Ann E Campbell

Publications of Ann E Campbell

• Regulation of PURA gene transcription by three promoters generating distinctly spliced 5-prime leaders: a novel means of fine control over tissue specificity and viral signals.

  Authors: Margaret J Wortman, Laura K Hanson, Luis Martínez-Sobrido, Ann E Campbell, Jonas A Nance, Adolfo García-Sastre, Edward M Johnson

  BMC molecular biology. 11/2010; 11:81.

  Purα is an evolutionarily conserved cellular protein participating in processes of DNA replication, transcription, and RNA transport; all involving binding to nucleic acids and altering conformationPurα is an evolutionarily conserved cellular protein participating in processes of DNA replication, transcription, and RNA transport; all involving binding to nucleic acids and altering conformation and physical positioning. The distinct but related roles of Purα suggest a need for expression regulated differently depending on intracellular and external signals. Here we report that human PURA (hPURA) transcription is regulated from three distinct and widely-separated transcription start sites (TSS). Each of these TSS is strongly homologous to a similar site in mouse chromosomal DNA. Transcripts from TSS I and II are characterized by the presence of large and overlapping 5'-UTR introns terminated at the same splice receptor site. Transfection of lung carcinoma cells with wild-type or mutated hPURA 5' upstream sequences identifies different regulatory elements. TSS III, located within 80 bp of the translational start codon, is upregulated by E2F1, CAAT and NF-Y binding elements. Transcription at TSS II is downregulated through the presence of adjacent consensus binding elements for interferon regulatory factors (IRFs). Chromatin immunoprecipitation reveals that IRF-3 protein binds hPURA promoter sequences at TSS II in vivo. By co-transfecting hPURA reporter plasmids with expression plasmids for IRF proteins we demonstrate that several IRFs, including IRF-3, down-regulate PURA transcription. Infection of NIH 3T3 cells with mouse cytomegalovirus results in a rapid decrease in levels of mPURA mRNA and Purα protein. The viral infection alters the degree of splicing of the 5'-UTR introns of TSS II transcripts. Results provide evidence for a novel mechanism of transcriptional control by multiple promoters used differently in various tissues and cells. Viral infection alters not only the use of PURA promoters but also the generation of different non-coding RNAs from 5'-UTRs of the resulting transcripts.

• Murine cytomegalovirus US22 protein pM140 protects its binding partner, pM141, from proteasome-dependent but ubiquitin-independent degradation.

  Authors: Lisa L Bolin, Laura K Hanson, Jacquelyn S Slater, Julie A Kerry, Ann E Campbell

  Journal of virology. 12/2009;

  Stable assembly of murine cytomegalovirus (MCMV) virions in differentiated macrophages is dependent upon the expression of US22 family gene M140. The M140 protein (pM140) exists in complex withStable assembly of murine cytomegalovirus (MCMV) virions in differentiated macrophages is dependent upon the expression of US22 family gene M140. The M140 protein (pM140) exists in complex with products of neighboring US22 genes. Here we report that pM140 protects its binding partner, pM141, from ubiquitin-independent proteasomal degradation. Protection is conferred by a stabilization domain mapping to amino acids 306-380 within pM140, and this domain is functionally independent from the region that confers binding of pM140 to pM141. The M140 protein thus contains multiple domains that collectively confer a structure necessary to function in virion assembly in macrophages.

• Murine Cytomegalovirus Capsid Assembly is Dependent upon US22 Family Gene M140 in Infected Macrophages.

