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Interest in natural remedies has grown recently due to a variety of public health concerns such as microbial antibiotic resistance. This global health concern necessitates innovative approaches to combat bacterial infections. Building upon established therapeutic uses of essential oils, this research focused on the volatile constituents of essential oils. The volatile antimicrobial activity of these constituents was studied by employing a derivative of a modified disk diffusion assay for quantitative comparisons. This study emphasizes the significance and value of exploring natural compounds as alternatives to traditional antibiotics and provides insights into their mechanisms and applications in contending with bacterial pathogens.

Necroptosis mediated by Z-nucleic-acid-binding protein (ZBP)1 (also called DAI or DLM1) contributes to innate host defense against viruses by triggering cell death to eliminate infected cells. During infection, vaccinia virus (VACV) protein E3 prevents death signaling by competing for Z-form RNA through an N-terminal Zα domain. In the absence of this E3 domain, Z-form RNA accumulates during the early phase of VACV infection, triggering ZBP1 to recruit receptor interacting protein kinase (RIPK)3 and execute necroptosis. The C-terminal E3 double-strand RNA-binding domain must be retained to observe accumulation of Z-form RNA and induction of necroptosis. Substitutions of Zα from either ZBP1 or the RNA-editing enzyme double-stranded RNA adenosine deaminase (ADAR)1 yields fully functional E3 capable of suppressing virus-induced necroptosis. Overall, our evidence reveals the importance of Z-form RNA generated during VACV infection as a pathogen-associated molecular pattern (PAMP) unleashing ZBP1/RIPK3/MLKL-dependent necroptosis unless suppressed by viral E3., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: β-lactam antibiotics are a class of broad-spectrum antibiotics consisting of all antibiotic agents that contain a β-lactam ring in their molecular structures. β-lactam antibiotics are only known to be isolated from fungi (e.g. Acremonium chrysogenum, Penicillium chrysogenum and Aspergillus nidulans) and bacteria (e.g. Streptomyces clavuligerus). We have shown that botanical extracts prepared from Larrea tridentata have strong antimicrobial activity against several bacteria, including members of Staphylococcus and Streptococcus genera., Methods: Through resistance studies, inhibitor assays, and ELISA testing, we demonstrated L. tridentata extracts may contain a β-lactam type antibiotic activity., Results: Based on the estimated β-lactam concentration within the extract, the antimicrobial activity of the L. tridentata extract was approximately 2000-8000-fold greater against Staphylococcus as compared to other β-lactams, penicillin or ampicillin. In the L. tridentata extract, this increased activity was found to be associated with the likely presence of a cofactor leading to increased potentiation of the β-lactam activity. This potentiation activity was also observed to enhance the activity of exogenously added natural penicillin antibiotics., Conclusions: Although constituents were not isolated in this study, the results obtained strongly support the presence of β-lactam type antibiotic activity and antibiotic potentiation activity present in ethanolic extracts prepared from L. tridentata.

Background: Essential oils are volatile and lipophilic liquid extracts made from plants as secondary metabolites that can be obtained by distillation. To date, several studies have investigated the direct antimicrobial activity of liquid essential oils. However, this study investigated the antimicrobial properties of the volatile substances present in various essential oils., Methods: A modified zone of inhibition protocol was developed using agar petri dishes with a center glass vial to allow evaporation and aerosolization of the potential active constituents from essential oils. In total, nineteen essential oils were tested against five Gram positive bacterial species, five Gram negative bacterial species and one fungi., Results: This study found potent antimicrobial activity from the volatile constituents of several essential oils. Rosemary, tea tree, and cassia volatiles were found to be the best broad-spectrum antibacterial agents, whereas clove volatiles had almost no antimicrobial activity., Conclusions: These results support the anecdotal historical evidence of the antimicrobial activity of the volatile constituents essential oils. Modern medical implications for this work may be related to the use of aromatic essential oils for respiratory or dermatological infections.

