Differential expression of rat beta-defensins

The innate immunity, utilizes a battery of broad-spectrum antibacterial cationic polypeptides (3-5 kDa) named alpha- and beta-defensins. Several beta-defensins have been isolated and shown to play a role in the defense of various tissues. Herein, we report the expression pattern of two rat beta-defensins, rBD-1 and rBD-2, in liver, kidney, lung, spleen, and brain using RT-PCR. To study polymorphism and verify gene identity, all cDNA products were sequenced. rBD-1 was expressed in the kidney, lung, brain, but not in spleen or liver, whereas rBD-2 was expressed in the lung, but not in the kidney or spleen. In addition, rBD-2 was expressed in the brain and liver. No polymorphism was found in the genes encoding rat beta-defensins. These findings demonstrate a different expression pattern for rBD-2 than what has been reported. We conclude that the rat may be a useful model to investigate the function and contribution of beta-defensins to host defense.     

Buoyancy studies in natural communities of square gas-vacuolate archaea in saltern crystallizer ponds

Background

Possession of gas vesicles is generally considered to be advantageous to halophilic archaea: the vesicles are assumed to enable the cells to float, and thus reach high oxygen concentrations at the surface of the brine.

Results

We studied the possible ecological advantage of gas vesicles in a dense community of flat square extremely halophilic archaea in the saltern crystallizer ponds of Eilat, Israel. We found that in this environment, the cells' content of gas vesicles was insufficient to provide positive buoyancy. Instead, sinking/floating velocities were too low to permit vertical redistribution.

Conclusion

The hypothesis that the gas vesicles enable the square archaea to float to the surface of the brines in which they live was not supported by experimental evidence. Presence of the vesicles, which are mainly located close to the cell periphery, may provide an advantage as they may aid the cells to position themselves parallel to the surface, thereby increasing the efficiency of light harvesting by the retinal pigments in the membrane.     

Did backcrossing contribute to the origin of hybrid edible bananas?

Background

Bananas and plantains (Musa spp.) provide a staple food for many millions of people living in the humid tropics. The cultivated varieties (cultivars) are seedless parthenocarpic clones of which the origin remains unclear. Many are believed to be diploid and polyploid hybrids involving the A genome diploid M. acuminata and the B genome M. balbisiana, with the hybrid genomes consisting of a simple combination of the parental ones. Thus the genomic constitution of the diploids has been classified as AB, and that of the triploids as AAB or ABB. However, the morphology of many accessions is biased towards either the A or B phenotype and does not conform to predictions based on these genomic formulae.

Scope

On the basis of published cytotypes (mitochondrial and chloroplast genomes), we speculate here that the hybrid banana genomes are unbalanced with respect to the parental ones, and/or that inter-genome translocation chromosomes are relatively common. We hypothesize that the evolution under domestication of cultivated banana hybrids is more likely to have passed through an intermediate hybrid, which was then involved in a variety of backcrossing events. We present experimental data supporting our hypothesis and we propose a set of experimental approaches to test it, thereby indicating other possibilities for explaining some of the unbalanced genome expressions. Progress in this area would not only throw more light on the origin of one of the most important crops, but provide data of general relevance for the evolution under domestication of many other important clonal crops. At the same time, a complex origin of the cultivated banana hybrids would imply a reconsideration of current breeding strategies.     

Nud1p, the yeast homolog of Centriolin, regulates spindle pole body inheritance in meiosis

Nud1p, a protein homologous to the mammalian centrosome and midbody component Centriolin, is a component of the budding yeast spindle pole body (SPB), with roles in anchorage of microtubules and regulation of the mitotic exit network during vegetative growth. Here we analyze the function of Nud1p during yeast meiosis. We find that a nud1-2 temperature-sensitive mutant has two meiosis-related defects that reflect genetically distinct functions of Nud1p. First, the mutation affects spore formation due to its late function during spore maturation. Second, and most important, the mutant loses its ability to distinguish between the ages of the four spindle pole bodies, which normally determine which SPB would be preferentially included in the mature spores. This affects the regulation of genome inheritance in starved meiotic cells and leads to the formation of random dyads instead of non-sister dyads under these conditions. Both functions of Nud1p are connected to the ability of Spc72p to bind to the outer plaque and half-bridge (via Kar1p) of the SPB.     

