Poxvirus replication

Replication of the viral DNA occurs only after a second stage uncoating and this step requires protein synthesis (Joklik, 1964) because the addition of cycloheximide at this stage of replication strongly inhibits DNA replication (Kates and McAuslan, 1961b). The double-stranded, crosslinked genome is replicated from each end semiconservatively (Esteban and Holowczak, 1977 Esteban et al., 

1977) and is localized within membrane-associated cytoplasmic viral factories (Cairns, 1960). DNA synthesis continues in these factories for a period up to 5 hr postinfection at which time limiting membranes begin to form around areas of condensed filaments of DNA. In the following period and up to 10 hr, this assembly results in the formation of spherical immature particles which then proceed to the slightly smaller, brick-shaped mature particles by means of continued eccentric condensation of the enclosed components (Sarov and Joklik, 1973). The mature particles diffuse away from the factories and are released from the cytoplasm, without lysis, as complex structures containing the undefined lateral bodies and a double outer membrane. 

All of the host cell macromolecular synthetic processes are inhibited by infection with members of the poxvirus group. The rapidity with which these events occur is dependent on the parameters of the infection, most importantly, the multiplicity of infection, the particular strain of virus, and the cell line used. 

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Most poxvirus virions appear to enter cells by pinocytosis, and then to uncoat within cytoplasmic vesicles. Poxvirus replication occurs in these discrete cytoplasmic inclusions, independent of the cell nucleus. Host biosynthetic processes are inhibited soon after poxvirus infection occurs. Virus transcription initiates almost immediately after entry, as DNA transcription is initiated by core enzymes while the genome is still in the core. Early gene products have to do with DNA synthesis, ensuring adequate levels of precursors for DNA synthesis and inhibition of host defense mechanisms.26 This... 

Li+ has significant inhibitory effects upon DNA viruses, in particular HSV which has been studied in depth. It was originally shown that Li+ inhibits viral replication in a dose-dependent, reversible manner in HSV-infected baby hamster kidney cells [240], and this has been found to be due to a Li+-induced decrease in the synthesis of viral DNA [241]. It is now well established that Li+ inhibits DNA synthesis in HSV types 1 and 2 and in several other DNA viruses, including measles, vaccinia, adenovirus, poxvirus, pseudorabies virus, Epstein-Barr virus, and the bovine, equine, and canine HV s [241]. Interestingly, Li+ has no effect on the replication of RNA viruses, such as influenza or encephalomyo-carditis virus. 

The oncolytic viruses include adenovirus, measles, reovirus, vesicular stomatitis virus (VSV),HSV,poxvirus, and vaccinia. Specific examples include (1) ONYX-015, which is an adenoviral oncolytic virus, administered to patients with liver metastases of colorectal cancer and pancreatic cancer [29], (2) Reolysin, which is an oncolytic reovirus administered to patients with glioma [30], and (3) MV-CEA, which is an oncolytic measles virus expressing carcinoembryonic antigen, administered to patients with ovarian cancer [31]. Some oncolytic viruses are wild type and are apparently not pathogenic in humans, such as the Newcastle disease virus (NDV), which is an RNA avian paramyxovirus. PV701, a naturally attenuated, replication-competent strain of NDV, has been administered to patients with advanced solid tumors [32], The applicability of oncolytic viruses as a therapy for clinical oncology trials is due to their potential selectivity the ability to kill tumor cells but not normal cells. However, the level of attenuation of viral replication in normal cells is limited for most oncolytic vectors. 

Idoxuiidlne, USF. Idoxuridinc. S-iodo-2 -dcoxyuridine (Stoxil, Herplex), was introduced in 1963 for the treatment of herpes simplex keratitis. - The drug is an iodinated analogue of thymidine that inhibits replication of a number of DNA viruses in vitro. The susceptible viruses include the herpesviruses and poxviruses (vaccinia). 

Ribavirin inhibits (he replication of a very wide variety of RNA and DNA viruses, including orthomyxoviruses, paramyxoviruses, arenaviruses, bunyavirusc,s, herpesviruses, adenoviruses, poxvirus, vaccinia, influenza virus, parainfluenza virus, and rhinovirus. In spite of the broad spectrum of activity of ribavirin, the drug has been approved for only one therapeutic indication—(he treatment of severe lower respiratory infections caused by RSV in carefully selected hospitalized infants and young children. 

The molluscum contagiosum members of the poxvirus (MCV) family characteristically induce nonin-flamed papular skin tumors in children and AIDS patients (Table 8). One of the MCV products acts as a broad-spectrum antagonist that blocks the response of a variety of leukocytes to both CXC and CC chemokines which may account for the failure of host inflammatory responses to MCV (Damon et al., 1998). This gene product also suppresses replication of hematopoietic progenitor cells (Krathwohl et al., 1997). 

A DNA-dependent RNA transcriptase is found in the core of poxviridae. Unlike all other DNA viruses, the poxvirus multiplies in cell cytoplasm with the other DNA viruses, transcription takes place in the nucleus, where the host cell polymerases are utilized. Thus the majority of DNA viruses need no replicative enzymes of their own. 

Figure 49-1 provides a schematic diagram of the replicative cycle of a DNA virus (A) and an RNA virus B). DNA viruses include poxviruses (smallpox), herpesviruses (chickenpox, shingles, oral and genital herpes), adenoviruses (conjunctivitis, sore throat), hepadnaviruses (hepatitis B virus [HBV]), and papiUomaviruses (warts). Typically, DNA viruses enter the host cell nucleus, where the viral DNA is transcribed into messenger RNA (mRNA) by host cell polymerase and mRNA is translated into virus-specific proteins. 

Idoxuridine (5-iodo-2 -deoxyuridine) is an iodinated thymidine analog that inhibits the in vitro replication of various DNA viruses, including herpesviruses and poxviruses. 

Streptovaricin B (307), tolypomycin and geldanamycin are examples of ansamycins which are chemically related to rifamycins. The streptovaricins and tolypomycins resemble rifamycins in RT-inhibitory activity [18]. Streptovaricin B inhibits the replication of poxviruses by inhibiting early stages of mRNA synthesis [18]. Inhibition of focus formation of MSV by streptovaricins is also reported [18]. 

Poxvirus vectors encompass vaccinia derived from the smallpox vaccine and more attenuated variants like ALVAC or MVA (Modified Vaccinia Ankara). Their genome consists of single-stranded DNA of 130 to 300 kb pairs. Replication is restricted to the cytoplasm of cells and high amounts of protein are synthesized by the cell following transduction. Most applications therefore involve intramuscular vaccination. 

An interesting question derived from these studies is why viruses utilize different strategies to modulate chemokine activity. While vCKBPs have been identified mainly in poxviruses, herpesviruses frequently encode vCKs and vCKRs. This may reflect the need to modulate different aspects of chemokine biology as a result of diverse viral replication mechanisms. 

The identification of the SECRET domain in five different poxvirus proteins is intr uit. This distribution may explain, in part, the variety of genes encoding TNFR homologues in poxvirus genomes, some of which (CrmB and CrmD) encode this additional chemokme-inhilntory activity. It may also provide the virus the abihty to differentially block chemokines involved in controllii distinct antiviral responses, inhibit chemokines at different sta of infection in the animal host or simultaneously inhibit chemokines and TNF. It is likely that as poxviruses with narrow host species specificity adapted to particular hosts (Le., VARY to humans or ECTV to mice), nes were selected to facilitate viral replication and transmission in each host. 

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