  Authors: Laura K Hanson, Jacquelyn S Slater, Victoria J Cavanaugh, William W Newcomb, Lisa L Bolin, Christine N. Nelson, Lisa D. Fetters, Qiyi Tang, Jay C Brown, Gerd G Maul, Ann E Campbell

  Journal of virology. 06/2009;

  Macrophages are an important target cell for infection with cytomegalovirus. A number of viral genes have now been identified that are either expressed specifically in this cell type, or function toMacrophages are an important target cell for infection with cytomegalovirus. A number of viral genes have now been identified that are either expressed specifically in this cell type, or function to optimize cytomegalovirus replication in this host cell. Among these is the murine cytomegalovirus (MCMV) US22 gene family member M140, a nonessential early gene whose deletion (RVDelta140) leads to significant impairment in virus replication in differentiated macrophages. We have now determined that the defect in replication is at the stage of viral DNA encapsidation. Although the rate of RVDelta140 genome replication and extent of DNA cleavage was comparable to revertant virus, deletion of M140 resulted in a significant reduction in the number of viral capsids in the nucleus, and the viral DNA remained sensitive to DNase treatment. These data are indicative of incomplete virion assembly. Steady state levels of both the major capsid protein (M86) and tegument protein M25 were reduced in the absence of the M140 protein (pM140). This effect may be related to the localization of pM140 to an aggresome-like, microtubule organizing center-associated structure, known to target misfolded and over-expressed proteins for degradation. It appears therefore that pM140 indirectly influences MCMV capsid formation in differentiated macrophages by regulating the stability of viral structural proteins.

• Inhibition of norovirus replication by morpholino oligomers targeting the 5'-end of the genome.

  Authors: Karin Bok, Victoria J Cavanaugh, David O Matson, Lorenzo González-Molleda, Kyeong-Ok Chang, Carmelann Zintz, Alvin W Smith, Patrick Iversen, Kim Y Green, Ann E Campbell

  Virology. 10/2008;

  Noroviruses are an important cause of non-bacterial epidemic gastroenteritis, but no specific antiviral therapies are available. We investigated the inhibitory effect of phosphorodiamidiateNoroviruses are an important cause of non-bacterial epidemic gastroenteritis, but no specific antiviral therapies are available. We investigated the inhibitory effect of phosphorodiamidiate morpholino oligomers (PMOs) targeted against norovirus sequences. A panel of peptide-conjugated PMOs (PPMOs) specific for the murine norovirus (MNV) genome was developed, and two PPMO compounds directed against the first AUG of the ORF1 coding sequence near the 5'-end of the genome proved effective in inhibiting MNV replication in cells. A consensus PPMO (designated Noro 1.1), designed to target the corresponding region of several diverse human norovirus genotypes, decreased the efficiency of protein translation in a cell-free luciferase reporter assay and inhibited Norwalk virus protein expression in replicon-bearing cells. Our data suggest that PPMOs directed against the relatively conserved 5'-end of the norovirus genome may show broad antiviral activity against this genetically diverse group of viruses.

• The salivary glands as a privileged site of cytomegalovirus immune evasion and persistence.

  Authors: Ann E Campbell, Victoria J Cavanaugh, Jacquelyn S Slater

  Medical microbiology and immunology. 07/2008; 197(2):205-13.

  The salivary glands (SG) provide a haven for persistent cytomegalovirus replication, and in this regard are a privileged site of virus immune evasion. The murine cytomegalovirus (MCMV) model hasThe salivary glands (SG) provide a haven for persistent cytomegalovirus replication, and in this regard are a privileged site of virus immune evasion. The murine cytomegalovirus (MCMV) model has provided insight into the immunological environment of the SG and the unqiue virus-host relationship of this organ. In response to MCMV infection, a robust T cell-mediated immune response is elicited, comprised predominantly of CD8+ T cells that phenotypically and functionally appear activated. However, they fail to clear virus by an unknown evasion mechanism that is independent of inhibitory NKG2A- or Programmed Death 1-mediated signaling. Virus is eventually eliminated from the SG by effector CD4+ T cells expressing antiviral cytokines. However, this mechanism is severely dampened by high levels of the immunosuppressive cytokine IL-10, selectively expressed by SG CD4+ T cells.

• Upregulation of CD94/NKG2A receptors and Qa-1b ligand during murine cytomegalovirus infection of salivary glands.

  Authors: Victoria J Cavanaugh, David H Raulet, Ann E Campbell

  The Journal of general virology. 06/2007; 88(Pt 5):1440-5.

