Kinetic Analysis of Permeation of Mitochondria-Targeted Antioxidants Across Bilayer Lipid Membranes

Mitochondria-targeted antioxidants consisting of a quinone part conjugated with a lipophilic cation via a hydrocarbon linker were previously shown to prevent oxidative damage to mitochondria in vitro and in vivo. In the present work, we studied the permeation of a series of compounds of this type across a planar bilayer phospholipid membrane. For this purpose, relaxation of the electrical current after a voltage jump was measured. With respect to the characteristic time of the relaxation process reflecting the permeation rate, hydrophobic cations can be ranked in the following series: 10(plastoquinonyl) decylrhodamine 19 (SkQR1) > 10-(6'-plastoquinonyl) decyltriphenylphosphonium (SkQ1) > 10-(6'-methylplastoquinonyl) decyltriphenylphosphonium (SkQ3) > 10-(6'-ubiquinonyl) decyltriphenylphosphonium (MitoQ). Thus, the permeation rate increased with (1) an increase in the size of the hydrophobic cation and (2) an increase in hydrophobicity of the quinone moiety. SkQ1 containing plastoquinone was shown to be more permeable through the membrane compared to MitoQ containing ubiquinone, which might be the reason for more pronounced beneficial action of SkQ1 in vitro and in vivo. The above approach can be recommended for the search for new antioxidants or other compounds targeted to mitochondria.     

Lipids revert inert Abeta amyloid fibrils to neurotoxic protofibrils that affect learning in mice

Although soluble oligomeric and protofibrillar assemblies of Abeta-amyloid peptide cause synaptotoxicity and potentially contribute to Alzheimer's disease (AD), the role of mature Abeta-fibrils in the amyloid plaques remains controversial. A widely held view in the field suggests that the fibrillization reaction proceeds 'forward' in a near-irreversible manner from the monomeric Abeta peptide through toxic protofibrillar intermediates, which subsequently mature into biologically inert amyloid fibrils that are found in plaques. Here, we show that natural lipids destabilize and rapidly resolubilize mature Abeta amyloid fibers. Interestingly, the equilibrium is not reversed toward monomeric Abeta but rather toward soluble amyloid protofibrils. We characterized these 'backward' Abeta protofibrils generated from mature Abeta fibers and compared them with previously identified 'forward' Abeta protofibrils obtained from the aggregation of fresh Abeta monomers. We find that backward protofibrils are biochemically and biophysically very similar to forward protofibrils: they consist of a wide range of molecular masses, are toxic to primary neurons and cause memory impairment and tau phosphorylation in mouse. In addition, they diffuse rapidly through the brain into areas relevant to AD. Our findings imply that amyloid plaques are potentially major sources of soluble toxic Abeta-aggregates that could readily be activated by exposure to biological lipids.     

Effect of weak static and low-frequency alternating magnetic fields on the fission and regeneration of the planarian dugesia (Girardia) tigrina

The effect of weak static (DC) and alternating (AC) magnetic fields (MFs), as well as combined (AC/DC) collinear MFs on the intensity of morphogenesis processes in the planarian Dugesia (Girardia) tigrina has been studied. It was found that combined MFs produce a stimulating effect on the fission and regeneration of planarians. Both components of the combined MFs, the direct (DC) and the alternating (AC), are important in the realization of the effects of weak MFs. The practically complete absence of one of the components (DC) reverses the sign of the effect. It was shown that the presence of concomitant background MFs does not substantially influence the effects of combined MFs with a very small AC component (100 nT). The effect of the "zero" field is significant and comparable in magnitude with the effects of combined MFs at effective frequencies. Narrow zones of effective amplitudes (in the region of tens and hundreds of nT) of the AC component of the combined MFs, with the DC component close to the value of the geomagnetic field were found, which alternate with regions where the response of the biological object to the influence is absent.     

Tjp3/zo-3 is critical for epidermal barrier function in zebrafish embryos

TJP3/ZO-3 is a scaffolding protein that tethers tight junction integral membrane proteins to the actin cytoskeleton and links the conserved Crumbs polarity complex to tight junctions. The physiological function of TJP3/ZO-3 is not known and mice lacking TJP3/ZO-3 show no apparent phenotype. Here we show that Tjp3/Zo-3 is a component of tight junctions present in the enveloping cell layer of zebrafish embryos. Silencing tjp3/zo-3 using morpholinos leads to edema, loss of blood circulation and tail fin malformations in the embryos. The ultrastructure of tight junctions of the enveloping cell layer is disrupted, without affecting the asymmetric distribution of plasma membrane proteins. Morphants show a loss of the epidermal barrier, as assessed by an increased permeability of the enveloping cell layer to low molecular weight tracers and a higher sensitivity of the embryos to osmotic stress. Subjecting wild-type embryos to osmotic stress mimicks the morphant phenotype, consistent with the phenotype being a direct consequence of failed osmoregulation. Thus, Tjp3/Zo-3 is critical for barrier function of the enveloping cell layer and osmoregulation in early stages of zebrafish development.     