Herpes simplex virus type-1 (HSV-1), one of the most widely spread human viruses in the Herpesviridae family, causes herpes labialis (cold sores) and keratitis (inflammation of the cornea). Conventional treatment for HSV-1 infection includes pharmaceutical drugs, such as acyclovir and docosonal, which are efficacious but maintain the potential for the development of viral drug resistance. Extracts from the carnivorous pitcher plant, Sarracenia purpurea, have previously been shown to inhibit the replication of HSV-1. In this study, we demonstrate that S. purpurea extracts can inhibit the replication of HSV-1 by two distinct mechanisms of action. These extracts directly inhibit extracellular virions or viral attachment to the human host cell as well as inhibiting the expression of viral immediate-early, early and late genes when added at various times post-infection. This botanical has previously been shown to inhibit the replication of poxviruses through the inhibition of early viral gene transcription. These results support a broader anti-viral activity of S. purpurea extracts against both pox and herpes viruses.

Astragalus membranaceus is a staple of Traditional Chinese Medicine being one of the oldest medicinal herbs listed in the material medica of Chinese herbal medicine. Chinese herbalists have used Astragalus to help the human body fight a variety of diseases. Modern herbalists utilize Astragalus primarily as an immunostimulant to prevent common infection and aid in the recovery following infection. Historically, the biological activities associated with Astragalus have been accounted for, at least in part, to several constituents present in the botanical including saponins and polysaccharides. We propose that in addition to these constituents, compounds from endophytic (or epiphytic) bacteria present in (or on) the roots of Astragalus may have an important biological role. Lipopolysaccharides and lipoproteins are major components of Gram-negative bacteria and highly potent activators of the innate immune response. Our data supports a direct correlation between the level of immune gene induction and the level of lipopolysaccharides/lipoproteins present in the Astragalus extract. We demonstrate that extracts from Astragalus specifically activate Toll-like and NOD-like receptors involved in the recognition and response to bacterial constituents and that removal of the lipopolysaccharide/lipoprotein from the Astragalus extract reduced the level of this response. The results support that many immune enhancing botanicals have established a symbiotic relationship with Gram-negative bacteria and that the immune enhancing effect of these botanical extracts on the body may not only be due to endogenous plant compounds, but endophytic (or epiphytic) bacterial components as well., (Copyright ©2020, Yale Journal of Biology and Medicine.)

Staphylococcus aureus is an opportunistic pathogen. Over- and improper-use of pharmaceuticals against S. aureus has led to the development of antibiotic resistance, including methicillin-resistant S. aureus (MRSA). This study examined the efficacy of botanical extracts as an alternative form of treatment to S. aureus and MRSA, including penicillin/methicillin-resistant S. aureus (Pen R ), and multidrug resistant S. aureus (MDR). Initial screening of botanicals was done via a minimum inhibitory concentration procedure. In addition, a temporal growth curve was performed in order to quantify the growth of the bacteria in the presence of the extracts. Results demonstrated 13 botanicals that had varying activities against S. aureus, Pen R , and MDR. These botanicals were separated into mild, moderate, and highly efficacious based on the concentration needed to inhibit bacterial growth. These results propose a comparison of botanical-derived antimicrobial extracts that may be utilized against S. aureus and different antibiotic resistant strains of MRSA., (© 2018 John Wiley & Sons, Ltd.)

