The level of genetic variation of human immunodeficiency virus type 1 (HIV-1), a member of the lentivirus genus of the Retroviridae family, is high relative to that of retroviruses in some other genera. The high error rates of purified HIV-1 reverse transcriptase in cell-free systems suggest an explanation for this high genetic variation. To test whether the in vivo rate of mutation during reverse transcription of HIV-1 is as high as predicted by cell-free studies, and therefore higher than that rates of mutation of retroviruses in other genera, we developed an in vivo assay for detecting forward mutations in HIV-1, using the lacZ alpha peptide gene as a reporter for mutations. This system allows the rates and types of mutations that occur during a single cycle of replication to be studied. We found that the forward mutation rate for HIV-1 was 3.4 x 10(-5) mutations per bp per cycle. Base substitution mutations predominated; G-to-A transition mutations were the most common base substitution. The in vivo mutation rates for HIV-1 are three and seven times higher than those previously reported for two other retroviruses, spleen necrosis virus and bovine leukemia virus, respectively. In contrast, our calculated in vivo mutation rate for HIV-1 is about 20-fold lower than the error rate of purified HIV-1 reverse transcriptase, with the same target sequence. This finding indicates that HIV-1 reverse transcription in vivo is not as error prone as predicted from the fidelity of purified reverse transcriptase in cell-free studies. Our data suggest that the fidelity of purified HIV-1 reverse transcriptase may not accurately reflect the level of genetic variation in a natural infection.

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Lower in vivo mutation rate of human immunodeficiency virus type 1 than that predicted from the fidelity of purified reverse transcriptase.

A number of hormones and growth factors have been shown to stimulate target cells via second messenger pathways that in turn regulate the phosphorylation of specific nuclear factors. The second messenger cyclic AMP, for example, regulates a striking number of physiologic processes, including intermediary metabolism, cellular proliferation, and neuronal signaling, by altering basic patterns of gene expression. Our understanding of cyclic AMP signaling in the nucleus has expanded considerably over the past decade, owing in large part to the characterization of cyclic AMP–responsive promoter elements, transcription factors that bind them, and signal-dependent coactivators that mediate target gene induction. More importantly, these studies have revealed new insights into biological problems as diverse as biological clocks and long-term memory. The purpose of this review is to describe the components of the cyclic AMP response unit and to analyze how these components cooperate to induce target gene expression ...

Transcriptional Regulation by Cyclic AMP

Most of the nuclear genome of warm-blooded vertebrates is a mosaic of very long (much greater than 200 kilobases) DNA segments, the isochores; these isochores are fairly homogeneous in base composition and belong to a small number of major classes distinguished by differences in guanine-cytosine (GC) content. The families of DNA molecules derived from such classes can be separated and used to study the genome distribution of any sequence which can be probed. This approach has revealed (i) that the distribution of genes, integrated viral sequences, and interspersed repeats is highly nonuniform in the genome, and (ii) that the base composition and ratio of CpG to GpC in both coding and noncoding sequences, as well as codon usage, mainly depend on the GC content of the isochores harboring the sequences. The compositional compartmentalization of the genome of warm-blooded vertebrates is discussed with respect to its evolutionary origin, its causes, and its effects on chromosome structure and function.

https://science.sciencemag.org/content/sci/228/4702/953.full.pdf

The mosaic genome of warm-blooded vertebrates.

https://www.cell.com/cell/pdf/0092-8674(86)90778-6.pdf

Identification and characterization of conserved and variable regions in the envelope gene of HTLV-III/LAV, the retrovirus of AIDS

