Exploring a role for heteromerization in GPCR signalling specificity

The critical involvement of GPCRs (G-protein-coupled receptors) in nearly all physiological processes, and the presence of these receptors at the interface between the extracellular and the intracellular milieu, has positioned these receptors as pivotal therapeutic targets. Although a large number of drugs targeting GPCRs are currently available, significant efforts have been directed towards understanding receptor properties, with the goal of identifying and designing improved receptor ligands. Recent advances in GPCR pharmacology have demonstrated that different ligands binding to the same receptor can activate discrete sets of downstream effectors, a phenomenon known as 'ligand-directed signal specificity', which is currently being explored for drug development due to its potential therapeutic advantage. Emerging studies suggest that GPCR responses can also be modulated by contextual factors, such as interactions with other GPCRs. Association between different GPCR types leads to the formation of complexes, or GPCR heteromers, with distinct and unique signalling properties. Some of these heteromers activate discrete sets of signalling effectors upon activation by the same ligand, a phenomenon termed 'heteromer-directed signalling specificity'. This has been shown to be involved in the physiological role of receptors and, in some cases, in disease-specific dysregulation of a receptor effect. Hence targeting GPCR heteromers constitutes an emerging strategy to select receptor-specific responses and is likely to be useful in achieving specific beneficial therapeutic effects.     

Relationship between reproductive behavior and new shoot development in 5-year-old branches of olive trees (Olea europaea L.)

The development of new shoots plays a central role in the complex interactions determining vegetative and reproductive growth in woody plants. To explore this role we evaluated the new shoots in the olive tree, Olea europaea L., and the effect of fruiting on new shoot growth and subsequent flowering. Five-year-old branches served as canopy subunits in order to obtain a global, whole-tree view of new shoot number, size and morphological origin. The non-bearing trees had many more shoots than the fruit-bearing trees, and a greater number of longer shoots. In both bearing conditions, however, the majority of shoots were less than 4 cm long, with shoots of progressively longer lengths present in successively decreasing frequencies. Six major shoot types were defined on the basis of apical or lateral bud origin and of parent shoot age. On fruit-bearing trees, the new shoots originated predominantly from the shoot apex, while on non-fruiting trees, they formed mainly from axillary buds, but in both cases, they tended to develop on younger parent shoots. The previous bearing condition of the tree was the main determinant for subsequent inflorescence development, which was independent of both shoot type and length. Thus, reproductive behavior strongly affected both the amount and type of new branching, but subsequent flowering level was more influenced by previous bearing than by the potential flowering sites on new shoots.     

ATM-mediated phosphorylation of polynucleotide kinase/phosphatase is required for effective DNA double-strand break repair

The cellular response to double-strand breaks (DSBs) in DNA is a complex signalling network, mobilized by the nuclear protein kinase ataxia-telangiectasia mutated (ATM), which phosphorylates many factors in the various branches of this network. A main question is how ATM regulates DSB repair. Here, we identify the DNA repair enzyme polynucleotide kinase/phosphatase (PNKP) as an ATM target. PNKP phosphorylates 5'-OH and dephosphorylates 3'-phosphate DNA ends that are formed at DSB termini caused by DNA-damaging agents, thereby regenerating legitimate ends for further processing. We establish that the ATM phosphorylation targets on human PNKP-Ser 114 and Ser 126-are crucial for cellular survival following DSB induction and for effective DSB repair, being essential for damage-induced enhancement of the activity of PNKP and its proper accumulation at the sites of DNA damage. These findings show a direct functional link between ATM and the DSB-repair machinery.     

Gibbs free energy as a measure of complexity correlates with time within <Emphasis Type="Italic">C. elegans</Emphasis> embryonic development

We investigate free energy behavior in the nematode Caenorhabditis elegans during embryonic development. Our approach utilizes publicly available gene expression data, which gives us a picture of developmental changes in protein concentration and, resultantly, chemical potential and free energy. Our results indicate a clear global relationship between Gibbs free energy and time spent in development and provide thermodynamic indicators of the large-scale biological events of cell division and differentiation.     

Structural insights on biologically relevant cationic membranes by ESR spectroscopy

Cationic bilayers have been used as models to study membrane fusion, templates for polymerization and deposition of materials, carriers of nucleic acids and hydrophobic drugs, microbicidal agents and vaccine adjuvants. The versatility of these membranes depends on their structure. Electron spin resonance (ESR) spectroscopy is a powerful technique that employs hydrophobic spin labels to probe membrane structure and packing. The focus of this review is the extensive structural characterization of cationic membranes prepared with dioctadecyldimethylammonium bromide or diC14-amidine to illustrate how ESR spectroscopy can provide important structural information on bilayer thermotropic behavior, gel and fluid phases, phase coexistence, presence of bilayer interdigitation, membrane fusion and interactions with other biologically relevant molecules.     

Mechanism of enzymatic crosslinking of fibrinogen molecules

The mechanism of enzymatic covalent crosslinking of fibrinogen molecules under the effect of plasma transglutaminase (factor XIIIa) has been studied. Using the methods of elastic and dynamic light scattering combined with viscosimetry and electrophoresis of the reduced samples, it has been shown that fibrinogen oxidation strongly accelerates self-assembly of the covalently crosslinked fibrinogen polymers. IR-spectroscopy data indicate that the degree of fibrinogen oxidation positively correlates with the amount of ε/(γ-glu)lys isopeptide covalent bonds whose formation is catalyzed by the factor XIIIa. The results of this and our previous studies cast doubt on the widespread concept that native fibrinogen is the substrate for factor XIIIa. In our opinion, only structurally defective fibrinogen molecules that have undergone oxidative structural conversion in the D-domain region are involved in the enzymatic reaction leading to the formation of covalent ε/(γ-glu)lys isopeptide bonds and self-assembly of fibrinogen polymers.