Organic compounds with a caffeoyl moiety (e.g. caffeic acid, rosmarinic acid, chicoric acid, etc.) have antiviral properties towards herpes simplex (HSV), influenza and immunodeficiency viruses (HIV). This study evaluated the HSV antiviral properties of caffeic acid when paired with a variety of metal and other inorganic ions. The results demonstrated that the antiviral activity of caffeic acid increased upwards of 100-fold by the addition of cations, such as Fe 3+ , and anionic molecules, such as molybdate and phosphate. Cellular toxicity tests of the caffeic acid chelates showed that they have low toxicities with selectivity indices (TD 50 /EC 50 ) for Fe 3+ , MoO 4 2- , and PO 4 3- chelates being 1700, >540, and >30, respectively. Caffeic acid paired with Fe 3+ was tested against eight strains of viruses, including those from different families. The caffeic acid chelates were mostly effective against HSV1 and HSV2, but they also had moderate activity against vaccinia virus and a VSV-Ebola pseudotyped virus. All the viruses that were strongly impacted by the caffeic chelates require heparan sulfate proteoglycans for cellular attachment, so it is likely that caffeic chelates target and interfere with this mechanism. Since the caffeic acid chelates target an extra-cellular process, they might be able to be combined with existing medications, such as acyclovir, that target an intracellular process to achieve greater viral control., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The classic presentation of acute gastrointestinal (GI) parasitic infection including fever, diarrhea, nausea, and/or abdominal pain is not always a reliable measure. In these cases, parasitic infections require sensitive stool testing to be identified and treated effectively. The standard practice for stool analysis typically requires 3 separate stool specimens to be collected on 3 separate days to overcome the possibility of misdiagnosis due to intermittent shedding. Nonetheless, this practice is not often utilized due to very short periods of sample viability, which often leads to patient misdiagnosis. In this study, a patient with a history that eluded to possible acute GI parasitic infection, but without classic symptomology, was screened using a sodium acetate acetic acid formalin (SAF)-fixative triple-faeces test. Even though the patient's symptoms did not strongly suggest stool analysis, her history of travel was clinically relevant. The faeces test revealed the patient was positive for Blastocystis hominis, Dientamoeba fragilis trophs, Endolimax nana trophs, Entamoeba coli trophs, a-hemolytic Streptococcus, Enterobacter cloacae, Klebsiella pneumoniae, and yeast. Subsequently, the patient was treated for GI infection and received significant symptom relief. This case supports the concept that physicians should fully evaluate a patient's clinical presentation and symptomology, including history and environmental exposures, and include highly sensitive testing for proper evaluation of potential GI parasitic infections.

Context • Research on the schizophrenia spectrum is primarily focused on pharmaceutical interventions, although alternative treatments have been gaining increasing popularity in recent years because patients are seeking treatments that are effective and have reduced side effects. A significant body of evidence already exists supporting the effectiveness of homeopathy to treat a wide array of illnesses. Objective • The research team intended to demonstrate the need for using both alternative and conventional treatments to improve clinical outcomes in the treatment of schizoaffective disorder. Design • The research team performed 2 case studies. Setting • The study took place at Arizona Natural Health Center (Tempe, AZ, USA), an outpatient clinic where Dr Tara Peyman worked as a naturopathic doctor from 2008 to 2014. Participants • The participants were a 23-y-old female (case 1) and a 34-y-old female (case 2), both of whom had been diagnosed with schizoaffective disorder of the bipolar type. Intervention • Individualized homeopathic treatment was initiated for the 2 patients, who previously had received medication of atypical antipsychotics and mood stabilizers. Outcome Measures • A Likert scale was used to evaluate the intensity of each patient's symptoms at each follow-up, based on self-reporting, using a scale from 1 to 10, with a score of 10 being the highest. Results • During the course of treatment, both patients' symptoms normalized, and they regained their ability to hold jobs, attend school, and maintain healthy relationships with their families and partners while requiring fewer pharmaceutical interventions. Conclusions • The 2 current case reports demonstrate a successful integrative approach to the treatment of schizoaffective disorder. They illustrate the value of individualized homeopathic prescriptions with proper case management in the successful treatment of that disorder. Future large-scale, double-blind, placebo-controlled studies should investigate individualized homeopathic treatments for mental health concerns, because the diseases cause great economic and social burden.

