Bernard Roizman

Herpes simplex virus infections

Herpes simplex virus (HSV) is a member of the herpesviridae family. Recognised since ancient Greek times, the virus frequently infects human beings, causing a range of diseases from mild uncomplicated mucocutaneous infection to those that are life threatening. In the past 50 years, substantial advances in our knowledge of the molecular biology of HSV have led to insights into disease pathogenesis and management. This review provides a contemporary interpretation of the biological properties, function, epidemiology, and treatment of HSV diseases.

Characterization of herpes simplex virus strains differing in their effects on social behaviour of infected cells.

Summary Established (laboratory) strains and fresh isolates of herpes simplex virus from patients with skin and genital lesions were classified into four groups depending on their effects on the social interaction among infected hep-2 cells. The groups comprised strains causing (1) rounding of cells but no adhesion or fusion, (2) loose aggregation of rounded cells, (3) tight adhesion of rounded cells, and (4) fusion of cells into polykaryocytes. Protype strains from each group were found to differ with respect to immunologic specificity, buoyant density in CsCl solutions and stability at 40°.

The order Herpesvirales

The taxonomy of herpesviruses has been updated by the International Committee on Taxonomy of Viruses (ICTV). The former family Herpesviridae has been split into three families, which have been incorporated into the new order Herpesvirales. The revised family Herpesviridae retains the mammal, bird and reptile viruses, the new family Alloherpesviridae incorporates the fish and frog viruses, and the new family Malacoherpesviridae contains a bivalve virus. Three new genera have been created in the family Herpesviridae, namely Proboscivirus in the subfamily Betaherpesvirinae and Macavirus and Percavirus in the subfamily Gammaherpesvirinae. These genera have been formed by the transfer of species from established genera and the erection of new species, and other new species have been added to some of the established genera. In addition, the names of some nonhuman primate virus species have been changed. The family Alloherpesviridae has been populated by transfer of the genus Ictalurivirus and addition of the new species Cyprinid herpesvirus 3. The family Malacoherpesviridae incorporates the new genus Ostreavirus containing the new species Ostreid herpesvirus 1.

Human Herpesviruses: Biology, Therapy, and Immunoprophylaxis

Part I. Introduction Part II. Basic Virology and Viral Gene Effects on Host Cell Functions Part II. Basic Virology and Viral Gene Effects on Host Cell Functions Part II. Basic Virology and Viral Gene Effects on Host Cell Functions Part III. Pathogenesis, Clinical Disease, Host Response, and Epidemiology Part III. Pathogenesis, Clinical Disease, Host Response, and Epidemiology Part III. Pathogenesis, Clinical Disease, Host Response, and Epidemiology Part III. HHV- 6a, 6b and 7 Ann Arvin and Richard Whitley Part III. Pathogenesis, Clinical Disease, Host Response, and Epidemiology Part IV. Non-Human Primate Herpesviruses Ann Arvin, Patrick Moore and Richard Whitley Part V. Subversion of Adaptive Immunity Richard Whitley and Ann Arvin Part VI. Antiviral Therapy Richard Whitley Part VII. Vaccines and Immunotherapy Ann Arvin and Koichi Yamanishi Part VIII. Herpes as Therapeutic Agents Richard Whitley and Bernard Roizman.

A generalized technique for deletion of specific genes in large genomes: a gene 22 of herpes simplex virus 1 is not essential for growth

We describe a general method for inactivation and deletion of genes at specific sites in large DNA genomes. In the first step of the procedure, the herpes simplex virus thymidine kinase is inserted into the genome at a specific site. In the second step, the thymidine kinase gene is desired sequences flanking the insertion site are deleted. Both steps involve recombination of the genomes with cloned chimeric fragments and utilize the available selection for or against thymidine kinase to select the desired genomes. We have applied the procedure to inactivate and to delete portions of an alpha gene of herpes simplex virus 1 specifying protein 22. The recombinant virus carrying the thymidine kinase inserted into the gene 22 and viruses exhibiting 0.1 kb and 0.7 kb deletions in the gene 22 specify new alpha polypeptides with molecular weights approximately 30% of the wild-type gene 22 product and grown normally in Vero cell cultures.

An interferon-related gene signature for DNA damage resistance is a predictive marker for chemotherapy and radiation for breast cancer

Individualization of cancer management requires prognostic markers and therapy-predictive markers. Prognostic markers assess risk of disease progression independent of therapy, whereas therapy-predictive markers identify patients whose disease is sensitive or resistant to treatment. We show that an experimentally derived IFN-related DNA damage resistance signature (IRDS) is associated with resistance to chemotherapy and/or radiation across different cancer cell lines. The IRDS genes STAT1, ISG15, and IFIT1 all mediate experimental resistance. Clinical analyses reveal that IRDS(+) and IRDS(−) states exist among common human cancers. In breast cancer, a seven–gene-pair classifier predicts for efficacy of adjuvant chemotherapy and for local-regional control after radiation. By providing information on treatment sensitivity or resistance, the IRDS improves outcome prediction when combined with standard markers, risk groups, or other genomic classifiers.