  Following acute infection, murine cytomegalovirus (MCMV) replicates persistently in the salivary glands, despite the vigorous response of activated CD8 T cells that infiltrate this gland.Following acute infection, murine cytomegalovirus (MCMV) replicates persistently in the salivary glands, despite the vigorous response of activated CD8 T cells that infiltrate this gland. Virus-specific CD8 T lymphocytes isolated from this organ were found to express the inhibitory CD94/NKG2A receptor that, in some virus models, confers an inhibitory response to cytotoxic T lymphocytes (CTLs). In response to MCMV infection, expression of the CD94/NKG2A ligand, Qa-1b, increased dramatically in the submandibular gland (SMG) prior to upregulation of H-2Dd. However, there was no net negative impact on virus-specific T-cell function, as virus titres were similar in CD94- and CD94+ mice. CD94/NKG2A expression, also known to inhibit apoptosis, did not influence the numbers of accumulated T, NK and NK T cells. These data indicate that expression of inhibitory CD94/NKG2A receptors does not account for the failure of MCMV-specific CTLs to clear the SMG of infection.

• Characterization and regulation of essential murine cytomegalovirus genes m142 and m143.

  Authors: Laura K Hanson, Bridget L Dalton, Laura F Cageao, Rachel E Brock, Jacquelyn S Slater, Julie A Kerry, Ann E Campbell

  Virology. 05/2005; 334(2):166-77.

  US22 gene family members m142 and m143 are essential for replication of murine cytomegalovirus (MCMV). Their transcripts are produced with immediate-early kinetics, but little else is known aboutUS22 gene family members m142 and m143 are essential for replication of murine cytomegalovirus (MCMV). Their transcripts are produced with immediate-early kinetics, but little else is known about these viral genes. Unlike their transcripts, the m142 and m143 gene products (pm142, pm143) were not expressed until early times post-infection, with levels increasing over the course of infection. Both pm142 and pm143 were predominantly cytoplasmic, but cellular fractionation studies confirmed that the proteins were present in the nucleus as well. In addition, pm142 was detected within the virion. Both the m142 and m143 promoters were strongly upregulated by viral infection or by MCMV IE1. However, UV-inactivated virus and IE3 upregulated only the m142 promoter. When tested for transcriptional transactivating activity, neither m142 nor m143 demonstrated significant activity, either alone or in combination with the major immediate-early gene products. This failure to transactivate, along with their essential nature, makes m142 and m143 unique among the immediate-early genes of the US22 gene family.

• Complex formation among murine cytomegalovirus US22 proteins encoded by genes M139, M140, and M141.

  Authors: Zaruhi Karabekian, Laura K Hanson, Jacquelyn S Slater, Neel K Krishna, Lisa L Bolin, Julie A Kerry, Ann E Campbell

  Journal of virology. 04/2005; 79(6):3525-35.

  The murine cytomegalovirus (MCMV) proteins encoded by US22 genes M139, M140, and M141 function, at least in part, to regulate replication of this virus in macrophages. Mutant MCMV having one or moreThe murine cytomegalovirus (MCMV) proteins encoded by US22 genes M139, M140, and M141 function, at least in part, to regulate replication of this virus in macrophages. Mutant MCMV having one or more of these genes deleted replicates poorly in macrophages in culture and in the macrophage-dense environment of the spleen. In this report, we demonstrate the existence of stable complexes formed by the products of all three of these US22 genes, as well as a complex composed of the products of M140 and M141. These complexes form in the absence of other viral proteins; however, the pM140/pM141 complex serves as a requisite binding partner for the M139 gene products. Products from all three genes colocalize to a perinuclear region of the cell juxtaposed to or within the cis-Golgi region but excluded from the trans-Golgi region. Interestingly, expression of pM141 redirects pM140 from its predominantly nuclear residence to the perinuclear, cytoplasmic locale where these US22 proteins apparently exist in complex. Thus, complexing of these nonessential, early MCMV proteins likely confers a function(s) independent of each individual protein and important for optimal replication of MCMV in its natural host.