Vaccinia virus (VACV) encodes an innate immune evasion protein, E3, which contains an N-terminal Z-nucleic acid binding (Zα) domain that is critical for pathogenicity in mice. Here we demonstrate that the N terminus of E3 is necessary to inhibit an IFN-primed virus-induced necroptosis. VACV deleted of the Zα domain of E3 (VACV-E3LΔ83N) induced rapid RIPK3-dependent cell death in IFN-treated L929 cells. Cell death was inhibited by the RIPK3 inhibitor, GSK872, and infection with this mutant virus led to phosphorylation and aggregation of MLKL, the executioner of necroptosis. In 293T cells, induction of necroptosis depended on expression of RIPK3 as well as the host-encoded Zα domain-containing DNA sensor, DAI. VACV-E3LΔ83N is attenuated in vivo, and pathogenicity was restored in either RIPK3- or DAI-deficient mice. These data demonstrate that the N terminus of the VACV E3 protein prevents DAI-mediated induction of necroptosis., Competing Interests: The authors declare no conflict of interest.

Context • Small intestinal bacterial overgrowth (SIBO) is commonly defined as an increased number of bacteria and/or an abnormal type of bacteria in the small intestine. Conventional treatment for SIBO is typically focused on antibiotics to eradicate the bacterial overgrowth. Numerous studies have demonstrated the antimicrobial activity of herbs, and a diet low in fermentable oligo-, di-, and monosaccharides and polyols (FODMAPs) has been shown to enhance antibiotic therapy. Objective • The current case study intended to evaluate the benefits of an alternative, multifaceted approach-including botanical and homeopathic therapies in conjunction with a low-FODMAP diet-in the treatment of SIBO and its associated symptoms. Design • The research team performed a case study. Setting • The study was conducted at SCNM Medical Center (Tempe, AZ, USA). Participant • The participant was a female patient at the SCNM Medical Center with chronic, daily, severe abdominal bloating and pain that particularly worsened after meals and by the end of the day. The patient also had a significant history of chronic constipation that had begun approximately 10 y prior to her experiencing the daily abdominal pain. Intervention • Based on a lactulose breath test for hydrogen and methane, the research team diagnosed the patient with a case of mild SIBO. The treatment approach was multifaceted, involving a low-FODMAP diet, antimicrobial botanical therapy, and homeopathic medicine. Results • The patient's abdominal pain and bloating resolved with the treatment of the SIBO, although her underlying constipation, which was likely associated with other factors, remained. Conclusions • This case study supports an alternative, multifaceted approach to the treatment of SIBO and commonly associated symptoms.

Context • Allergies are a common affliction, whether they are respiratory, food related, or dermatological. People often resort to continuous use of over-the-counter medications, such as antihistamines, to manage their symptoms. Controversy still remains over testing serum immunoglobulin (Ig) G to diagnose food allergies. Objective • This study intended to examine the benefits of treatment of a pediatric patient with natural supplements and an elimination diet for IgG food allergies. Design • The research team reported a case study. Setting • The study was conducted at Southwest Naturopathic Medical Center (Tempe, AZ, USA). Participant • The participant was a 10-y-old Caucasian female who had diagnoses of allergic rhinitis and reactive bronchospasm, the second of which was exacerbated by allergens such as wheat, perfumes, and seasonal flora. Intervention • Following testing for IgE- and IgG-reactive foods, the patient was treated with natural supplements to reduce her allergic responses and was instructed to make dietary changes to eliminate the IgG-reactive foods. Outcome Measures • The patient's symptom severity was tracked starting 1 mo after her initial visit to Southwest Naturopathic Medical Center. The severity was based on the patient's subjective reports about her congestion to her mother and on her mother's observations of the effect of symptoms on her attention and school performance. The bronchospasm severity was based on the frequency of a sensation of wheezing and chest tightness, the frequency of inhaler use, and the occurrence of any exacerbation of symptoms with acute respiratory illness Results • After 1 mo, in which the patient used the natural supplements, she experienced a 90% improvement in coughing; a 70% improvement in nasal congestion; less chest tightness; and no need for use of loratadine, diphenhydramine, or albuterol. At the 8-mo follow-up visit, her nasal congestion was reported to be entirely gone. Conclusions • The case demonstrates the effectiveness of natural supplements and a diet eliminating IgG-reactive foods in the treatment and management of pediatric allergic rhinitis and reactive bronchospasm.

