Calsyntenins Mediate TGN Exit of APP in a Kinesin-1-Dependent Manner

Kinesin motors are required for the export of membranous cargo from the trans-Golgi network (TGN), yet information about how kinesins are recruited to forming transport intermediates is sparse. Here we show that the Kinesin-1 docking protein calsyntenin-1 localizes to the TGN in vivo and directly and specifically recruits Kinesin-1 to Golgi/TGN membranes as well as to dynamic post-Golgi carriers. Overexpression of various calsyntenin chimeras and kinesin light chain 1 (KLC1) at high levels caused the formation of aberrant membrane stacks at the endoplasmic reticulum (ER) or the Golgi, disrupted overall Golgi structure and blocked exit of calsyntenin from the TGN. Intriguingly, this blockade of calsyntenin exit strongly and selectively impeded TGN exit of amyloid precursor protein (APP). Using live cell microscopy we found that calsyntenins exit the TGN in Kinesin-1-decorated tubular structures which may serve as carriers for calsyntenin-1-mediated post-TGN transport of APP. Abrogation of this pathway via virus-mediated knockdown of calsyntenin-1 expression in primary cultured neurons caused a marked elevation of APP C-terminal fragments. Together, these results indicate a role for calsyntenin-1 in Kinesin-1-dependent TGN exit and post-Golgi transport of APP-containing organelles and further suggest that distinct intracellular routes may exhibit different capacities for proteolytic processing of APP.     

Influence of enzyme supplementation of diets and cooking–flaking of maize on digestive traits and growth performance of broilers from 1 to 21 days of age

A trial was conducted to study the influence of cooking–flaking (C-F) of maize and enzyme supplementation (ES) of the diet on mucosa morphometry, digestive organ weight, dietary component digestibility and growth performance of broilers from 1 to 21 days of age. There were 4 treatments arranged factorially with 2 heat processings of maize (raw and C-F) and 2 levels (0 and 500 mg/kg) of an enzyme complex with xylanase, protease and α-amylase activity. Dietary treatment did not affect any productive trait from 1 to 21 days of age. However, from 1 to 4 days of age, body weight gain (P<0.01) and feed conversion ratio (P<0.001) were improved by ES of the diet. Intestinal viscosity increased with C-F of maize and decreased with ES of the diet (P<0.001). The effects of ES on intestinal viscosity were more pronounced with C-F than with raw maize (P<0.001) and at younger than older ages (P<0.05). Cooking–flaking of maize increased total tract apparent digestibility of dry matter and organic matter (P<0.05) and of neutral detergent fibre and starch (P<0.001) and tended to improve ether extract (EE) digestibility (P<0.10). In addition, ES improved nitrogen retention (P<0.01) and EE digestibility (P<0.001). Digestibility of neutral detergent fibre and starch increased linearly (P<0.001) with age whereas nitrogen retention and organic matter, EE and gross energy digestibility decreased from 4 to 8 days of age and increased thereafter (P<0.001). The relative weight (g/kg body weight) of the pancreas decreased (P<0.01) and that of the liver increased (P<0.001) with C-F of maize. The relative weight of the proventriculus, gizzard and liver reached a maximum before 6 days of age and that of the small intestine and pancreas at approximately 7.8 days of age (P<0.001). Enzymes increased villus height (P<0.05) but did not affect villus width or villus surface area. It is concluded that C-F of maize improved dietary component digestibility but had no effect on broiler performance at 21 days of age. In addition, ES improved nitrogen retention, EE digestibility and productive performance of broilers but only from 1 to 4 days of age.     