• Age-related impaired type 1 T cell responses to influenza: reduced activation ex vivo, decreased expansion in CTL culture in vitro, and blunted response to influenza vaccination in vivo in the elderly.

  Authors: Yuping Deng, Yu Jing, Ann E Campbell, Stefan Gravenstein

  Journal of immunology (Baltimore, Md. : 1950). 04/2004; 172(6):3437-46.

  The objective of this study was to analyze the changes in the type 1 T cell response, including the CD4+ Th1 and CD8+ T cell responses, to influenza in the elderly compared with those in youngThe objective of this study was to analyze the changes in the type 1 T cell response, including the CD4+ Th1 and CD8+ T cell responses, to influenza in the elderly compared with those in young adults. PBMC activated ex vivo with influenza virus exhibited an age-related decline in type 1 T cell response, shown by the decline in the frequency of IFN-gamma-secreting memory T cells specific for influenza (IFN-gamma+ ISMT) using ELISPOT or intracellular cytokine staining. The reduced frequency of IFN-gamma+ ISMT was accompanied by a reduced level of IFN-gamma secretion per cell in elderly subjects. Tetramer staining, combined with IFN-gamma ELISPOT, indicated that the decline in IFN-gamma+, influenza M1-peptide-specific T cells was not due to attrition of the T cell repertoire, but, rather, to the functional loss of ISMT with age. In addition, the decline in type 1 T cell response was not due to an increase in Th2 response or defects in APCs from the elderly. The expansion of influenza-specific CD8+ T cells in CTL cultures was reduced in the elderly. Compared with young subjects, frail elderly subjects also exhibited a blunted and somewhat delayed type 1 T cell response to influenza vaccination, which correlated positively with the reduced IgG1 subtype and the total Ab response. Taken together, these data demonstrate that there is a decline in the type 1 T cell response to influenza with age that may help explain the age-related decline in vaccine efficacy and the increases in influenza morbidity and mortality.

• Role of murine cytomegalovirus US22 gene family members in replication in macrophages.

  Authors: Carine Ménard, Markus Wagner, Zsolt Ruzsics, Karina Holak, Wolfram Brune, Ann E Campbell, Ulrich H. Koszinowski

  Journal of virology. 06/2003; 77(10):5557-70.

  The large cytomegalovirus (CMV) US22 gene family, found in all betaherpesviruses, comprises 12 members in both human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV). Conserved sequenceThe large cytomegalovirus (CMV) US22 gene family, found in all betaherpesviruses, comprises 12 members in both human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV). Conserved sequence motifs suggested a common ancestry and related functions for these gene products. Two members of this family, m140 and m141, were recently shown to affect MCMV replication on macrophages. To test the role of all US22 members in cell tropism, we analyzed the growth properties in different cell types of MCMV mutants carrying transposon insertions in all 12 US22 gene family members. When necessary, additional targeted mutants with gene deletions, ATG deletions, and ectopic gene revertants were constructed. Mutants with disruption of genes M23, M24, m25.1, m25.2, and m128 (ie2) showed no obvious growth phenotype, whereas growth of M43 mutants was reduced in a number of cell lines. Genes m142 and m143 were shown to be essential for virus replication. Growth of mutants with insertions into genes M36, m139, m140, and m141 in macrophages was severely affected. The common phenotype of the m139, m140, and m141 mutants was explained by an interaction at the protein level. The M36-dependent macrophage growth phenotype could be explained by the antiapoptotic function of the gene that was required for growth on macrophages but not for growth on other cell types. Together, the comprehensive set of mutants of the US22 gene family suggests that individual family members have diverged through evolution to serve a variety of functions for the virus.

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• Vigorous innate and virus-specific cytotoxic T-lymphocyte responses to murine cytomegalovirus in the submaxillary salivary gland.

  Authors: Victoria J Cavanaugh, Yuping Deng, Mark P Birkenbach, Jacquelyn S Slater, Ann E Campbell

  Journal of virology. 03/2003; 77(3):1703-17.