Monkeypox virus (MPXV) infection fails to activate the host anti-viral protein, PKR, despite lacking a full-length homologue of the vaccinia virus (VACV) PKR inhibitor, E3. Since PKR can be activated by dsRNA produced during a viral infection, we have analyzed the accumulation of dsRNA in MPXV-infected cells. MPXV infection led to less accumulation of dsRNA than VACV infection. Because in VACV infections accumulation of abnormally low amounts of dsRNA is associated with mutations that lead to resistance to the anti-poxvirus drug isatin beta-thiosemicarbazone (IBT), we investigated the effects of treatment of MPXV-infected cells with IBT. MPXV infection was eight-fold more resistant to IBT than wild-type vaccinia virus (wtVACV). These results demonstrate that MPXV infection leads to the accumulation of less dsRNA than wtVACV, which in turn likely leads to a decreased capacity for activation of the dsRNA-dependent host enzyme, PKR., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The botanical, Astragalus membranaceus, is a therapeutic in traditional Chinese medicine. Limited literature exists on the overall in vivo effects of A. membranaceus on the human body. This study evaluates the physiological responses to A. membranaceus by measuring leukocyte, platelet, and cytokine responses as well as body temperature and blood pressure in healthy individuals after the in vivo administration of A. membranaceus. A dose-dependent increase in monocytes, neutrophils, and lymphocytes was measured 8-12 hours after administration and an increase in the number of circulating platelets was seen as early as 4 hours. A dynamic change in the levels of circulating cytokines was observed, especially in interferon-γ and tumor necrosis factor-α, IL-13, IL-6, and soluble IL-2R. Subjective symptoms reported by participants were similar to those typically experienced in viral type immune responses and included fatigue, malaise, and headache. Systolic and diastolic blood pressure were reduced within 4 hours after administration, while body temperature mildly increased within 8 hours after administration. In general, all responses returned to baseline values by 24 hours. Collectively, these results support the role of A. membranaceus in priming for a potential immune response as well as its effect on blood flow and wound healing.

Background: Resurgences of Staphylococcus aureus infection continue globally, with antibiotic resistance increasing dramatically, making these infections more difficult to treat. S. aureus epidemics impose public health threats, and economic burdens on health care costs worldwide, presenting challenges modern medicine struggles to control., Objective: In order to answer today's call for effective treatments against S. aureus, we evaluated and compared various botanical extracts that have historically been suggested as useful for their antimicrobial properties against S. aureus., Design: Briefly, S. aureus cultures were treated with selected botanical extracts and the minimum inhibitory concentration (MIC) determined. In addition, to obtain more quantitative measures on bacterial growth, 24-hour growth studies were done to examine the temporal activity and stability of various botanicals on bacterial replication., Results: The antimicrobial activity observed for the botanical extracts used in this comparative evaluation of efficacy included both bacteriostatic and bacteriocidal activity against S. aureus. Highly effective botanicals including Salvia officinalis, Eucalyptus globulus, Coleus forskohlii, Coptis chinensis, Turnera diffusa, and Larrea tridentata exhibited MIC values ranging from 60 to 300 μg/mL and a 10(6)-fold reduction in bacterial replication. Arctostaphylos uva-ursi and Allium sativum were slightly less effective, exhibiting MIC values ranging from 90 to 400 μg/mL and a 10(5)-fold reduction, while Anemopsis californica gave MIC value of 360 μg/mL and a 10(4)-fold reduction in bacterial replication. Many botanicals, especially at lower doses, had an initial inhibitory effect followed by a recovery in bacterial replication. Such botanicals included E. globulus, C. chinensis, T. diffusa, A. californica, and Berberis vulgaris., Conclusions: Our data demonstrate that S. officinalis, E. globulus, C. forskohlii, A. uva-ursi, C. chinensis, T. diffusa, A. californica, A. sativum, and L. tridentata all show promising direct antimicrobial activity against S. aureus. For many of these botanicals, strong bacteriocidal activity was observed at higher concentrations, but even at lower concentrations, bacteriostatic activity was evident. Other botanicals including B. vulgaris, Baptisia tinctoria, and Glycyrrhiza glabra showed moderate activity against S. aureus, while Schisandra chinensis, Echinacea angustifolia, and Polygonum multiflorum were shown to be ineffective.