Molecular cloning of thehom—thrC—thrB cluster fromBacillus sp. ULM1: Expression of thethrC gene inEscherichia coli and corynebacteria,and evolutionary relationships of the threonine genes

A 6.5 kb DNA fragment containing the gene (thrC) encoding threonine synthase, the last enzyme of the threonine biosynthetic pathway, has been cloned from the DNA ofBacillus sp. ULM1 by complementation ofEscherichia coli andBrevibacterium lactofermentum thrC auxotrophs. Complementation studies showed that thethrB gene (encoding homoserine kinase) is found downstream from thethrC gene, and analysis of nucleotide sequences indicated that thehom gene (encoding homoserine dehydrogenase) is located upstream of thethrC gene. The organization of this cluster of genes is similar to theBacillus subtilis threonine operon (hom—thrC—thrB). An 1.9 kbBclI, fragment from theBacillus sp. ULM1 DNA insert that complementedthrC mutations both inE. coli and in corynebacteria was sequenced, and an ORF encoding a protein of 351 amino acids was found corresponding to a protein of 37462 Da. ThethrC gene showed a low G+C content (39.4%) and the encoded threonine synthase is very similar to theB. subtilis enzyme. Expression of the 1.9 kbBclI DNA fragment inE. coli minicells resulted in the formation of a 37 kDa protein. The upstream region of this gene shows promoter activity inE. coli but not in corynebacteria. A peptide sequence, including a lysine that is known to bind the pyridoxal phosphate cofactor, is conserved in all threonine synthase sequences and also in the threonine and serine dehydratase genes. Amino acid comparison of nine threonine synthases revealed evolutionary relationships between different groups of bacteria. Dedicated to Dr. J. Spížek on the occasion of his 60th birthday     

Mothers that produce sons and daughters are genetically different in red deer

Sex ratio theory, and in particular Fisher principle, assumes parental control over the sex of offspring through the action of autosomal genes with Mendelian segregation. In spite of the importance of Fisher's principle in evolutionary biology, the number of studies looking for possible loci involved in sex ratio bias is, at best, very low. Newly developed genetic tools frequently allow evolutionary biologists to manage genetic data. Here we encourage the application of association tools to databases that include genetic information for autosomal loci and offspring sex to improve our knowledge on sex ratio evolution. As an example we use microsatellite markers to scan autosomal chromosomes and look for linked genetic regions associated with offspring sex in red deer (Cervus elaphus). We found a microsatellite marker (CelJP38) mapped in chromosome 27 for which females producing sons and daughters were genetically different. To the best of our knowledge, this is the first study that shows a genetic signal that points out an association between mother genotype and offspring sex in natural populations of a mammal.     

Factors affecting the production of SCEs by maleic hydrazide in root-tip chromosomes of Allium cepa

We have investigated the influence of pH on the induction of chromatid-type aberrations and sister-chromatid exchanges (SCEs) by maleic hydrazide (MH) in root-tip cells of Allium cepa. For both cytogenetic endpoints, the lower the pH of the treatment solution, the higher were the frequencies of chromosome alterations detected at metaphase. We have further studied the persistence of lesions giving rise to SCEs during successive cell cycles, as well as the influence of BrdU concentration in the post-treatment medium on the yield of MH-induced SCEs. Our results suggest that the cytogenetic action of MH in many respects resembles that of bifunctional alkylating agents.     

Influence of Residue 22 on the Folding, Aggregation Profile, and Toxicity of the Alzheimer's Amyloid β Peptide

Several biophysical techniques have been used to determine differences in the aggregation profile (i.e., the secondary structure, aggregation propensity, dynamics, and morphology of amyloid structures) and the effects on cell viability of three variants of the amyloid β peptide involved in Alzheimer's disease. We focused our study on the Glu²² residue, comparing the effects of freshly prepared samples and samples aged for at least 20 days. In the aged samples, a high propensity for aggregation and β-sheet secondary structure appears when residue 22 is capable of establishing polar (Glu²² in wild-type) or hydrophobic (Val²² in E22V) interactions. The Arctic variant (E22G) presents a mixture of mostly disordered and α-helix structures (with low β-sheet contribution). Analysis of transmission electron micrographs and atomic force microscopy images of the peptide variants after aging showed significant quantitative and qualitative differences in the morphology of the formed aggregates. The effect on human neuroblastoma cells of these Aβ_12-28 variants does not correlate with the amount of β-sheet of the aggregates. In samples allowed to age, the native sequence was found to have an insignificant effect on cell viability, whereas the Arctic variant (E22G), the E22V variant, and the slightly-aggregating control (F19G-F20G) had more prominent effects.