Tenotomy decreases reporter protein synthesis via the 3'-untranslated region of the beta-myosin heavy chain mRNA

We tested thehypothesis that the -myosin heavy chain (-MHC) 3'-untranslatedregion (UTR) mediates decreased protein expression after tenotomy ofthe rat soleus. We also tested the hypothesis that decreased proteinexpression is the result of RNA-protein interactions within the 3'-UTR.-MHC was chosen for study because of its critical role in thefunction of postural muscles such as soleus. Adult rat soleus muscleswere directly injected with luciferase (LUC) reporter constructscontaining either the -MHC or SV40 3'-UTR. After 48 h oftenotomy, there was no significant effect on LUC expression in the SV403'-UTR group. In the -MHC 3'-UTR group, LUC expression was 37.3 ± 4% (n = 5, P = 0.03) of that in shamcontrols. Gel mobility shift assays showed that a protein factorspecifically interacts with the -MHC 3'-UTR and that tenotomysignificantly increases the level of this interaction (25 ± 7%,n = 5, P = 0.02). Thus the -MHC3'-UTR is directly involved in decreased protein expression that isprobably due to increased RNA-protein binding within the UTR.

     

Nonhomologous synapsis of the XY during early pachynema in In(X)1H male mice

It has been previously supposed that meiotic synapsis is restricted to homology during early, but not late pachynema. The synaptic begavior of an inverted X chromosome, In(X)1H as reflected in the synaptonemal complexes of the sex chromosomes has been examined in microspread spermatocytes by electron microscopy and evidence of extensive nonhomologus synapsis between the X and Y during early pachynema has been obtained.     

Gamma interferon blocks gammaherpesvirus reactivation from latency in a cell type-specific manner

Gammaherpesviruses are important pathogens whose lifelong survival in the host depends critically on their capacity to establish and reactivate from latency, processes regulated by both viral genes and the host immune response. Previous work has demonstrated that gamma interferon (IFN-gamma) is a key regulator of chronic infection with murine gammaherpesvirus 68 (gammaHV68), a virus that establishes latent infection in B lymphocytes, macrophages, and dendritic cells. In mice deficient in IFN-gamma or the IFN-gamma receptor, gammaHV68 gene expression is altered during chronic infection, and peritoneal cells explanted from these mice reactivate more efficiently ex vivo than cells derived from wild-type mice. Furthermore, treatment with IFN-gamma inhibits reactivation of gammaHV68 from latently infected wild-type peritoneal cells, and depletion of IFN-gamma from wild-type mice increases the efficiency of reactivation of explanted peritoneal cells. These profound effects of IFN-gamma on chronic gammaHV68 latency and reactivation raise the question of which cells respond to IFN-gamma to control chronic gammaHV68 infection. Here, we show that IFN-gamma inhibited reactivation of peritoneal cells and spleen cells harvested from mice lacking B lymphocytes, but not wild-type spleen cells, suggesting that IFN-gamma may inhibit reactivation in a cell type-specific manner. To directly test this hypothesis, we expressed the diphtheria toxin receptor specifically on either B lymphocytes or macrophages and used diphtheria toxin treatment to deplete these specific cells in vivo and in vitro after establishing latency. We demonstrate that macrophages, but not B cells, are responsive to IFN-gamma-mediated suppression of gammaHV68 reactivation. These data indicate that the regulation of gammaherpesvirus latency by IFN-gamma is cell type specific and raise the possibility that cell type-specific immune deficiency may alter latency in distinct and important ways.     

CD40 engagement on dendritic cells, but not on B or T cells, is required for long-term control of murine gammaherpesvirus 68