Treatment of intracranial gliomas in immunocompetent mice using herpes simplex viruses that express murine interleukins

This report describes a test of the hypothesis that the oncolytic effect of genetically engineered, replication competent herpes simplex viruses (HSV) depends both on cell destruction by the virus and an immune response to the tumor cells induced in an immunocompetent animal system. The oncolytic vector was a HSV recombinant virus in which both copies of the γ134.5 gene were replaced with the murine genes encoding the cytokine interleukin-4 (IL-4) or interleukin-10 (IL-10). The hypothesis predicted that if an immune response plays a role in survival following intratumoral treatment of tumor-bearing animals with HSV, expression of IL-4 should prolong survival whereas expression of IL-10 should reduce it. The results are that (1) these cytokines can be expressed by HSV in productively infected cells both in vitro and in vivo; (2) HSV-expressing IL-4 or IL-10 genes were able to infect and destroy glioma cells in vitro; (3) intracerebral inoculation of HSV expressing either IL-4 or IL-10 into syngeneic murine glioma GL-261 cells implanted in the brains of immunocompetent C57BL/6 mice produced dramatically opposite physiologic responses. The IL-4 HSV significantly prolonged survival of tumor bearers, whereas tumor-bearing mice that received the IL-10 HSV had a median survival that was identical to that of saline treated controls; (4) immunohistochemical analyses of mouse brains at 3 and 7 days after virus inoculation showed marked accumulation of inflammatory cells composed primarily of macro- phages/microglia, with various proportions of CD8+ and CD4+ T cells, but few B lymphocytes. We conclude that the cytokines expressed from genes encoded in the viral genome influence HSV therapy of tumors and this is probably due to the host immune response. Thus, cytokine expression may be an important adjunct to tumor therapy utilizing genetically engineered HSV.

Regulation of α genes of herpes simplex virus: Expression of chimeric genes produced by fusion of thymidine kinase with α gene promoters

Abstract We report a system for investigating promoters of eucaryotic cell and virus genes based on analyses of the regulation of herpes simplex virus 1 (HSV-1) thymidine kinases whose structural gene sequences have been fused to the promoter of the gene under study. In infected cells, the polypeptides specified by HSV-1 form at least three groups, α, β and γ, whose synthesis is coordinately regulated and sequentially ordered at the transcriptional level. To identify the DNA sequence responsible for the regulation of transcription of α genes, we fused the sequence encoding the 5′ end of an α gene to the structural gene sequence of the thymidine kinase, a β gene. The resultant recombinant DNA was inserted into the viral genome and was also used to convert Ltk − cells to tk + phenotype. In cells infected with recombinant virus, the thymidine kinase gene was regulated and expressed as an α gene—that is, it was transcribed and processed in the absence of prior infected cell protein synthesis. Moreover, mRNA selected by hybridization to sequences encoding the thymidine kinase contains at its 5′ terminus sequences homologous to the donor sequence encoding the 5′ terminus of the α mRNA. In converted tk + cells, the fused thymidine kinase gene, like the wild-type gene, is stimulated by superinfection with the tk − virus. However, the stimulation is many times greater and is due to non-α-gene products, whereas in cells converted by the wild-type gene, the stimulation is by a gene products. We conclude that the α genes are identified for transcription by sequences at or near those encoding the 5′ terminus of the mRNA, and transposition of these sequences to a β gene is all that is required to convert it to an α gene. Transcription of α genes appears to be regulated by non-α-gene products, which could be contained within the structure of the virion. In converted Ltk + cells, the thymidine kinase gene uses its own promoter.

Role of ICP0 in the strategy of conquest of the host cell by herpes simplex virus 1.

