Immunohistochemical Characterisation of Cell-Type Specific Expression of CK1δ in Various Tissues of Young Adult BALB/c Mice

Background

Casein kinase 1 delta (CK1δ) phosphorylates many key proteins playing important roles in such biological processes as cell growth, differentiation, apoptosis, circadian rhythm and vesicle transport. Furthermore, deregulation of CK1δ has been linked to neurodegenerative diseases and cancer. In this study, the cell specific distribution of CK1δ in various tissues and organs of young adult BALB/c mice was analysed by immunohistochemistry.

Methodology/Principal Findings

Immunohistochemical staining of CK1δ was performed using three different antibodies against CK1δ. A high expression of CK1δ was found in a variety of tissues and organ systems and in several cell types of endodermal, mesodermal and ectodermal origin.

Conclusions

These results give an overview of the cell-type specific expression of CK1δ in different organs under normal conditions. Thus, they provide evidence for possible cell-type specific functions of CK1δ, where CK1δ can interact with and modulate the activity of key regulator proteins by site directed phosphorylation. Furthermore, they provide the basis for future analyses of CK1δ in these tissues.     

An automated quantitative image analysis tool for the identification of microtubule patterns in plants

High throughput confocal imaging poses challenges in the computational image analysis of complex subcellular structures such as the microtubule cytoskeleton. Here, we developed CellArchitect, an automated image analysis tool that quantifies changes to subcellular patterns illustrated by microtubule markers in plants. We screened microtubule‐targeted herbicides and demonstrate that high throughput confocal imaging with integrated image analysis by CellArchitect can distinguish effects induced by the known herbicides indaziflam and trifluralin. The same platform was used to examine 6 other compounds with herbicidal activity, and at least 3 different effects induced by these compounds were profiled. We further show that CellArchitect can detect subcellular patterns tagged by actin and endoplasmic reticulum markers. Thus, the platform developed here can be used to automate image analysis of complex subcellular patterns for purposes such as herbicide discovery and mode of action characterisation. The capacity to use this tool to quantitatively characterize cellular responses lends itself to application across many areas of biology.      

Chromosome-scale genome assembly of the transformation-amenable common wheat cultivar ‘Fielder’

We have established a high-quality, chromosome-level genome assembly for the hexaploid common wheat cultivar ‘Fielder’, an American, soft, white, pastry-type wheat released in 1974 and known for its amenability to Agrobacterium tumefaciens-mediated transformation and genome editing. Accurate, long-read sequences were obtained using PacBio circular consensus sequencing with the HiFi approach. Sequence reads from 16 SMRT cells assembled using the hifiasm assembler produced assemblies with N50 greater than 20 Mb. We used the Omni-C chromosome conformation capture technique to order contigs into chromosome-level assemblies, resulting in 21 pseudomolecules with a cumulative size of 14.7 and 0.3 Gb of unanchored contigs. Mapping of published short reads from a transgenic wheat plant with an edited seed-dormancy gene, TaQsd1, identified four positions of transgene insertion into wheat chromosomes. Detection of guide RNA sequences in pseudomolecules provided candidates for off-target mutation induction. These results demonstrate the efficiency of chromosome-scale assembly using PacBio HiFi reads and their application in wheat genome-editing studies.     

A homozygous <Emphasis Type="Italic">RAB3GAP2</Emphasis> mutation causes Warburg Micro syndrome

Warburg Micro syndrome and Martsolf syndrome are clinically overlapping autosomal recessive conditions characterized by congenital cataracts, microphthalmia, postnatal microcephaly, and developmental delay. The neurodevelopmental and ophthalmological phenotype is more severe in Warburg Micro syndrome in which cerebral malformations and severe motor and mental retardation are common. While biallelic loss-of-function mutations in RAB3GAP1 are present in the majority of patients with Warburg Micro syndrome; a hypomorphic homozygous splicing mutation of RAB3GAP2 has been reported in a single family with Martsolf syndrome. Here, we report a novel homozygous RAB3GAP2 small in-frame deletion, c.499_507delTTCTACACT (p.Phe167_Thr169del) that causes Warburg Micro syndrome in a girl from a consanguineous Turkish family presenting with congenital cataracts, microphthalmia, absent visually evoked potentials, microcephaly, polymicrogyria, hypoplasia of the corpus callosum, and severe developmental delay. No RAB3GAP2 mutations were detected in ten additional unrelated patients with RAB3GAP1-negative Warburg Micro syndrome, consistent with further genetic heterogeneity. In conclusion, we provide evidence that RAB3GAP2 mutations are not specific to Martsolf syndrome. Rather, our findings suggest that loss-of-function mutations of RAB3GAP1 as well as functionally severe RAB3GAP2 mutations cause Warburg Micro syndrome while hypomorphic RAB3GAP2 mutations can result in the milder Martsolf phenotype. Thus, a phenotypic severity gradient may exist in the RAB3GAP-associated disease continuum (the “Warburg–Martsolf syndrome”) which is presumably determined by the mutant gene and the nature of the mutation.     