Effects of neuropeptide F on regeneration in <Emphasis Type="Italic">Girardia tigrina</Emphasis> (Platyhelminthes)

The effects of neuropeptide F (NPF; from Moniezia expansa) on the regeneration of Girardia tigrina were studied. The animals were decapitated and incubated in water (control) or NPF. The dynamics of the proliferation of the neoblasts in the developing tissue were studied during the course of regeneration by monitoring the mitotic index (MI). The effects of incubation in FMRFamide and GYIRFamide on the MI were also tested. The course of cephalic regeneration was followed with in vivo computer-assisted morphometry for up to 7 days. The development of the regenerating nervous system and the musculature was visualised by immunostaining with a primary antiserum to the C-terminal decapeptide of NPF (YFAIIGRPRFa) and tetramethylrhodamine-isothiocyanate-conjugated phalloidin, which stains F-actin in muscle filaments. The study showed that NPF had a stimulatory effect on the mitotic activity of the neoblasts. FMRFamide and GYIRFamide did not have this effect. NPF also stimulated the growth of the regenerating head and the growing nervous system and musculature. NPF is postulated to have a morphogenetic action in the regenerating animals. This work was supported by two grants from the Finnish Academy of Science (nos. 202685, 2004) and (no. 112090, 2006) to M.G., an RFBR grant (07-04-00452a) to N.K. and a Wellcome Trust grant (069411) to A.G.M. for which we express our gratitude.     

A new UV-B absorbing mycosporine with photo protective activity from the lichenized ascomycete Collema cristatum.

A novel photo protective mycosporine was isolated from the lichenized ascomycete Collema cristatum. Biological activity was measured in terms of protection against UV-B induced membrane destruction and pyrimidine dimer formation in cultured human keratinocytes, and prevention of UV-B induced erythema. It was found that the pure isolated compound prevented UV-B induced cell destruction in a dose-dependent manner, that the compound partially prevented pyrimidine dimer formation and completely prevented UV-B induced erythema when applied to the skin prior to irradiation.     

N-glycosylation modulates the cell-surface expression and catalytic activity of corin

N-Glycosylation may influence the subcellular localization and biological activity of the pro-ANP convertase, corin. In HEK293-corin cells, the inhibition of N-glycosylation, with tunicamycin, reduced the cell-surface expression of murine corin, but did not alter the total expression. Therefore, tunicamycin treatment likely caused the intracellular accumulation of non-glycosylated corin. Tunicamycin treatment also significantly reduced corin activity (pro-ANP cleavage) in these cells. We developed an assay to measure the effect of N-glycosylation on corin activity, independent of its effect on corin localization. We determined that the reduction in corin activity was due to a direct effect of N-glycosylation, and was not secondary to the effect of N-glycosylation on corin cell-surface expression. Our data provide evidence that N-glycosylation is essential for the cell-surface expression of murine corin and modulates its functional activity. N-Glycosylation represents a possible mechanism for the regulation of native corin on the surface of cardiomyocytes.     

COP9 Limits Dendritic Branching via Cullin3-Dependent Degradation of the Actin-Crosslinking BTB-Domain Protein Kelch

Components of the COP9 signalosome (CSN), a key member of the conserved 26S proteasome degradation pathway, have been detected to be altered in patients of several debilitating syndromes. These findings suggest that CSN acts in neural circuits, but the exact function of CSN in brain remains unidentified. Previously, using Drosophila peripheral nervous system (PNS) as a model system, we determined that CSN is a critical regulator of dendritic morphogenesis. We found that defects in CSN led to the strikingly contrast phenotype of either reducing or stimulating dendritic branching. In particular, we have reported that CSN stimulates dendritic branching via Cullin1-mediated proteolysis. Here we describe that CSN inhibits dendritic arborization in PNS neurons acting via control of Cullin3 function: loss of Cullin3 causes excessive dendritic branching. We also identified a downstream target for Cullin3-dependent degradation in neurons – the actin-crosslinking BTB-domain protein Kelch. Inappropriate accumulation of Kelch, either due to the impaired Cullin3-dependent turnover, or ectopic expression of Kelch, leads to uncontrolled dendritic branching. These findings indicate that the CSN pathway modulates neuronal network in a multilayer manner, providing the foundation for new insight into the CSN role in human mental retardation disorders and neurodegenerative disease.