Objectives: Silver has historically and extensively been used as a broad-spectrum antimicrobial agent. However, the Food and Drug Administration currently does not recognize colloidal silver as a safe and effective antimicrobial agent. The goal of this study was to further evaluate the antimicrobial efficacy of colloidal silver., Design: Several strains of bacteria, fungi, and viruses were grown under multicycle growth conditions in the presence or absence of ionic colloidal silver in order to assess the antimicrobial activity., Results: For bacteria grown under aerobic or anaerobic conditions, significant growth inhibition was observed, although multiple treatments were typically required. For fungal cultures, the effects of ionic colloidal silver varied significantly between different genera. No viral growth inhibition was observed with any strains tested., Conclusions: The study data support ionic colloidal silver as a broad-spectrum antimicrobial agent against aerobic and anaerobic bacteria, while having a more limited and specific spectrum of activity against fungi.

The NS5A nonstructural protein of hepatitis C virus (HCV) has been shown to inhibit the cellular interferon (IFN)-induced protein kinase R (PKR). PKR mediates the host IFN-induced antiviral response at least in part by inhibiting mRNA translation initiation through phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha). We thus examined the effect of NS5A inhibition of PKR on mRNA translation within the context of virus infection by using a recombinant vaccinia virus (VV)-based assay. The VV E3L protein is a potent inhibitor of PKR. Accordingly, infection of IFN-pretreated HeLa S3 cells with an E3L-deficient VV (VVDeltaE3L) resulted in increased phosphorylation levels of both PKR and eIF2alpha. IFN-pretreated cells infected with VV in which the E3L locus was replaced with the NS5A gene (VVNS5A) displayed diminished phosphorylation of PKR and eIF2alpha in a transient manner. We also observed an increase in activation of p38 mitogen-activated protein kinase in IFN-pretreated cells infected with VVDeltaE3L, consistent with reports that p38 lies downstream of the PKR pathway. Furthermore, these cells exhibited increased phosphorylation of the cap-binding initiation factor 4E (eIF4E), which is downstream of the p38 pathway. Importantly, these effects were reduced in cells infected with VVNS5A. NS5A was also found to inhibit activation of the p38-eIF4E pathway in epidermal growth factor-treated cells stably expressing NS5A. NS5A-induced inhibition of eIF2alpha and eIF4E phosphorylation may exert counteracting effects on mRNA translation. Indeed, IFN-pretreated cells infected with VVNS5A exhibited a partial and transient restoration of cellular and viral mRNA translation compared with IFN-pretreated cells infected with VVDeltaE3L. Taken together, these results support the role of NS5A as a PKR inhibitor and suggest a potential mechanism by which HCV might maintain global mRNA translation rate during early virus infection while favoring cap-independent translation of HCV mRNA during late infection.