  To better understand the immunological mechanisms that permit prolonged shedding of murine cytomegalovirus (MCMV) from the salivary gland, the phenotypic and functional characteristics of leukocytesTo better understand the immunological mechanisms that permit prolonged shedding of murine cytomegalovirus (MCMV) from the salivary gland, the phenotypic and functional characteristics of leukocytes infiltrating the submaxillary gland (SMG) were analyzed in infected BALB/c mice. A robust innate immune response, comprised of CD11c+ major histocompatibility complex class II+ CD11b- CD8alpha+ dendritic cells and gamma/delta T-cell receptor-bearing CD3+ T cells was prominent through at least 28 days postinfection. Concurrently, a dramatic increase in pan-NK (DX5+) CD3+ and CD8+ T cells was observed, while CD4+ T cells, known to be essential for viral clearance from this tissue, increased slightly. The expression particularly of gamma interferon but also of interleukin-10 and CC chemokines was extraordinarily high in the SMG in response to MCMV infection. The gamma interferon was produced primarily by CD4+ and CD8+ T lymphocytes and DX5+ CD3+ T cells. The SMG CD8+ T cells were highly cytolytic ex vivo, and a significant proportion of these cells were specific to an immunodominant MCMV peptide. These peptide-specific clones were not exhausted by the presence of high virus titers, which persisted in the SMG despite the strength of the cell-mediated responses. In contrast, MCMV replication was efficiently cleared from the draining cervical and periglandular lymph nodes, a tissue displaying a substantially weaker antiviral response. Our data indicated that vigorous innate and acquired immune responses are elicited, activated, and retained in response to mucosal inflammation from persistent MCMV infection of the submaxillary gland.

• Identification and characterization of novel murine cytomegalovirus M112-113 (e1) gene products.

  Authors: Gina M Ciocco-Schmitt, Zaruhi Karabekian, Earl W Godfrey, Richard M Stenberg, Ann E Campbell, Julie A Kerry

  Virology. 04/2002; 294(1):199-208.

  The human cytomegalovirus (HCMV) UL112-113 gene products play important roles in viral DNA replication and transcriptional regulation. In this report, we characterize two novel transcriptsThe human cytomegalovirus (HCMV) UL112-113 gene products play important roles in viral DNA replication and transcriptional regulation. In this report, we characterize two novel transcripts originating from the homologous M112-113 (e1) region of the murine cytomegalovirus (MCMV) genome. These transcripts of 2.0 and 2.4 kb represent alternatively spliced products of the e1 gene region. Analysis of the e1 proteins demonstrates the presence of a previously unidentified 87-kDa protein that is likely encoded by the 2.4-kb transcript. All four protein products derived from the e1 gene region are expressed with early kinetics, are coordinately regulated, and localize predominantly to the nucleus of MCMV-infected cells. The expression pattern and localization of the e1 proteins show significant similarity to those of the HCMV UL112-113 proteins, signifying that MCMV e1 will serve as a useful model for assessing the role of this early gene region during viral infection.

• Inhibition of norovirus replication by morpholino oligomers targeting the 5′-end of the genome

  Authors: Karin Bok, Victoria J. Cavanaugh, David O. Matson, Lorenzo González-Molleda, Kyeong-Ok Chang, Carmelann Zintz, Alvin W. Smith, Patrick Iversen, Kim Y. Green, Ann E. Campbell

  Virology.

  Noroviruses are an important cause of non-bacterial epidemic gastroenteritis, but no specific antiviral therapies are available. We investigated the inhibitory effect of phosphorodiamidiateNoroviruses are an important cause of non-bacterial epidemic gastroenteritis, but no specific antiviral therapies are available. We investigated the inhibitory effect of phosphorodiamidiate morpholino oligomers (PMOs) targeted against norovirus sequences. A panel of peptide-conjugated PMOs (PPMOs) specific for the murine norovirus (MNV) genome was developed, and two PPMO compounds directed against the first AUG of the ORF1 coding sequence near the 5′-end of the genome proved effective in inhibiting MNV replication in cells. A consensus PPMO (designated Noro 1.1), designed to target the corresponding region of several diverse human norovirus genotypes, decreased the efficiency of protein translation in a cell-free luciferase reporter assay and inhibited Norwalk virus protein expression in replicon-bearing cells. Our data suggest that PPMOs directed against the relatively conserved 5′-end of the norovirus genome may show broad antiviral activity against this genetically diverse group of viruses.

