Canonical Wnt signaling: high-throughput RNAi widens the path

The canonical Wnt signaling pathway is highly conserved in evolution, widely used throughout animal development, and frequently hyperactive in cancer. Although Wnt signaling has been the subject of extensive genetic analysis in the past, some 200 genes have now been identified as candidate modulators of this pathway by a recent study using high-throughput RNAi screening.     

Using morphological characters of subfossil daphniid postabdominal claws to improve taxonomic resolution within species complexes

Daphnia subfossils from lake sediments are useful for exploring the impacts of environmental stressors on aquatic ecosystems. Unfortunately, taxonomic resolution of Daphnia remains is coarse, as only a small portion of the animal is preserved, and so the identification of daphniid subfossils typically relies upon postabdominal claws. Daphniid claws can be assigned to one of two species complexes: D. longispina or D. pulex. Both complexes contain species with differing environmental optima, and therefore improved taxonomic resolution of subfossil daphniid claws would aid paleolimnological analyses. To identify morphological features that may be used to help differentiate between species within complexes, we used species presence/absence data from net tows to select lakes in central Ontario (Canada) containing only a single species from a particular complex, then used remains preserved in surface sediments of these lakes to isolate four Daphnia species: D. ambigua and D. mendotae from the D. longispina complex, and D. pulicaria and D. catawba from the D. pulex complex. Our analyses demonstrate that, within the D. longispina complex, postabdominal claw length (PCL) and spinule length can be used to distinguish D. mendotae from D. ambigua. In addition, within the D. pulex complex, there are differences between D. pulicaria and D. catawba in the relative lengths of the proximal and middle combs on the postabdominal claw. However, the number of stout spines on the middle comb is an unreliable character for differentiating species. Overall, our data demonstrate that greater resolution within Daphnia species complexes is possible using postabdominal claws; however, the process is arduous, and applicability will likely decrease with the number of taxa present.     

One-step refolding and purification of disulfide-containing proteins with a C-terminal MESNA thioester

Background  

Expression systems based on self-cleavable intein domains allow the generation of recombinant proteins with a C-terminal thioester. This uniquely reactive C-terminus can be used in native chemical ligation reactions to introduce synthetic groups or to immobilize proteins on surfaces and nanoparticles. Unfortunately, common refolding procedures for recombinant proteins that contain disulfide bonds do not preserve the thioester functionality and therefore novel refolding procedures need to be developed.     

Integrating gene expression and GO classification for PCA by preclustering

Background  

Gene expression data can be analyzed by summarizing groups of individual gene expression profiles based on GO annotation information. The mean expression profile per group can then be used to identify interesting GO categories in relation to the experimental settings. However, the expression profiles present in GO classes are often heterogeneous, i.e., there are several different expression profiles within one class. As a result, important experimental findings can be obscured because the summarizing profile does not seem to be of interest. We propose to tackle this problem by finding homogeneous subclasses within GO categories: preclustering.     

Calcium content of littoral Cladocera in three softwater lakes of the Canadian Shield

Aqueous calcium (Ca) concentrations are declining in softwater lakes of the Canadian Shield largely because of decades of acid deposition and afforestation following timber harvesting. Populations of pelagic cladoceran taxa with high Ca requirements, especially Daphnia spp., are declining in response to reduced aqueous Ca availability. However, the Ca content, and thus the requirements of littoral cladoceran taxa are unknown; therefore, the potential vulnerability of this major component of lake ecosystems to ongoing regional Ca decline remains uncertain. Here, we identify the body Ca concentration of nine littoral cladoceran taxa collected from three lakes in the Muskoka-Haliburton region of Ontario, Canada. Ca content differed among taxonomic groups, ranging from 3.6 mg g⁻¹ for Acroperus harpae to 23.2 mg g⁻¹ for Disparalona spp. Perhaps surprisingly, some littoral microcrustacean taxa have Ca burdens comparable to the Ca-rich daphniids. Therefore, there may be differential responses to ongoing lake water Ca declines among taxa within littoral communities as has been observed among open-water taxa, with potential ecological repercussions for near-shore food webs.     

