Quantitation of luciferase activity.Relative intensities of in vivo bioluminescence were represented as an overlapping pseudocolor images from violet (least intense) to red (most intense). For the detection of MHV-68 antigen, a polyclonal rabbit antiserum was used that had been generated in rabbits injected with purified MHV-68 particles. Venn diagram depicts the relative numbers of proteins and overlap of the control and infected proteomic data sets. B cells can be de novo infected in vitro, albeit with low efficiency, and latent B cell lines have been established (Usherwood et al., 1996b; Forrest and Speck, 2008; Liang et al., 2011). Medina, C. 1A). A total of 10 to 20 μg total RNA was hybridized overnight with the γ6 riboprobe set specific for selected γHV68 transcripts (41).
The frequency of cells that spontaneously reactivated from latency was calculated by Poisson distribution analysis of mean data from 3 experiments. Gammaherpesviruses share an ability to transform lymphocytes in natural or experimental hosts. The observation that none of the cycKO virus-infected mice died was surprising, since the cycKO virus previously showed no defect in acute replication or pathogenesis (4, 5, 21), with a single exception for a low-dose inoculation following intranasal infection (19). However, these initial studies have not been extended and a robust model for γHV68-induced lymphomas is still needed. Notably, both the Y120F and Y129F M2 mutants were expressed to similar levels as wild type M2 (Fig. Second, although the absence of mLANA correlated with increased p53 induction during MHV68 lytic replication, it is not known if p53 functions remain intact or are dominantly inhibited in the infected cell. In addition, individuals who are still EBV seronegative in adulthood are potentially resistant to both MS and EBV infection (46).
The granule exocytosis pathway has been shown to be important for regulation of infection with a number of viruses (reviewed in reference 57). Transfection of homologous RTA proteins into MHV-68-latently infected cells. Data were collected from 2 to 5 independent experiments and are presented as means ± standard errors of the means (SEM). Unless otherwise stated, mice were age and sex matched and infected at 7 to 10 weeks of age with 106 PFU of γHV68 by intraperitoneal injection in 0.5 ml of complete Dulbecco modified Eagle medium (38). Unattached virus was then removed by washing cells with medium containing 2% newborn calf serum (MM). While much is known about the functional properties of the v-cyclins, their role in the pathogenesis of gamma-2 herpesviruses has not been defined. A third interaction is that IFN-I transcriptionally suppresses M2 to inhibit viral reactivation from latency, both directly  and by restoring STAT-1/2 expression to allow STAT-1-dependent transcriptional suppression of ORF50, the main viral lytic transactivator .
At 24 h posttransfection, cells were washed with 1× phosphate-buffered saline (PBS) and subjected to reporter assays. falciparum infection. In ongoing experiments, we try to elucidate the biological mechanisms underlying the observed phenotypes. Thioglycolate inoculation increased the total number of PECs by about 20-fold but did not affect the frequency of cells that reactivate gammaHV68, consistent with a bone marrow reservoir for latent gammaHV68. J Virol 73, 54279–4283. Total RNA was harvested from the lungs of two mice at 1, 3, 5, and 7 days postinfection (dpi). Herpesviruses have also been shown to antagonize IFN responses through numerous mechanisms.
The correct insert orientation was identified by DNA sequencing. The inflammasome is a critical mediator of inflammatory responses that restricts the pathogenesis of numerous viruses and bacteria (11). When the mechanisms of v-cyclin-mediated reactivation are considered, one attractive possibility is that the same functions required for cell cycle manipulation are required to regulate reactivation. The tegument is an electron-dense structure existing between the nucleocapsids and the envelope and constructed with viral- and cellular-encoded proteins and RNAs (26). Examples of human-herpesvirus regulatory proteins that induce PML degradation or PML-NB disruption are herpes simplex virus 1 (HSV-1) ICP0 (13); varicella-zoster virus (VZV) ORF61 (14); human cytomegalovirus (CMV) IE1 (15); Epstein-Barr virus (EBV) proteins EBNA1, Zta, and EBNA-LP (16–18); and Kaposi’s sarcoma-associated herpesvirus (KSHV) LANA2 (19). W. We found that mice previously infected with γHV-68 developed more severe EAE when compared to uninfected EAE mice and showed pathological features that recapitulate human MS.
M. While these viruses are normally effectively contained by the immune system, they can cause a number of clinical problems when patients are immunocompromised. For instance, Epstein-Barr virus (EBV) carries a gene encoding viral interleukin 10 (vIL-10), while MHV68 uses a strategy of upregulating IL-10 via the unique M2 protein to drive B cell proliferation (6, 7). We independently validated the mLANA-Hsc70 interaction through coimmunoprecipitation and in vitro glutathione S-transferase (GST) pulldown assays. Murine gammaherpesvirus 68 (MHV-68) is a member of the gammaherpesvirus subfamily and is closely related to Kaposi’s sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus. We have found that MHV-68 ORF52 encodes an abundant tegument protein conserved among gammaherpesviruses.