• Murine cytomegalovirus independently inhibits priming of helper and cytotoxic T lymphocytes

  Authors: Jacquelyn S. Slater, W. Stewart Futch, Victoria J. Cavanaugh, Ann E. Campbell

  Virology.

  Murine cytomegalovirus (MCMV) inhibits antigen-specific cytotoxic T lymphocyte (CTL) priming in vivo (Campbell et al., 1989). To address the mechanism of this immune suppression, two possibilitiesMurine cytomegalovirus (MCMV) inhibits antigen-specific cytotoxic T lymphocyte (CTL) priming in vivo (Campbell et al., 1989). To address the mechanism of this immune suppression, two possibilities were considered: (1) MCMV directly interferes with in vivo priming of CTL precursors (CTLp), or (2) MCMV suppresses T helper cell functions necessary for CTL priming. We therefore quantitated T helper cell function in MCMV-infected, SV40-immune mice and assessed dependency of SV40-specific CTLp priming on T helper cell activity. MCMV infection of H-2b mice significantly suppressed the frequency of IL-2 producing T helper cells generated in SV40-immune mice. This suppression was not due to alterations in the number or percentage of CD4 lymphocytes. The helper cell deficiency correlated with suppressed SV40-specific CTL activity. However, CTLp priming in vivo was found to be independent of CD4 T helper cells and IL-2. Therefore, the suppressive effects of MCMV on helper and cytotoxic T cell functions are independent, implying that MCMV directly inhibits an event in lymphocyte priming common to both helper and cytotoxic T cells.

• Characterization and regulation of essential murine cytomegalovirus genes m142 and m143

  Authors: Laura K. Hanson, Bridget L. Dalton, Laura F. Cageao, Rachel E. Brock, Jacquelyn S. Slater, Julie A. Kerry, Ann E. Campbell

  Virology.

  US22 gene family members m142 and m143 are essential for replication of murine cytomegalovirus (MCMV). Their transcripts are produced with immediate-early kinetics, but little else is known aboutUS22 gene family members m142 and m143 are essential for replication of murine cytomegalovirus (MCMV). Their transcripts are produced with immediate-early kinetics, but little else is known about these viral genes. Unlike their transcripts, the m142 and m143 gene products (pm142, pm143) were not expressed until early times post-infection, with levels increasing over the course of infection. Both pm142 and pm143 were predominantly cytoplasmic, but cellular fractionation studies confirmed that the proteins were present in the nucleus as well. In addition, pm142 was detected within the virion. Both the m142 and m143 promoters were strongly upregulated by viral infection or by MCMV IE1. However, UV-inactivated virus and IE3 upregulated only the m142 promoter. When tested for transcriptional transactivating activity, neither m142 nor m143 demonstrated significant activity, either alone or in combination with the major immediate-early gene products. This failure to transactivate, along with their essential nature, makes m142 and m143 unique among the immediate-early genes of the US22 gene family.

• Murine cytomegalovirus-induced suppression of antigen-specific cytotoxic T lymphocyte maturation

  Authors: Ann E. Campbell, Jacquelyn S. Slater, W.Stewart Futch

  Virology.

