Identification of gammaherpesvirus infection in free-ranging black bears (Ursus americanus).

Herpesvirus infection was investigated in black bears (Ursus americanus) with neurological signs and brain lesions of nonsuppurative encephalitis of unknown cause. Visible cytopathic effects (CPE) could only be observed on days 3-5 post-infection in HrT-18G cell line inoculated with bear tissue extracts. The observed CPE in HrT-18G cells included syncytia, intranuclear inclusions, and cell detachments seen in herpesvirus infection in vitro. Herpesvirus-like particles were observed in viral culture supernatant under the electron microscope, however, capsids ranging from 60 nm to 100 nm in size were often observed in viral cultures within the first two passages of propagation. Herpesvirus infection in the bear tissues and tissue cultures were detected by PCR using degenerate primers specific to the DNA polymerase gene (DPOL) and glycoprotein B gene (gB). DNA sequencing of the amplicon revealed that the detected herpesvirus has 94-95% identity to Ursid gammaherpesvirus 1 (UrHV-1) DNA sequences of DPOL. Phylogenetic analysis of DPOL sequences indicates that black bear herpesviruses and UrHV-1 are closely related and have small distances to members of Rhadinovirus. Interestingly, black bear herpesvirus infections were also found in bears without neurological signs. The DPOL DNA sequence of black bear herpesviruses detected in neurological bears were similar to the those detected in the non-neurological bears. However, the gB DNA sequence detected from the neurological bear is different from non-neurological bear and has only 64.5%-70% identity to each other. It is possible that at least two different types of gammaherpesviruses are present in the U. americanus population or several gammaherpesviruses exist in ursine species.

The Molluscum Contagiosum Virus protein MC163 localizes to the mitochondria and dampens mitochondrial mediated apoptotic responses.

Apoptosis is a powerful host cell defense to prevent viruses from completing replication. Poxviruses have evolved complex means to dampen cellular apoptotic responses. The poxvirus, Molluscum Contagiosum Virus (MCV), encodes numerous host interacting molecules predicted to antagonize immune responses. However, the function of the majority of these MCV products has not been characterized. Here, we show that the MCV MC163 protein localized to the mitochondria via an N-terminal mitochondrial localization sequence and transmembrane domain. Transient expression of the MC163 protein prevented mitochondrial membrane permeabilization (MMP), an event central to cellular apoptotic responses, induced by either Tumor Necrosis Factor alpha (TNF-α) or carbonyl cyanide 3-chlorophenylhydrazone (CCCP). MC163 expression prevented the release of a mitochondrial intermembrane space reporter protein when cells were challenged with TNF-α. Inhibition of MMP was also observed in cell lines stably expressing MC163. MC163 expression may contribute to the persistence of MCV lesions by dampening cellular apoptotic responses.

Gain- and loss-of-function mutations in Zat10 enhance the tolerance of plants to abiotic stress.

C(2)H(2)-zinc finger proteins that contain the EAR repressor domain are thought to play a key role in modulating the defense response of plants to abiotic stress. Constitutive expression of the C(2)H(2)-EAR zinc finger protein Zat10 in Arabidopsis was found to elevate the expression of reactive oxygen-defense transcripts and to enhance the tolerance of plants to salinity, heat and osmotic stress. Surprisingly, knockout and RNAi mutants of Zat10 were also more tolerant to osmotic and salinity stress. Our results suggest that Zat10 plays a key role as both a positive and a negative regulator of plant defenses.

The zinc-finger protein Zat12 plays a central role in reactive oxygen and abiotic stress signaling in Arabidopsis.

Plant acclimation to environmental stress is controlled by a complex network of regulatory genes that compose distinct stress-response regulons. In contrast to many signaling and regulatory genes that are stress specific, the zinc-finger protein Zat12 responds to a large number of biotic and abiotic stresses. Zat12 is thought to be involved in cold and oxidative stress signaling in Arabidopsis (Arabidopsis thaliana); however, its mode of action and regulation are largely unknown. Using a fusion between the Zat12 promoter and the reporter gene luciferase, we demonstrate that Zat12 expression is activated at the transcriptional level during different abiotic stresses and in response to a wound-induced systemic signal. Using Zat12 gain- and loss-of-function lines, we assign a function for Zat12 during oxidative, osmotic, salinity, high light, and heat stresses. Transcriptional profiling of Zat12-overexpressing plants and wild-type plants subjected to H(2)O(2) stress revealed that constitutive expression of Zat12 in Arabidopsis results in the enhanced expression of oxidative- and light stress-response transcripts. Under specific growth conditions, Zat12 may therefore regulate a collection of transcripts involved in the response of Arabidopsis to high light and oxidative stress. Our results suggest that Zat12 plays a central role in reactive oxygen and abiotic stress signaling in Arabidopsis.

Cytosolic ascorbate peroxidase 1 is a central component of the reactive oxygen gene network of Arabidopsis.

Reactive oxygen species (ROS), such as O2- and H2O2, play a key role in plant metabolism, cellular signaling, and defense. In leaf cells, the chloroplast is considered to be a focal point of ROS metabolism. It is a major producer of O2- and H2O2 during photosynthesis, and it contains a large array of ROS-scavenging mechanisms that have been extensively studied. By contrast, the function of the cytosolic ROS-scavenging mechanisms of leaf cells is largely unknown. In this study, we demonstrate that in the absence of the cytosolic H2O2-scavenging enzyme ascorbate peroxidase 1 (APX1), the entire chloroplastic H2O2-scavenging system of Arabidopsis thaliana collapses, H2O2 levels increase, and protein oxidation occurs. We further identify specific proteins oxidized in APX1-deficient plants and characterize the signaling events that ensue in knockout-Apx1 plants in response to a moderate level of light stress. Using a dominant-negative approach, we demonstrate that heat shock transcription factors play a central role in the early sensing of H2O2 stress in plants. Using knockout plants for the NADPH oxidase D protein (knockout-RbohD), we demonstrate that RbohD might be required for ROS signal amplification during light stress. Our study points to a key role for the cytosol in protecting the chloroplast during light stress and provides evidence for cross-compartment protection of thylakoid and stromal/mitochondrial APXs by cytosolic APX1.