Response of macroinvertebrates to warming,nutrient addition and predation in large-scale mesocosm tanks

There is increasing concern about the effect of climate change on aquatic systems. We examined changes in macroinvertebrate communities caused by increased temperature (3°C above ambient during summer only and continuous 3°C above ambient all year round), influences of fish (Gasterosteus aculeatus L.) and addition of nutrients (nitrogen and phosphorus) in 48 large-scale (3000 l) tanks over a 2 year period. While numbers of Isopoda, Chaoborus, Corixidae, Ephemeroptera, Notonectidae and Odonata were reduced by the presence of fish, nutrient addition caused isopods, corixids, mayflies and odonates to increase in abundance. Impacts of temperature increase were surprisingly low, with only gastropods increasing in heated tanks, suggesting that, overall abundances of most macroinvertebrate taxa will not be severely affected by the predicted temperature rise. To determine if taxa were sampled representatively during the experiment, net sweep samples taken towards the end of the experiment were compared with final macroinvertebrate abundances when the complete contents of each tank were harvested. We found that net sweeping is an appropriate semi-quantitative method for most taxa in mesocosm tanks. However, mites, coleopteran adults and larvae, dipterans and Chaoborus were not adequately sampled. This might explain why we could not detect any treatment effects of temperature, fish or nutrients on mites, coleopterans and dipterans and calls for different sampling techniques for these taxa, especially in ponds with vegetation stands.     

Morphology and structural organization of spiracles in femaleArgas (Persicargas) walkerae (Acari: Argasidae)

Scanning electron microscopical investigations of fractures and corrosion casts of the spiracles from femaleA. walkerae ticks revealed a four-part structure, consisting of spiracular plate, ostium and macula forming the external closure, followed by the subostial space and the vestibulum of regulable volume, as well as the atrial chamber as the innermost part from which the main tracheal trunks originate. On the average, the spiracular plate was 158 m long and 188 m at the broadest width. It consisted of a thin, highly perforated external and a thick internal layer, which enclosed the interpedicellar space with numerous stout pedicels. In its posterior region, the spiracular plate was covered by the macula, which was up to 80 m in length and 110 m in width. The interpedicellar cavity opened into the subostial space measuring 95.5 m in length and 159.6m in width, which proceeded into the 112-m long vestibulum. The roof of the vestibulum was flexible and could be everted and inverted. Inverted, the roof formed a quadratic bulge with numerous deep cuticular folds, which confined the lumen of the vestibulum either partially or completely. In corrosion casts, the roof was everted to a length of up to 89.3 m. In the posterior part of the vestibulum, as well as in the initial fourth of the artrial chamber, numerous anvil-, cone-or drop-like cuticular projections were arranged in wedge-like fashion. The atrial chamber was almost spherical with a diameter of 138.4 m. Five main tracheal trunks of different luminal diameter as well as numerous channels opened into the atrial chamber.     

Common ragweed (Ambrosia artemisiifolia L.): allergenicity and molecular characterization of pollen after plant exposure to elevated NO2

Ragweed pollen is the main cause of allergenic diseases in Northern America, and the weed has become a spreading neophyte in Europe. Climate change and air pollution are speculated to affect the allergenic potential of pollen. The objective of this study was to investigate the effects of NO₂, a major air pollutant, under controlled conditions, on the allergenicity of ragweed pollen. Ragweed was exposed to different levels of NO₂ throughout the entire growing season, and its pollen further analysed. Spectroscopic analysis showed increased outer cell wall polymers and decreased amounts of pectin. Proteome studies using two‐dimensional difference gel electrophoresis and liquid chromatography–tandem mass spectrometry indicated increased amounts of several Amb a 1 isoforms and of another allergen with great homology to enolase Hev b 9 from rubber tree. Analysis of protein S‐nitrosylation identified nitrosylated proteins in pollen from both conditions, including Amb a 1 isoforms. However, elevated NO₂ significantly enhanced the overall nitrosylation. Finally, we demonstrated increased overall pollen allergenicity by immunoblotting using ragweed antisera, showing a significantly higher allergenicity for Amb a 1. The data highlight a direct influence of elevated NO₂ on the increased allergenicity of ragweed pollen and a direct correlation with an increased risk for human health.