Principles of Biochemistry, Volume 3 - A. Lehninger 1985

Molecular Mechanisms of Genetic Information Transfer
DNA Replication and Transcription
In RNA-containing viruses, DNA is transcribed using reverse transcriptase

Some oncogenic RNA animal Viruses, such as Rous Sarcoma virus, possess a unique enzyme—an RNA-dependent DNA polymerase, commonly known as Reverse Transcriptase. Upon infecting a host Cell, this enzyme can catalyze the synthesis of DNA complementary to the viral RNA, which serves as a template. This process generates DNA containing Cancer-causing genes, which is frequently integrated into The Genome of the eukaryotic host cell, where it may remain dormant—that is, unexpressed—for many generations (Fig. 28-24). Under specific conditions, such latent viral genes can be activated to drive viral Replication, while under others, they may trigger the malignant transformation of The Cell.

The existence of reverse transcriptases in RNA tumor viruses was predicted as early as 1962 by Howard Temin of the University of Wisconsin, and their presence was ultimately demonstrated in 1970 by Temin independently alongside David Baltimore of the Massachusetts Institute of Technology. Their discovery attracted widespread attention primarily because it provided definitive proof of Genetic information flowing in the reverse direction, from RNA to DNA. This breakthrough clarified how oncogenes carried by RNA viruses manage to integrate into the host cell genome. It also necessitated a revision of the Central dogma of molecular biology (Fig. 28-25). RNA viruses that encode reverse transcriptase are also referred to as Retroviruses (“retro” being Latin for “backward”).

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Fig. 28-24. The Role of reverse transcriptase in the synthesis of complementary DNA on a single-stranded viral RNA template within an animal cell. The resulting cDNA can integrate into the host cell genome.

Fig. 28-25. Extended interpretation of the central dogma of Molecular Genetics, incorporating the flow of genetic information from RNA to DNA following the discovery of reverse transcriptases.

The discovery of reverse transcriptase resolved a long-standing question: how genetic information from oncogenic RNA viruses can become incorporated into host cell DNA. Accumulating evidence now indicates that the DNA of many animal species contains genes derived from RNA viruses, even in animals that have never been exposed to such viruses. These observations suggest that The genes of certain RNA viruses were transcribed into DNA and integrated into the Chromosomes of these animals' ancestors, perhaps during the Cytology/cytology/16.html">Early stages of their biological evolution. Subsequently, they were passed down through generations via the replication of the total cellular DNA, including the viral-origin oncogenes embedded within it. Indeed, according to one theory of cancer origin, we all harbor “silent,” unexpressed oncogenes in our chromosomes that entered our ancestors' genomes via RNA viruses thousands or millions of years ago. This theory further proposes that such oncogenes are normally quiescent, but when activated—for instance, by exposure to carcinogenic agents—they are transcribed and translated to produce factors that drive the transformation of normal human Cells into malignant ones.

Viral reverse transcriptases, much like all DNA and RNA polymerases, contain Zn²⁺ ions. While they exhibit peak catalytic activity when using their own viral RNA as a template, they are also capable of synthesizing DNA complementary to A wide variety of other RNAs. Reverse transcriptases require a primer, direct the Synthesis of the new DNA chain in the 5'→3' direction, and closely resemble DNA polymerases in many other respects.

As we will see later, reverse transcriptase has become an invaluable biochemical tool for investigating DNA-RNA relationships and for DNA Cloning, much like Restriction Endonucleases (Section 27.24). This enzyme made it possible to artificially synthesize DNA complementary to any RNA template, whether mRNA, tRNA, or rRNA. Synthetic DNA produced in this manner is called complementary DNA (cDNA). For instance, reverse transcriptase can be used to generate a synthetic Gene (i.e., cDNA) encoding one of the Hemoglobin polypeptide chains, starting from its mRNA. mRNAs encoding hemoglobin chains are readily isolated from erythrocytes. In this and many other cases where isolating a natural eukaryotic gene for a specific polypeptide is difficult, yet its mRNA is accessible, a synthetic gene can be produced from that mRNA using reverse transcriptase. Later, we will explore how cDNAs are utilized in recombinant DNA cloning (Chapter 30).