Molecular phylogenetics at the population/species interface in cave spiders of the southern Appalachians (Araneae:Nesticidae:Nesticus)

This paper focuses on the relationship between population genetic structure and speciation mechanisms in a monophyletic species group of Appalachian cave spiders (Nesticus). Using mtDNA sequence data gathered from 256 individuals, I analyzed patterns of genetic variation within and between populations for three pairs of closely related sister species. Each sister-pair comparison involves taxa with differing distributional and ecological attributes; if these ecological attributes are reflected in basic demographic differences, then speciation might proceed differently across these sister taxa comparisons. Both frequency-based and gene tree analyses reveal that the genetic structure of the Nesticus species studied is characterized by similar and essentially complete population subdivision, regardless of differences in general ecology. These findings contrast with results of prior genetic studies of cave-dwelling arthropods that have typically revealed variation in population structure corresponding to differences in general ecology. Species fragmentation through both extrinsic and intrinsic evolutionary forces has resulted in discrete, perhaps independent, populations within morphologically defined species. Large sequence divergence values observed between populations suggest that this independence may extend well into the past. These patterns of mtDNA genealogical structure and divergence imply that species as morphological lineages are currently more inclusive than basal evolutionary or phylogenetic units, a suggestion that has important implications for the study of speciation mechanisms.      

Chronic stress and its effects on adrenal cortex apoptosis in pregnant rats

The model of chronic intermittent stress by immobilization during pregnancy may produce alterations in the mechanisms that maintain adrenal gland homeostasis. In earlier investigations using this model, significant variations in plasma prolactin and corticosterone levels, and adrenal gland weights were observed. We hypothesized that chronic stress causes changes in apoptosis in the adrenal glands of pregnant rats. We identified and quantified apoptotic cells in the adrenal cortex and examined their ultrastructural characteristics using transmission electron microscopy. Adrenal glands of pregnant rats at gestation days 12, 17 and 21 were studied for control and experimental (stressed) rats. Immunolabelling techniques, stereological analysis and image quantification of adrenal gland sections were combined to determine differences in apoptosis in the different cell populations of the adrenal cortex. The apoptotic index of the experimental rats showed a significant reduction at gestation day 17, while at days 12 and 21 there were no differences from controls. Moreover, the apoptotic index of the reticular zones in control and experimental animals showed a significant increase compared to the glomerular and fascicular zones at the three gestation times studied. Chronic stress by immobilization reduced the caspase-dependent apoptotic index at gestation day 17, which may be related to variations in plasma concentrations of estrogens and prolactin.     

HistoChoice® as an alternative to formalin fixation of undecalcified bone specimens

We compared histochemical and immunohistochemical staining as well as fluorochrome labeling in murine bone specimens that were fixed with 10% neutral buffered formalin to those fixed with HistoChoice®. We showed that sections from undecalcified tibiae fixed for 4 h in HistoChoice® resulted in enhanced toluidine blue and Von Kossa histochemical staining compared to formalin fixation. HistoChoice® produced comparable or improved staining for alkaline phosphatase. Acid phosphatase localization was better in formalin fixed specimens, but osteoclasts were visuralized more easily in HistoChoice® fixed specimens. As expected, immunohistochemical labeling was antibody dependent; some antibodies labeled better in HistoChoice® fixed specimens while others were better in formalin fixed specimens. Toluidine blue, Von Kossa, and alkaline phosphatase staining of sections fixed for 12 h produced sections that were similar to 4 h fixed sections. Fixation for 12 h preserved acid phosphatase activity better. Increasing fixation to 12 h affected immunolocalization differentially. Bone sialoprotein labeling in HistoChoice® fixed specimens was comparable to formalin fixed samples. On the other hand, after 12 h formalin fixation, osteocalcin labeling was comparable to HistoChoice®. For most histochemical applications, fixing murine bone specimens for 4 h with HistoChoice® yielded superior staining compared to formalin fixation. If immunohistochemical localization is desired, however, individual antibodies must be tested to determine which fixation process retains antigenicity better. In addition, there was no detectable difference in the intensity of fluorochrome labeling using either fixative. Finally, fixation duration did not alter the intensity of labeling.