Scanning electron microscopy of cloned astrocytic lines derived from ethylnitrosourea-induced rat gliomas

Six cloned astrocytoma cell lines derived from four ethylnitrosourea-induced F-344 rat gliomas were viewed by scanning electron microscopy in vitro, and two were examined in vivo after transplantation to the intracerebral site. All clones consisted of stellate cells that were reasonably homogeneous within individual glioma lines. Cell membrane features common to all tumor lines included microvilli, blebs, ruffles, and miniridges, mainly confined to perikarya, and filopodia emanating chiefly from cell processes. One cell line demonstrated a profuse, and another cell line a moderate, degree of microvillous development and cell surface roughening, which in one tumor correlated with rapid in vitro cell doubling time. Both cell lines maintained these topographical appearances when transplanted into brain. These results extend the SEM observations of astrocytomas, particularly in cloned ethylnitrosourea-induced tumors in rats. The confirm that distinct variations in cell membrane topography do occur among tumors of this type, probably irrespective of their origin in humans or rats, and irrespective of their mode of genesis as spontaneous, chemically-induced, or virally-induced tumors.     

Quantitative trait loci for foliar terpenes in a global eucalypt species

Terpenes are a diverse group of plant secondary metabolites that mediate a plethora of ecological interactions in many plant species. Despite increasing research into the genetic control of important adaptive traits in some plant species, the genetic control of terpenes in forest tree species is still relatively poorly studied. In this study, we use quantitative genetic and quantitative trait loci (QTL) analysis to investigate the genetic control of foliar terpenes in an ecologically and commercially important eucalypt species, Eucalyptus globulus. We show a moderate to high within-family broad-sense heritability and significant genetic basis to the variation in 14 of the 16 terpenes assayed. This is the first report of QTL for terpenes in this species. Eleven QTL influenced the terpenes overall. One QTL on linkage group 6 affected six of the seven different sesquiterpenes assayed (plus one monoterpene), which, in combination with highly significant correlations between these compounds, argues that their variation is influenced by a QTL with pleiotropic effect early in the biosynthetic pathway. We examine the homology of these QTL to those found in a closely related eucalypt, Eucalyptus nitens, and provide evidence that both common and unique QTL influence terpene levels.     

Wood construction more strongly shapes deadwood microbial communities than spatial location over 5 years of decay

Diverse communities of fungi and bacteria in deadwood mediate wood decay. While rates of decomposition vary greatly among woody species and spatially distinct habitats, the relative importance of these factors in structuring microbial communities and whether these shift over time remains largely unknown. We characterized fungal and bacterial diversity within pieces of deadwood that experienced 6.3–98.8% mass loss while decaying in common garden ‘rotplots’ in a temperate oak-hickory forest in the Ozark Highlands, MO, USA. Communities were isolated from 21 woody species that had been decomposing for 1–5 years in spatially distinct habitats at the landscape scale (top and bottom of watersheds) and within stems (top and bottom of stems). Microbial community structure varied more strongly with wood traits than with spatial locations, mirroring the relative role of these factors on decay rates on the same pieces of wood even after 5 years. Co-occurring fungal and bacterial communities persistently influenced one another independently from their shared environmental conditions. However, the relative influence of wood construction versus spatial locations differed between fungi and bacteria, suggesting that life history characteristics of these clades structure diversity differently across space and time in decomposing wood.     

PEATmoss (Physcomitrella Expression Atlas Tool): a unified gene expression atlas for the model plant Physcomitrella patens

Physcomitrella patens is a bryophyte model plant that is often used to study plant evolution and development. Its resources are of great importance for comparative genomics and evo‐devo approaches. However, expression data from Physcomitrella patens were so far generated using different gene annotation versions and three different platforms: CombiMatrix and NimbleGen expression microarrays and RNA sequencing. The currently available P. patens expression data are distributed across three tools with different visualization methods to access the data. Here, we introduce an interactive expression atlas, Physcomitrella Expression Atlas Tool (PEATmoss), that unifies publicly available expression data for P. patens and provides multiple visualization methods to query the data in a single web‐based tool. Moreover, PEATmoss includes 35 expression experiments not previously available in any other expression atlas. To facilitate gene expression queries across different gene annotation versions, and to access P. patens annotations and related resources, a lookup database and web tool linked to PEATmoss was implemented. PEATmoss can be accessed at https://peatmoss.online.uni-marburg.de     

Maximizing antibody production in a targeted integration host by optimization of subunit gene dosage and position

Historically, therapeutic protein production in Chinese hamster ovary (CHO) cells has been accomplished by random integration (RI) of expression plasmids into the host cell genome. More recently, the development of targeted integration (TI) host cells has allowed for recombination of plasmid DNA into a predetermined genomic locus, eliminating one contributor to clone-to-clone variability. In this study, a TI host capable of simultaneously integrating two plasmids at the same genomic site was used to assess the effect of antibody heavy chain and light chain gene dosage on antibody productivity. Our results showed that increasing antibody gene copy number can increase specific productivity, but with diminishing returns as more antibody genes are added to the same TI locus. Random integration of additional antibody DNA copies in to a targeted integration cell line showed a further increase in specific productivity, suggesting that targeting additional genomic sites for gene integration may be beneficial. Additionally, the position of antibody genes in the two plasmids was observed to have a strong effect on antibody expression level. These findings shed light on vector design to maximize production of conventional antibodies or tune expression for proper assembly of complex or bispecific antibodies in a TI system.     