NFAT transcription factors play a central role in initiating T-cell activation through the induction of immediate-early T-cell specific genes including interleukin-2 (IL-2). NFAT transcription factors bind to a sequence in the IL-2 enhancer known as the antigen receptor response element 2 (ARRE-2). Multiple proteins exhibiting ARRE-2 binding activity have been isolated, including a heterodimer from stimulated T-cell nuclear extracts consisting of Mr = 90 000 (NF90) and Mr = 45 000 (NF45) subunits. The subunits of this heterodimer have been cloned, and NF90 was found to encode a protein containing two domains that are predicted to form motifs capable of binding to double-stranded RNA. Using in vitro translated polypeptides, we have demonstrated that NF90 specifically binds to double-stranded RNA. Furthermore, NF90 was phosphorylated in a double-stranded RNA-dependent manner likely by the interferon-induced, double-stranded RNA-dependent protein kinase, PKR. The NF90 protein was found to be expressed not only in T-cells, but also in nonimmune HeLa cells. In HeLa cells, the protein was almost exclusively localized to the ribosome salt wash fraction of cell lysates.

Several viruses have been shown to code for proteins that specifically bind to double-stranded RNA or RNA with large amounts of secondary structure. These proteins have been implicated in providing interferon resistance to viruses and in inhibiting induction of apoptosis by viruses, and have been suggested to be involved in regulation of viral and cellular protein synthesis in infected cells. This article describes methods for detecting and analyzing viral double-stranded RNA-binding proteins., (Copyright 1998 Academic Press.)

An epidemiologic association between viral infections and the onset of asthma and allergy has been documented. Also, evidence from animal and human studies has suggested an increase in antigen-specific immunoglobulin E (IgE) production during viral infections, and elevated levels of IgE are characteristic of human asthma and allergy. Here, we provide molecular evidence for the roles of viral infection and of activation of the antiviral protein kinase (PKR) (double-stranded-RNA [dsRNA]-activated protein kinase) in the induction of IgE class switching. The presence of dsRNA, a known component of viral infection and an activator of PKR, induced IgE class switching as detected by the expression of germ line epsilon in the human Ramos B-cell line. Furthermore, dsRNA treatment of Ramos cells resulted in the activation of PKR and in vivo activation of the NF-kappaB complex. Interestingly, infection of Ramos cells with rhinovirus (common cold virus) serotypes 14 and 16 resulted in the induction of germ line epsilon expression. To further evaluate the role of PKR in the viral induction of IgE class switching, we infected Ramos cells with two different vaccinia virus (cowpox virus) strains. Infection with wild-type vaccinia virus failed to induce germ line epsilon expression; however, a deletion mutant of vaccinia virus (VP1080) lacking the PKR-inhibitory polypeptide E3L induced the expression of germ line epsilon. Collectively, the results of our study define a common molecular mechanism underlying the role of viral infections in IgE class switching and subsequent induction of IgE-mediated disorders such as allergy and asthma.

An inhibitor of eIF-2a phosphorylation was identified in various plant species. The plant protein (termed PKI) specifically cross-reacts with monoclonal antiserum that recognizes the glycosylated, active form of a M(r) 87 kD protein analog (p67) from reticulocytes. Northern blot analysis using a probe to the reticulocyte inhibitor cDNA further supports the presence of analogous transcripts in plant tissue. PKI specifically inhibits the phosphorylation of the plant encoded eIF-2 alpha kinase (pPKR) as well as plant and human eIF-2 alpha phosphorylation. The interaction between PKI and pPKR is indicated by their copurification on dsRNA agarose, despite evidence showing that PKI does not bind dsRNA. Further, wheat PKI inhibits human PKR phosphorylation but activity is recovered by immuno-depletion of PKI from wheat germ fractions. PKI is temporally regulated during plant growth and development. It is maximally present in extracts from dormant seeds, however, it is not detectable soon after leaf emergence at approximately 48 h post-imbibition. PKI levels are again detectable at the mid-milk stage in seed development. Protein levels of pPKR in ribosomal salt wash and cytosolic extracts from healthy plant tissue remain essentially constant throughout the life cycle. In contrast, pPKR activity levels based upon autophosphorylation vary significantly and are inversely correlated with PKI protein levels. Phosphorylation of eIF-2 alpha is a classical mechanism for the downregulation of protein synthesis suggesting that inhibition of pPKR activity by PKI may contribute to the dramatic and rapid increase in protein synthesis observed during seed germination.