Exchange factors,effectors, GAPs and motor proteins: common thermodynamic and kinetic principles for different functions

In this article, we review the properties of several classes of proteins that interact with ATPases and GTPases involved in energy and signal transduction. We show that certain common basic thermodynamic principles apply to the manner in which the nucleotide hydrolases interact with their partner molecules, and that the principles involved in signal transduction can be quantitatively modified to give systems with the known properties of energy transducing systems. As an example, actin can be described as an exchange factor for myosin, with its exchange activity being specific for ATP or inorganic phosphate in the myosin.ADP.P(i) complex, in contrast to the unspecific exchange activity of guanosine nucleotide exchange factors operating on GTPases involved in signal transduction and regulatory processes. These common aspects are reflected in shared structural features, suggesting an evolutionary relationship between such systems.     

A bacterial artificial chromosome-based genetic linkage map of the nematode Pristionchus pacificus

To understand the evolution of developmental processes, nonmodel organisms in the nematodes, insects, and vertebrates are compared with established model systems. Often, these comparisons suffer from the inability to apply sophisticated technologies to these nonmodel species. In the nematode Pristionchus pacificus, cellular and genetic analyses are used to compare vulva development to that of Caenorhabditis elegans. However, substantial changes in gene function between P. pacificus and C. elegans limit the use of candidate gene approaches in studying P. pacificus mutations. To facilitate map-based cloning of mutations in P. pacificus, we constructed a BAC-based genetic linkage map. A BAC library of 13,440 clones was generated and completely end sequenced. By comparing BAC end and EST sequences between the "wild-type" strain P. pacificus var. California and the polymorphic strain P. pacificus var. Washington, 133 single-stranded conformational polymorphisms were identified. These markers were tested on a meiotic mapping panel of 46 randomly picked F(2) animals after a cross of the two strains, providing the first genetic linkage map of P. pacificus. A mapping strategy using two selected markers per chromosome was devised and the efficiency of this approach was illustrated by the mapping of the Ppa-unc-1/Twitchin gene.     

Significance of Agonistic Vocalization in the Croaking Gourami (Trichopsis vittatus,Teleostei)

Trichopsis vittatus emits high amplitude sounds during agonistic encounters with conspecifics. The sound producing organ is derived from the structural components of the pectoral fins. The study involved muting a sample of subjects by removing two pectoral fin tendons without any further restriction in movements and behaviour. Mute and unaltered males were then placed together in pairs and the following agonistic behavioural elements were determined: attacks, lateral displays, sound production and frontal displays. Soniferous males had a higher probability of winning contests when size differences were small. In pairs with big size ratios, the larger males were more often successful. Besides visual and acoustical signals, lateral line stimuli seem to play no role in threatening displays. These results demonstrate that vocalization during agonistic encounters is important for becoming dominant in specific circumstances. Because of correlation between main frequencies and size, sound emission might be a very effective method of assessing the physical strength of an opponent.     

Diaminothiazoles Modify Tau Phosphorylation and Improve the Tauopathy in Mouse Models

Although Tau accumulation is a feature of several neurodegenerative conditions, treatment options for these conditions are nonexistent. Targeting Tau kinases represents a potential therapeutic approach. Small molecules in the diaminothiazole class are potent Tau kinase inhibitors that target CDK5 and GSK3β. Lead compounds from the series have IC₅₀ values toward CDK5/p25 and GSK3β in the low nanomolar range and no observed toxicity in the therapeutic dose range. Neuronal protective effects and decreased PHF-1 immunoreactivity were observed in two animal models, 3×Tg-AD and CK-p25. Treatment nearly eliminated Sarkosyl-insoluble Tau with the most prominent effect on the phosphorylation at Ser-404. Treatment also induced the recovery of memory in a fear conditioning assay. Given the contribution of both CDK5/p25 and GSK3β to Tau phosphorylation, effective treatment of tauopathies may require dual kinase targeting.     

Folding and refolding of proteins in chromatographic beds

The correct folding of solubilized recombinant proteins is of key importance for their production in industry. On-column refolding of proteins is mainly achieved by three methods: size-exclusion chromatography, ion exchange chromatography and affinity chromatography using immobilized metal chelates. The principles of these methods were first laid down in the 1990s, but many recent improvements have been made to these processes including sophisticated changes to the mobile phase composition and the recycling of aggregates to improve yield. Advances have also been made in the use of immobilized metal affinity chromatography and by mimicking the natural folding process with artificial chaperones.     