Water structure around a left-handed Z-DNA fragment analyzed by cryo neutron crystallography

Even in high-quality X-ray crystal structures of oligonucleotides determined at a resolution of 1 Å or higher, the orientations of first-shell water molecules remain unclear. We used cryo neutron crystallography to gain insight into the H-bonding patterns of water molecules around the left-handed Z-DNA duplex [d(CGCGCG)]₂. The neutron density visualized at 1.5 Å resolution for the first time allows us to pinpoint the orientations of most of the water molecules directly contacting the DNA and of many second-shell waters. In particular, H-bond acceptor and donor patterns for water participating in prominent hydration motifs inside the minor groove, on the convex surface or bridging nucleobase and phosphate oxygen atoms are finally revealed. Several water molecules display entirely unexpected orientations. For example, a water molecule located at H-bonding distance from O6 keto oxygen atoms of two adjacent guanines directs both its deuterium atoms away from the keto groups. Exocyclic amino groups of guanine (N2) and cytosine (N4) unexpectedly stabilize waters H-bonded to O2 keto oxygens from adjacent cytosines and O6 keto oxygens from adjacent guanines, respectively. Our structure offers the most detailed view to date of DNA solvation in the solid-state undistorted by metal ions or polyamines.     

Modeling Cumulative Effects of Climate and Development on Moose,Wolf, and Caribou Populations

Wildlife models focused solely on a single strong influence (e.g., habitat components, wildlife harvest) are limited in their ability to detect key mechanisms influencing population change. Instead, we propose integrated modeling in the context of cumulative effects assessment using multispecies population dynamics models linked to landscape-climate simulation at large spatial and temporal scales. We developed an integrated landscape and population simulation model using ALCES Online as the model-building platform, and the model accounted for key ecological components and relationships among moose (Alces alces), grey wolves (Canis lupus nubilus), and woodland caribou (Rangifer tarandus caribou) in northern Ontario, Canada. We simulated multiple scenarios over 5 decades (beginning 2020) to explore sensitivity to climate change and land use and assessed effects at multiple scales. The magnitude of effect and the relative importance of key factors (climate change, roads, and habitat) differed depending on the scale of assessment. Across the full extent of the study area (654,311km² [ecozonal scale]), the caribou population declined by 26% largely because of climate change and associated predator-prey response, which led to caribou range recession in the southern part of the study area. At the caribou range scale (108,378 km²), which focused on 2 herds in the northern part of the study area, climate change led to a 10% decline in the population and development led to an additional 7% decline. At the project scale (8,331 km²), which was focused more narrowly on the landscape surrounding 4 proposed mines, the caribou population declined by 29% largely in response to simulated development. Given that observed caribou population dynamics were sensitive to the cumulative effects of climate change, land use, interspecific interactions, and scale, insights from the analysis might not emerge under a less complex model. Our integrated modeling framework provides valuable support for broader regional assessments, including estimation of risk to caribou and Indigenous food security, and for developing and evaluating potential caribou recovery strategies. © 2021 The Authors. The Journal of Wildlife Management published by Wiley Periodicals LLC on behalf of The Wildlife Society.     

As old as the hills: Pliocene palaeogeographical processes influence patterns of genetic structure in the widespread,common shrub Banksia sessilis

The impact of Quaternary glaciation on the development of phylogeographic structure in plant species is well documented. In unglaciated landscapes, phylogeographic patterns tend to reflect processes relating to persistence and stochasticity, yet other factors, associated with the palaeogeographical history of the landscape, including geomorphological events, can also have a significant influence. The unglaciated landscape of south‐western Western Australia is an ideal location to observe these ancient drivers of lineage diversification, with tectonic activity associated with the Darling Fault in the late Pliocene attributed to patterns of deep phylogeographic divergence in a widespread tree from this region. Interestingly, other species within this region have not shown this pattern and this palaeogeographical boundary therefore presents an opportunity to examine age and historical distribution of plant species endemic to this region. In this study, we assess patterns of genetic diversity and structure across 28 populations of the widespread shrub Banksia sessilis using three cpDNA markers and nine nuclear microsatellite markers. Sixteen cpDNA haplotypes were identified, comprising two major chloroplast DNA lineages that are estimated to have diverged in the Pliocene, approximately 3.3 million years ago. This timing coincides with major geomorphological processes in the landscape, including the separation of the Darling Plateau from the adjacent Swan Coastal Plain, as well as eustatic changes on the Swan Coastal Plain that are likely to have resulted in the physical isolation of historical plant lineages. Chloroplast lineages were broadly aligned with populations associated with older lateritic soils of the Darling Plateau and Geraldton sandplains or the younger sandy soils associated with the Swan Coastal Plain and Southern Coastline. This structural pattern of lateritic versus non‐lateritic division was not observed in the nuclear microsatellite data that identified three genetic clades that roughly corresponded to populations in the North, South, and Central portions of the distributions.