Manual of Standards for Diagnostic Tests and Vaccines

Bovine Leukemia Virus: Molecular Biology and Epidemiology

Short-term cultures of bovine leukemic lymphocytes release virus particles with biochemical properties of RNA oncogenic viruses. These particles, tentatively called bovine leukemia virus (BLV), have a high molecular weight RNA-reverse transcriptase complex and a density of 1.155 g/ml in sucrose solutions. Molecular hybridizations between BLV/[3H]cDNA and several viral RNAs show that BLV is not related to Mason-Pfizer monkey virus, simian sarcoma associated virus, feline leukemia virus, or avian myeloblastosis virus. These results were confirmed by hybridization between BLV 70S RNA and [3H]cDNA synthesized in the various viruses tested. The high preference of BLV reverse transciptase for Mg++ as the divalent cation suggests that BLV might be an atypical mammalian leukemogenic "type C" virus. DNA-DNA hybridization studies using BLV [3H]cDNA as a probe strongly suggest that the DNA of bovine leukemic cells contains viral sequences that cannot be detected in normal bovine DNA.

https://www.pnas.org/content/pnas/73/4/1014.full.pdf

Bovine leukemia virus: an exogenous RNA oncogenic virus

Integration of bovine leukemia proviral DNA in the genome of infected cells was investigated in cattle affected by either the persistent lymphocytosis or the lymph node tumor form of enzootic bovine leukosis. In persistent lymphocytosis, proviral DNA was found to be integrated at a large number of genomic sites in one-fourth to one-third of circulating leukocytes. In the lymph node tumor form, in contrast, proviral DNA was found to be integrated at one or very few sites in the genomes of a larger fraction of both circulating leukocytes and lymph node tumor cells.

Genomic integration of bovine leukemia provirus: comparison of persistent lymphocytosis with lymph node tumor form of enzootic

Direct inoculation of a cloned bovine leukemia virus (BLV) provirus into sheep has allowed study of the viral infectivity of genetic mutants in vivo. Three BLV variants cloned from BLV-induced tumors and 12 in vitro-modified proviruses were isolated and analyzed for viral expression in cell culture. The proviruses were then inoculated into sheep in order to assess viral infectivity in vivo. Of three variants cloned from BLV-induced tumors (344, 395, and 1345), one (344) was found infectious in vivo. This particular provirus was used to engineer 12 BLV mutants. A hybrid between the 5' region of the complete but noninfectious provirus 395 and the 3' end of mutant 344 was infectious in vivo, suggesting that the tax/rex sequences were altered in virus 395. As expected, several regions of the BLV genome appeared to be essential for viral infection: the protease, pol, and env genes. Even discrete modifications in the fusion peptide located at the NH2 end of the transmembrane gp30 glycoprotein destroyed the infectious potential. In contrast, mutations and deletions in the X3 region present between the env gene and the 3' tax/rex region did not interfere with viral infection in vivo. This region of unknown function could thus be used to introduce foreign sequences. A BLV recombinant carrying a ribozyme directed against the tax/rex sequences was still infectious in vivo. Cotransfection of two noninfectious mutants carrying deletions led to infection in two of four independent injections, the infectious virus being then a recombinant between the two deletants. The experimental approach described here should help to gain insight into essential mechanisms such as in vivo viral replication, cooperation between deletants for viral infectivity, and viral superinfections. The gene products in the X3 and X4 region which are dispensable for in vivo infection could be involved in leukemogenesis, and thus proviruses deleted in these sequences could constitute the basis for a live attenuated vaccine.

In vivo infection of sheep by bovine leukemia virus mutants.

Genomic integration of bovine leukemia provirus and lack of viral RNA expression in the target cells of cattle with different responses to BLV infection.

Marc Mammerickx

Papers

Bovine Leukemia Virus: Molecular Biology and Epidemiology

Bovine leukemia virus: an exogenous RNA oncogenic virus

Genomic integration of bovine leukemia provirus: comparison of persistent lymphocytosis with lymph node tumor form of enzootic

In vivo infection of sheep by bovine leukemia virus mutants.

Genomic integration of bovine leukemia provirus and lack of viral RNA expression in the target cells of cattle with different responses to BLV infection.