  Antigen-specific cytotoxic T lymphocyte (CTL) maturation was inhibited in mice acutely infected with murine cytomegalovirus(MCMV). When immunization with Simian virus 40 (SV40) either preceded orAntigen-specific cytotoxic T lymphocyte (CTL) maturation was inhibited in mice acutely infected with murine cytomegalovirus(MCMV). When immunization with Simian virus 40 (SV40) either preceded or followed infection with MCMV by 1 day, the frequency of SV40-specific CTL precursors among lymph node cells (LNC) was significantly reduced compared to noninfected mice. Replication of the herpesvirus in LNC could not be detected; however, MCMV rendered noninfectious by heat treatment was not suppressive to CTL development. Lymph node cells from nonimmunized, MCMV-infected mice contained a cell(s) present in low frequency which suppressed in vitro maturation of SV40 CTL in immune lymph nodes from mice not infected with MCMV. These suppressor cells did not affect the antigen- and interleukin-2-dependent growth nor cytotoxic activity of a mature, SV40-specific CTL clone. These results indicate that MCMV interferes with immunoregulatory functions required for development of antigen-specific CTL precursors to mature effector CTL. The immunosuppression is mediated at least in part by the MCMV-induced suppressor cell(s).

• Transcriptional Analysis of the Murine Cytomegalovirus HindIII-I Region: Identification of a Novel Immediate-Early Gene Region

  Authors: Laura K. Hanson, Bridget L. Dalton, Zaruhi Karabekian, Helen E. Farrell, William D. Rawlinson, Richard M. Stenberg, Ann E. Campbell

  Virology.

  Cytomegaloviruses likely encode numerous gene products involved in regulating virus–host cell interactions and pathogenesis. We previously identified a region of murine cytomegalovirus (MCMV) withinCytomegaloviruses likely encode numerous gene products involved in regulating virus–host cell interactions and pathogenesis. We previously identified a region of murine cytomegalovirus (MCMV) within HindIII-J and -I that regulates pathogenesis of the virus [open reading frames (ORFs) M139–M141] or is likely required for MCMV replication (ORFs m142 and m143). As a prerequisite for further studies on the structure and function of this gene region, we mapped the transcripts encoded within MCMV HindIII-I. Probes for ORFs M140 and M141 hybridized to 5.4- and 7.0-kb RNA, respectively, which were transcribed with early kinetics and were 3′ coterminal with HindIII-J ORF M139. Probes representing ORFs m142, m143, or m144 hybridized to 3′ coterminal transcripts of 1.8, 3.8, and 5.1 kb, respectively. ORFs m142 and m143 were transcribed with immediate-early kinetics but were most abundantly expressed at early times. Probes for the rightmost end of HindIII-I hybridized to a 5.1-kb early/late RNA corresponding to m144 and to a 1.8-kb early RNA transcribed from m145. All of the major transcripts were polyadenylated and therefore are likely coding. Additional minor transcripts of intermediate sizes were also detected. ORFs M139–m143 showed homology to the betaherpesvirus-specific HCMV US22 gene family. Because deletion of these viral genes results in attenuated or helper-dependent phenotypes, this conserved region of US22 family genes may have a role in virus replication as well as in the pathogenesis of betaherpesviruses in their natural hosts.

• Enhancement of interleukin-1 activity by murine cytomegalovirus infection of a macrophage cell line

  Authors: Margaret A Lewis, Jacquelyn S Slater, Laura I Leverone, Ann E Campbell

  Virology.

  The effect of murine cytomegalovirus (MCMV) infection on interleukin 1 (IL-1) secretion was assessed using the macrophage cell lines P388D1 and 1774A.1. The former proved to be nonpermissive for MCMVThe effect of murine cytomegalovirus (MCMV) infection on interleukin 1 (IL-1) secretion was assessed using the macrophage cell lines P388D1 and 1774A.1. The former proved to be nonpermissive for MCMV in that infectious virus and viral immediate early protein (pp89) were not expressed in these cells. MCMV infection of the P388D1 cells had no effect on release of biologically active IL-1. In contrast, J774A.1 cells, which were semi permissive for virus replication and pp89 expression, secreted enhanced levels of IL-1 activity following infection. The enhancement was evident when infection either preceded or followed lipopolysaccharide stimulation of the macrophages. The relative proportion of IL-1 α and β secreted from MCMV-infected cells was similar to noninfected controls. In addition, the bioactivity of intracellular IL-1 α escaping membranes of fixed cells was unaffected by virus infection. From these findings, we conclude that limited MCMV expression in the J774A.1 macrophage cell line enhances secretion of IL-1 α and β bioactivity.