After reading this section, you should be able to:
■ Describe in general terms the strategy used by retroviruses to synthesize their nucleic acids and proteins
■ Differentiate a segmented genome from the genome of a retrovirus
■ Describe the life cycle of HIV, noting the specific mechanisms it uses to accomplish each step
■ Defend the statement “all three enzymatic activities of reverse transcriptase are required for dsDNA to be synthesized from RNA”
■ Distinguish DNA-dependent DNA polymerases, DNA-dependent RNA polymerases, RNA-dependent RNA polymerases, and RNAdependent DNA polymerases in terms of their templates, products synthesized, and proofreading activity, as discussed in this and previous sections
Retroviruses have positive-strand RNA genomes. However, their genomes do not function as mRNA (figure ). Instead, retroviruses first convert their ssRNA genomes into dsDNA using a multifunctional enzyme called reverse transcriptase.
The dsDNA then integrates into the host’s DNA, where it can serve as a template for mRNA synthesis and synthesis of the plus-strand RNA genome. The host cell’s DNA-dependent RNA polymerase catalyzes both of these processes.
Retroviruses have a plus-strand RNA genome that is first converted into dsDNA by the enzyme reverse transcriptase. The viral dsDNA integrates into the host chromosome, where it serves as the template for synthesis of viral mRNA and viral genomes. Both are synthesized using the host cell’s DNA-dependent RNA polymerase
Numerous retroviruses have been identified and studied. However, human immunodeficiency virus (HIV), the cause of AIDS (acquired immune deficiency syndrome), is of particular interest. AIDS is now recognized as the greatest pandemic of the second half of the twentieth century. Because of its global importance, we focus exclusively on HIV in this section.
Acquired immune deficiency syndrome (AIDS) (section 38.3) HIV is a member of the genus Lentivirus within the family Retroviridae. In the United States, AIDS is caused primarily by HIV-1. HIV-1 is an enveloped virus. The envelope surrounds an outer shell, which encloses a somewhat cone-shaped core (figure ).
The core contains two copies of the HIV RNA genome and several enzymes, including the enzymes reverse transcriptase and integrase. Thus far 10 virus-specific proteins have been discovered in the HIV virion. After entering the body, the gp120 viral envelope protein binds to host cells that have a surface glycoprotein called CD4.
The virus requires a coreceptor in addition to CD4, and this varies depending on the host cell infected (see figure 38.10). Still being debated is how HIV enters a host cell. Initially it was thought that the HIV envelope fused with the cell’s plasma membrane and the virus released its core into the cytoplasm. However, evidence exists that virions enter by receptor-mediated endocytosis (figure).
Some scientists suggest that virions may enter by either method. Inside the infected cell, the core protein dissociates from the RNA, and the RNA is copied into a single strand of DNA by the reverse transcriptase enzyme. The RNA is next degraded by reverse transcriptase, and the DNA strand is duplicated to form a double-stranded DNA copy of the original RNA genome (figure ).
Reverse transcription is a critical step in the life cycle of HIV, and reverse transcriptase is a remarkable enzyme with multiple activities. It is an RNA-dependent DNA polymerase, a DNAdependent DNA polymerase, and a ribonuclease. This latter function is referred to as RNaseH activity.
Despite its versatility, reverse transcriptase lacks a function observed in other DNA polymerases: proofreading. Thus it makes many errors as it synthesizes DNA. Nonetheless, reverse transcription is an amazing process that involves the use of a host tRNA molecule as primer for initial steps in DNA synthesis (figure).
In the process, a newly synthesized, small negative-strand DNA molecule is transferred from one end of the RNA template to the other to prime the synthesis of the rest of the minus-strand DNA.
Later the full-length minus-strand DNA circularizes to allow completion of the plus-strand DNA. Once dsDNA is formed, a complex of dsDNA (the provirus), integrase enzyme, and other factors (including some host molecules) moves into the nucleus, where the proviral DNA is integrated into the cell’s DNA by integrase.
Once integrated, the provirus can force the cell to synthesize viral mRNA (figure). Several different mRNAs are transcribed using the host cells DNA-dependent RNA polymerase.
Some are full-length mRNAs and others are subgenomic mRNAs. The subgenomic mRNAs are formed by alternative splicing that brings different coding regions together, eliminating any intervening coding regions (rather than introns, as typically occurs during alternative splicing).
Translation of two of the mRNAs yields polyproteins that are cleaved to give rise to numerous proteins. One of the polyproteins is synthesized by ribosomal frameshifting followed by readthrough.
Ribosomal frameshifting is often used when viral genes overlap but are in different reading frames. It occurs at specific sites at which the ribosome pauses and then either continues in the same reading frame or shifts to the alteRNAtive reading frame.
In the case of HIV, the ribosome reaches a stop codon, pauses, and either stops to yield one protein or shifts reading frame and continues translating the mRNA to yield the larger polyprotein. Some of the earliest proteins synthesized are used to regulate cellular activities so that HIV genes are preferentially expressed.
Proteins needed to form HIV virions are made later. Eventually viral proteins and the complete HIV-1 RNA genome are assembled into new nucleocapsids that bud from the infected host cell (figure). After some time, the host cell dies, in part from repeated budding but by other processes as well.
Retrieve, Infer, Apply
■ What is the function of each of the following HIV products: gp120, reverse transcriptase, and integrase?
■ CD4 is found on several different immune system cells. Consult chapter 34. Why is the presence of this molecule on the surface of many immune system cells important to the development of AIDS?
■ What role does alternative splicing play in the life cycle of HIV-1? For what purposed
Retroviruses: Plus-Strand Viruses That Use Reverse Transcriptase in Their Life Cycles
■ Retroviruses replicate their genomes and synthesize mRNA via a dsDNA intermediate. The dsDNA is formed by a multifunctional enzyme called reverse transcriptase (figure).
■ HIV is an enveloped retrovirus with a cone-shaped core that contains two copies of its genome and several enzymes, including reverse transcriptase. Upon infection, its ssRNA genome is converted to dsDNA, which is then integrated into the host genome (figure).
■ HIV uses alternative splicing to generate numerous different mRNA molecules. One encodes two polyproteins. One of the polyproteins arises by a combined ribosomal frameshifting, readthrough mechanism.