In early studies, herpes simplex virus 1 (HSV-1) proteins were identified on the basis of two criteria. The first consisted of characterization of proteins contained in virions purified from cells whose proteins were labeled prior to infection. These proteins designated by the prefix VP were numbered in the order of decreasing apparent molecular weight or, conversely, ascending electrophoretic mobility in denaturing gels (147). The second criterion identified putative viral proteins accumulating in infected cells but absent from uninfected cells. These proteins, designated infected-cell proteins (ICPs), were also numbered in order of decreasing apparent molecular weight (73). One protein, however, while clearly apparent only in infected cells, varied with respect to electrophoretic mobility depending on the composition of the denaturing gel (74). This anomalously migrating protein was designated ICP0 (74). In subsequent studies, viral proteins were designated either by their known primary function (e.g., DNA polymerase, etc.) or the position of the gene along the unique long (UL) or short (US) components of the viral genome (105, 107). The original ICP designation, however, was retained primarily for five proteins recognized in early studies as being the products of α or immediate-early genes expressed after infection in the absence of de novo viral protein synthesis (74, 75). The five proteins, ICP0, ICP4, ICP22, ICP27, and ICP47, have been extensively studied, and for the most part, there is at least a semblance of concordance between the phenotype of cells infected with the mutant lacking the gene, the behavior of the protein in transduced cells, and the molecular functions expressed by the protein (reviewed in reference 132). ICP4 is an essential positive and negative regulator of gene expression (reviewed in reference 132). The protein blocks gene expression by binding to high-affinity DNA consensus sites located at transcription initiation sites of at least two genes. The mechanism of gene activation is less understood, although ICP4’s affinity for transcription factors and binding to highly degenerate or nonconsensus sites are suggestive of how it might act. ICP27 is also a multifunctional protein whose phenotype can be largely explained by its ability to block RNA splicing but not transport of unspliced RNA early in infection and by its activity as a chaperone of newly made viral mRNA across the nuclear membranes at late times after infection (reviewed in reference 137). Available data indicate that the carboxyl-terminal half of ICP22 enables full expression of a subset of late (γ2) viral genes by causing cdc2 cyclin-dependent kinase activity to survive the degradation of its physiologic partners cyclins A and B, by an aberrant partnership with the UL42 DNA polymerase processivity factor (2, 5, 6, 19, 141). Optimal transcription of late genes requires binding and posttranslational modification of topoisomerase IIα by the two proteins (7). The sole known mission of ICP47 is to bind to and preclude TAP1/TAP2 from enabling the transport of antigenic peptides into the endoplasmic reticulum for eventual presentation on the cell surface (50, 71, 161). Despite 3 decades of research and enormous interest, it is not known how the functions encoded in ICP0 account for its phenotype in either infected or transduced cells. However, available evidence suggests that ICP0 is a multifunctional protein and that its role in viral infection reflects the sum of its multiple and diverse functions (132). Studies published nearly 20 years ago established that ICP0 activates genes introduced into cells by transfection or infection (29, 30, 54, 118, 129). Furthermore, ICP0 also activates a specific subset of cellular genes, including several p53-responsive genes that it activates independently of p53 (72). ICP0 is considered a promiscuous transactivator, inasmuch as it activates transcription from HSV (16, 22, 104) and heterologous (58, 114, 143) promoter elements independently of a single cis-acting element (40). ICP0 activates transcription of viral genes in synergy with or independent of ICP4 (29, 31, 55, 129). ICP0 has been shown to interact with ICP4, and this interaction is believed to mediate cooperative activation of gene expression, as it maps to a region of ICP0 (residues 617 to 775) that contains the domain involved in synergy with ICP4 (residues 680 to 767) (33, 159). In most cell lines infected at low multiplicity with mutants lacking the gene encoding ICP0 (Δα0), viral yields are 10- to 100-fold lower than those from cells infected with wild-type virus (34, 136, 148). At higher multiplicities of infection, viral yields and protein expression are similar to those of wild-type virus (34, 136, 148). Exceptions are a few cell lines, exemplified by the line U20S, in which Δα0 mutants replicate as well as wild-type viruses (160). Because of the lethargy of Δα0 mutants, ICP0 has been held responsible for the establishment of latency and a myriad of other functions. Nevertheless, none of these phenotypic properties of ICP0 correlate directly with the emerging patterns of interaction of ICP0 with cellular proteins—the subject of this review.

Virulence of and establishment of latency by genetically engineered deletion mutants of herpes simplex virus I

We report the results of studies on the biologic properties of seven deletion mutants of herpes simplex virus 1 (HSV-1). The genes deleted from six of these mutants map in the S component of HSV-1 DNA and include those specifying the alpha protein 47, the glycoproteins G and E, the viral protein kinase, and two proteins whose functions are not yet known (open reading frames US2 and US11). The seventh virus [HSV-1(F) delta 305] contained a 700-bp deletion in the thymidine kinase gene. The results of intracerebral inoculation of Balb/c mice indicated that all but one of the deletion mutants in the S component were significantly attenuated. The PFU/LD50 ratios for these mutants ranged from 10(4)- to 10(5)-fold higher than that of the wild-type, HSV-1(F). The PFU/LD50 for mutant R7032, from which the glycoprotein E gene had been deleted, was less than 100-fold higher than that of the parent virus. All of the mutants, with one exception, were able to establish latency in mice; the exception, HSV-1(F) delta 305, was able to establish latency in rabbits.