CD4 T cells are not essential for primary clearance of replicating murine gammaherpesvirus 68 (MHV-68) but are required for effective long-term control. The virus reactivates in the lungs of major histocompatibility complex class II-deficient (CII-/-) mice that lack functional CD4 T cells. CD40 ligand (CD40L) is upregulated on activated CD4 T cells, and it is thought that CD40-CD40L interactions are an important component of CD4 T-cell help. Our previous studies have shown that agonistic antibodies to CD40 can substitute for CD4 T-cell function in the long-term control of MHV-68. In the present study, we sought to identify the CD40-positive cell type mediating this effect. To address this question, we adoptively transferred MHV-68 peptide-pulsed CII(-/-) dendritic cells (DC) that had been treated with an agonistic antibody to CD40 into MHV-68-infected CII(-/-) recipients. Viral reactivation was significantly lower in mice injected with anti-CD40-treated DC than in those injected with control DC or in mice that did not receive any DC. However, in similar experiments with B cells, anti-CD40 treatment had no effect. We also investigated the requirement for CD40 expression on T cells by adoptive transfer of T cells from CD40(+/+) or CD40(-/-) mice into T-cell-deficient recipients that were subsequently infected with MHV-68. The results showed that CD40 expression on T cells is not necessary for preventing viral reactivation. Taken together, our data suggest that CD40 engagement on DC, but not on T or B cells, is essential for effective long-term control of MHV-68.     

Antibody to herpes simplex virus type 2 as serological marker of sexual lifestyle in populations.

OBJECTIVES--To examine the epidemiology of antibody to herpes simplex virus type 2 and to assess its suitability as a serological marker of sexual behaviour in populations with high and low prevalences. DESIGN--Cross sectional survey. SETTING--Department of genitourinary medicine and blood donation centre in central London. SUBJECTS--Representative sample of 869 patients attending department between November 1990 and December 1991, and 1494 consecutive blood donors attending for donation between February and April 1992. METHOD--Participants had a blood sample taken for antibody testing with a novel type specific assay and completed a questionnaire. RESULTS--Prevalence of antibody differed significantly between the two groups (188/833 (22.7%) clinic attenders; 102/1347 (7.6%) blood donors). In both populations antibody was strongly associated with sex, sexual orientation, years of sexual activity, number of lifetime sexual partners, and past infection with sexually transmitted diseases after other factors were controlled for. Only 130 (45%) of all those with antibody had symptoms suggestive of genital herpes, and 79 (27.4%) had had genital herpes diagnosed. Of those without antibody to herpes simplex viruses type 1 and 2, 8.0% reported genital blisters or sores and 1.1% had had genital herpes diagnosed by a doctor. CONCLUSIONS--The strong relation between herpes simplex virus type 2 and sexual lifestyle suggests that the presence of antibody to the virus may be suitable for use as an objective, serological marker of patterns of sexual behaviour in different populations. These data show that only a minority of those infected with herpes simplex virus type 2 have a diagnosis of genital herpes or express clinical symptoms, making serological determinants of infection essential for epidemiological studies.     

Sphingosine Kinase 1 Is Regulated by Peroxisome Proliferator-activated Receptor α in Response to Free Fatty Acids and Is Essential for Skeletal Muscle Interleukin-6 Production and Signaling in Diet-induced Obesity

We previously demonstrated that sphingosine kinase 1 (Sphk1) expression and activity are up-regulated by exogenous palmitate (PAL) in a skeletal muscle model system and in diet-induced obesity in mice; however, potential functions and in vivo relevance of this have not been addressed. Here, we aimed to determine the mechanism by which PAL regulates SphK1 in muscle, and to determine potential roles for its product, sphingosine-1-phosphate (S1P), in muscle biology in the context of obesity. Cloning and analysis of the mouse Sphk1 promoter revealed a peroxisome proliferator-activated receptor (PPAR) α cis-element that mediated activation of a reporter under control of the Sphk1 promoter; direct interaction of PPARα was demonstrated by chromatin immunoprecipitation. PAL treatment induced the proinflammatory cytokine interleukin (IL)-6 in a manner dependent on SphK1, and this was attenuated by inhibition of the sphingosine-1-phosphate receptor 3 (S1PR3)_. Diet-induced obesity in mice demonstrated that IL-6 expression in muscle, but not adipose tissue, increased in obesity, but this was attenuated in Sphk1^−/− mice. Moreover, plasma IL-6 levels were significantly decreased in obese Sphk1^−/− mice relative to obese wild type mice, and muscle, but not adipose tissue IL-6 signaling was activated. These data indicate that PPARα regulates Sphk1 expression in the context of fatty acid oversupply and links PAL to muscle IL-6 production. Moreover, this function of SphK1 in diet-induced obesity suggests a potential role for SphK1 in obesity-associated pathological outcomes.