Bovine viral diarrhea virus: prevention of persistent fetal infection by a combination of two mutations affecting Erns RNase and Npro protease

Different genetically engineered mutants of bovine viral diarrhea virus (BVDV) were analyzed for the ability to establish infection in the fetuses of pregnant heifers. The virus mutants exhibited either a deletion of the overwhelming part of the genomic region coding for the N-terminal protease N(pro), a deletion of codon 349, which abrogates the RNase activity of the structural glycoprotein E(rns), or a combination of both mutations. Two months after infection of pregnant cattle with wild-type virus or either of the single mutants, the majority of the fetuses contained virus or were aborted or found dead in the uterus. In contrast, the double mutant was not recovered from fetal tissues after a similar challenge, and no dead fetuses were found. This result was verified with a nonrelated BVDV containing similar mutations. After intrauterine challenge with wild-type virus, mutated viruses, and cytopathogenic BVDV, all viruses could be detected in fetal tissue after 5, 7, and 14 days. Type 1 interferon (IFN) could be detected in fetal serum after challenge, except with wild-type noncytopathogenic BVDV. On days 7 and 14 after challenge, the largest quantities of IFN in fetal serum were induced by the N(pro) and RNase-negative double mutant virus. The longer duration of fetal infection with the double mutant resulted in abortion. Therefore, for the first time, we have demonstrated the essential role of both N(pro) and E(rns) RNase in blocking interferon induction and establishing persistent infection by a pestivirus in the natural host.     

A horizontally transferred nuclear gene is associated with microhabitat variation in a natural plant population

Horizontal gene transfer involves the non-sexual interspecific transmission of genetic material. Even if they are initially functional, horizontally transferred genes are expected to deteriorate into non-expressed pseudogenes, unless they become adaptively relevant in the recipient organism. However, little is known about the distributions of natural transgenes within wild species or the adaptive significance of natural transgenes within wild populations. Here, we examine the distribution of a natural plant-to-plant nuclear transgene in relation to environmental variation within a wild population. Festuca ovina is polymorphic for an extra (second) expressed copy of the nuclear gene (PgiC) encoding cytosolic phosphoglucose isomerase, with the extra PgiC locus having been acquired horizontally from the distantly related grass genus Poa. We investigated variation at PgiC in samples of F. ovina from a fine-scale, repeating patchwork of grassland microhabitats, replicated within spatially separated sites. Even after accounting for spatial effects, the distributions of F. ovina individuals carrying the additional PgiC locus, and one of the enzyme products encoded by the locus, are significantly associated with fine-scale habitat variation. Our results suggest that the PgiC transgene contributes, together with the unlinked ‘native’ PgiC locus, to local adaptation to a fine-scale mosaic of edaphic and biotic grassland microhabitats.     

An integrative approach combining ion mobility mass spectrometry,X-ray crystallography,and nuclear magnetic resonance spectroscopy to study the conformational dynamics of α1-antitrypsin upon ligand binding

Native mass spectrometry (MS) methods permit the study of multiple protein species within solution equilibria, whereas ion mobility (IM)-MS can report on conformational behavior of specific states. We used IM-MS to study a conformationally labile protein (α₁-antitrypsin) that undergoes pathological polymerization in the context of point mutations. The folded, native state of the Z-variant remains highly polymerogenic in physiological conditions despite only minor thermodynamic destabilization relative to the wild-type variant. Various data implicate kinetic instability (conformational lability within a native state ensemble) as the basis of Z α₁-antitrypsin polymerogenicity. We show the ability of IM-MS to track such disease-relevant conformational behavior in detail by studying the effects of peptide binding on α₁-antitrypsin conformation and dynamics. IM-MS is, therefore, an ideal platform for the screening of compounds that result in therapeutically beneficial kinetic stabilization of native α₁-antitrypsin. Our findings are confirmed with high-resolution X-ray crystallographic and nuclear magnetic resonance spectroscopic studies of the same event, which together dissect structural changes from dynamic effects caused by peptide binding at a residue-specific level. IM-MS methods, therefore, have great potential for further study of biologically relevant thermodynamic and kinetic instability of proteins and provide rapid and multidimensional characterization of ligand interactions of therapeutic interest.PDB Code(s): 4PYW     