Parietal endoderm secreted SPARC promotes early cardiomyogenesis in vitro

Cardiomyogenesis proceeds in the presence of signals emanating from extra-embryonic lineages emerging before and during early eutherian gastrulation. In embryonic stem cell derived embryoid bodies, primitive endoderm gives rise to visceral and parietal endoderm. Parietal endoderm undergoes an epithelial to mesenchymal transition shortly before first cardiomyocytes start to contract rhythmically. Here, we demonstrate that Secreted Protein, Acidic, Rich in Cysteine, SPARC, predominantly secreted by mesenchymal parietal endoderm specifically promotes early myocardial cell differentiation in embryoid bodies. SPARC enhanced the expression of bmp2 and nkx2.5 in embryoid bodies and fetal cardiomyocytes. Inhibition of either SPARC or Bmp2 attenuated in both cases cardiomyogenesis and downregulated nkx2.5 expression. Thus, SPARC directly affects cardiomyogenesis, modulates Bmp2 signaling, and contributes to a positive autoregulatory loop of Bmp2 and Nkx2.5 in cardiomyocytes.     

Cannibalism in a Population of the Medicinal Leech (<Emphasis Type="Italic">Hirudo medicinalis</Emphasis> L.)

Medicinal leeches (Hirudo medicinalis L.) were maintained in large ponds in a commercial leech farm at Biebertal, Germany. The feeding of hungry adult leeches was performed on representative individuals that were placed on cloth soaked with mammalian blood obtained from a local butchery (pig, Sus scrofa). In a second set of experiments, cane toads (Bufo marinus) were used as host organisms. The leeches rapidly attached to the toads, explored the body and sucked blood. After feeding, the fully engorged leeches were placed into the pond or an aquarium. In this artificial habitat, the satiated leeches were attacked by hungry conspecifics, sucked off, and killed. This observation demonstrates that H. medicinalis must be classified as a cannibalistic annelid.     

Analysis of the Growth Response of Air-Grown Rice Coleoptiles to Submergence

The effect of submergence of air-grown rice seedlings (Oryza sativa L. var. Sasanishiki) on coleoptile growth and ultrastructure, extensibility and chemical composition of the cell walls was investigated. The lag-time between start of submergence and the onset of the enhancement of growth was less than 4 h. The growth response was associated with a drastic thinning of the cell walls and an increase in wall extensibility. At the outer epidermal wall of both air-grown and submerged coleoptiles electron-dense (osmiophilic) particles were detected. During submergence, the net accumulation of cellulose and hemicellulose was reduced, but the increase in pectic substances was unaffected. Submergence caused an 80% inhibition of the net accumulation of wall-bound phenolics (ferulic- and diferulic acid) compared with air-grown controls. The osmotic concentration of the tissue saps was not affected by submergence. Our results support the hypothesis that rapid coleoptile elongation under water is caused by an inhibition of the formation of phenolic cross-links between matrix polysaccharides via diferulate, which results in a mechanical stiffening of the cell walls in the air-grown coleoptile.     

On sexual agglutination and mating type substances in isogamous dioecious Chlamydomonads: IV. Unilateral inactivation of the sex contact capacity in compatible and incompatible taxa by α-mannosidase and snake venom protease

Sex cell contact at fertilization is analysed in the mating type reaction of isogamous Chlamydomonas species. Contact is based upon a complementarity between special mating type substances, sex and species specific glycoprotein complexes. In three related taxa, the contact capactiy of their (+) gamete type is sensitive to snake venom protease (α-protease) and depends decisively on terminal α-glycosidically bound mannose residues. Enzymatic removal of these residues by α-mannosidase incapacitates live (+) gametes and inactivates isolated (+) mating type substance. (+) gametes inactivated by α-mannosidase or α-protease do not agglutinate with (?) gametes nor respond to isolated (?) mating type substance. Isolated (?) substance is adsorbed to and inactivated by the homologous (+) gametes. (+) gametes incapacitated by α-mannosidase or α-protease do not adsorb nor inactivate the isolated (?) substance. The agglutinability of live (?) gametes and the contact capacity of isolated (?) mating type substance is not affected by α-mannosidase or α-protease. The mannose residues react only within the species-typical complementarity. Some additional feature(s) of the (+) mating type substances must effect their species specificity and account for gametic isolation and sexual incompatibility between species.