrane and C-terminal domains of gB1. Cells expressing the truncated gB1 protein were subjected to stepwise methotrexate selection, and a cell line was isolated in which the gB1 gene copy number had been amplified 10-fold and the level of expression of gB1 had increased over 60-fold. The truncated gB1 protein was purified from medium conditioned by the amplified cell line. N-terminal amino acid sequence analysis of this purified protein identified the signal peptide cleavage site and predicted the cleavage of a 30-amino-acid signal sequence from the primary protein. The immunogenicity of the truncated gB1 protein was also tested in mice, and high levels of antibody and protection from virus challenge were observed.

Expression of cell-associated and secreted forms of herpes simplex virus type 1 glycoprotein gB in mammalian cells.

Milligram amounts of highly purified hepatitis A virus (HAV) were obtained from persistently infected cell cultures The HAV polypeptides were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose for detection by an enzyme-linked immunotransfer blot procedure The HAV nucleotide-derived amino acid sequence was subjected to computer analysis to identify potential immunogenic regions within the HAV capsid polypeptides Synthetic peptides corresponding to selected regions of each of the larger putative capsid polypeptides were coupled to keyhole limpet hemocyanin and used to immunize rabbits Four of six anti-HAV peptide sera were strongly reactive Antipeptide serum generated against amino acids (aa) 75 through 82 reacted with the 27,000-molecular-weight (MW) polypeptide; serum against aa 279 through 285 reacted with the 29,000-MW HAV polypeptide; and sera against aa 591 through 602 and 606 through 618 reacted with the 33,000-MW HAV polypeptide These reactions enabled the identification of the gene order of the larger HAV P1 region gene products Our data indicate the following molecular weights: HAV VP2 or 1B, 27,000; HAV VP3 or 1C, 29,000; and HAV VP1 or 1D, 33,000

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Papers

Multiple monoclonal B cell expansions and c-myc oncogene rearrangements in acquired immune deficiency syndrome-related lymphoproliferative disorders. Implications for lymphomagenesis

Primary structure and gene organization of human hepatitis A virus.

Presence of Chromosomal Abnormalities and Lack of AIDS Retrovirus DNA Sequences in AIDS-associated Kaposi's Sarcoma

Expression of cell-associated and secreted forms of herpes simplex virus type 1 glycoprotein gB in mammalian cells.

Structure of the hepatitis A virion: peptide mapping of the capsid region.