Relationships among fire frequency, rainfall and vegetation patterns in the wet–dry tropics of northern Australia: an analysis based on NOAA-AVHRR data

Aim To quantify the regional‐scale spatio‐temporal relationships among rainfall, vegetation and fire frequency in the Australian wet–dry tropics (AWDT). Location Northern Australia: Cape York Peninsula, central Arnhem, central Kimberly, Einasleigh Uplands, Gulf Fall Uplands and northern Kimberley. Methods Monthly ‘fraction of photosynthetic active radiation absorbed by green vegetation’ (fAPAR) was decomposed into monthly evergreen (EG) and monthly raingreen (RG) components using time‐series techniques applied to monthly normalized difference vegetation index (NDVI) data from Advanced Very High Resolution Radiometer (AVHRR) imagery. Fire affected areas were independently mapped at the same spatio‐temporal resolution from AVHRR imagery. Weather station records were spatially interpolated to create monthly rainfall surfaces. Vegetation structural classes were derived from a digitized map of northern Australian vegetation communities (1 : 1,000,000). Generalized linear models were used to quantify relationships among the fAPAR, EG and RG signals, vegetation structure, rainfall and fire frequency, for the period November 1996–December 2001. Results The fAPAR and EG signals are positively correlated with annual rainfall and canopy cover, notably: EG_{closed forest} > EG_{open heathland} > EG_{open forest} > EG_woodland > EG_{open woodland} > EG_{low woodland} > EG_{low open woodland} > EG_{open grassland}. Vegetation height and fAPAR are positively correlated, excluding the special case of open heathland. The RG signal is highest where intermediate annual rainfall and strong seasonality in rainfall coincide, and is associated with vegetation structure as follows: RG_{open grassland} > RG_woodland > RG_{open forest} > RG_{open heathland} > RG_{low woodland} > RG_{open woodland} > RG_{low open woodland} > RG_{closed forest}. Monthly RG tracks monthly rainfall. Annual proportion of area burnt (PB) is maximal where high RG coincides with low EG (open grassland, several woodland communities). PB is minimal in vegetation where both RG and EG are low (low open woodland); and in vegetation where EG is high (closed forest, open heathland). Conclusions The RG–EG scheme successfully reflects digitally mapped tree and grass covers in relation to rainfall. RG–EG patterns are strongly associated with fire frequency patterns. PB is maximal in areas of high RG, where high biomass production during the wet season supports abundant fine fuel during the dry season. PB is minimal in areas with high EG, where relatively moist fuel limits fire ignition; and in areas with low EG and RG, where a relative short supply of fuel limits fire spread.     

Coronavirus replication complex formation utilizes components of cellular autophagy

The coronavirus mouse hepatitis virus (MHV) performs RNA replication on double membrane vesicles (DMVs) in the cytoplasm of the host cell. However, the mechanism by which these DMVs form has not been determined. Using genetic, biochemical, and cell imaging approaches, the role of autophagy in DMV formation and MHV replication was investigated. The results demonstrated that replication complexes co-localize with the autophagy proteins, microtubule-associated protein light-chain 3 and Apg12. MHV infection induces autophagy by a mechanism that is resistant to 3-methyladenine inhibition. MHV replication is impaired in autophagy knockout, APG5-/-, embryonic stem cell lines, but wild-type levels of MHV replication are restored by expression of Apg5 in the APG5-/-cells. In MHV-infected APG5-/-cells, DMVs were not detected; rather, the rough endoplasmic reticulum was dramatically swollen. The results of this study suggest that autophagy is required for formation of double membrane-bound MHV replication complexes and that DMV formation significantly enhances the efficiency of replication. Furthermore, the rough endoplasmic reticulum is implicated as the possible source of membranes for replication complexes.