Change in sugar, sterol and fatty acid composition in banana meristems caused by sucrose-induced acclimation and its effects on cryopreservation

To understand the mechanisms of sucrose‐induced acclimation in relation to plant cryopreservation, sugars, sterols, fatty acids of different lipid fractions (neutral lipids, glycolipids and sphingolipids and phospholipids), as well as free fatty acids were analyzed in proliferating meristem cultures of different banana varieties. The four banana varieties that were selected show different post‐thaw shoot regeneration rates (0–53.4%). All mentioned parameters were analyzed using (1) control meristems that were cultured on a normal sucrose concentration (0.09 M), which resulted in low survival after cryopreservation; and (2) 2‐week sucrose precultured meristems (0.4 M). This sucrose preculture, essential for regeneration after cryopreservation, resulted in a significant increase of each of seven sugars detected. The ratio of stigmasterol/sitosterol (St/Si) in sucrose‐pretreated meristems significantly increased. The sucrose pretreatment also resulted in a significant increase of total fatty acid content of the neutral lipid fraction and of the glycolipid and sphingolipid fraction, as well as the total free fatty acid content. The individual fatty acid content of the phospholipids was differently changed by the sucrose pretreatment for the given varieties studied. In most cases, sucrose pretreatment resulted in an increase of the double bond index (DBI) in the neutral lipids and a decrease of DBI in the glycolipids and sphingolipids, in phospholipids as well as in free fatty acids. Principal component analysis of all collected data revealed that (1) for the control material, sucrose and total sugar contents were closely linked to the post‐thaw shoot regeneration, suggesting that sucrose and total sugar may be main limiting factors to survive cryopreservation; (2) accumulation of large quantities of sugars (glucose, fructose, sucrose and total sugar) in sucrose‐pretreated material cannot explain the differences in survival after cryopreservation of the four banana varieties. We assume that a minimal amount of sugars is needed in meristem cultures to survive cryopreservation. Still, other limiting factors do influence the survival following the sucrose pretreatment. We observed that the parameters which are closely linked to the post‐thaw shoot regeneration are a minimal change in the ratios of St/Si, the minimal change of the DBI of phospholipids and free fatty acids, as well as linoleic acid content (C18:2); and (3) inositol, raffinose, myristic acid (C14:0) and oleic acid (C18:1) were present in small quantities; however, they could be correlated to survival after cryopreservation, suggesting that they may be also involved in cryopreservation process.     

Long-lived alphaMUPA transgenic mice exhibit pronounced circadian rhythms

Robust biological rhythms have been shown to affect life span. Biological clocks can be entrained by two feeding regimens, restricted feeding (RF) and caloric restriction (CR). RF restricts the time of food availability, whereas CR restricts the amount of calories with temporal food consumption. CR is known to retard aging and extend life span of animals via yet-unknown pathways. We hypothesize that resetting the biological clock could be one possible mechanism by which CR extends life span. Because it is experimentally difficult to uncouple calorie reduction from temporal food consumption, we took advantage of the murine urokinase-like plasminogen activator (alphaMUPA) transgenic mice overexpressing a serine protease implicated in brain development and plasticity; they exhibit spontaneously reduced eating and increased life span. Quantitative real-time PCR analysis revealed that alphaMUPA mice exhibit robust expression of the clock genes mPer1, mPer2, mClock, and mCry1 but not mBmal1 in the liver. We also found changes in the circadian amplitude and/or phase of clock-controlled output systems, such as feeding behavior, body temperature, and enteric cryptdin expression. A change in the light-dark regimen led to modified clock gene expression and abrogated circadian patterns of food intake in wild-type (WT) and alphaMUPA mice. Consequently, food consumption of WT mice increased, whereas that of alphaMUPA mice remained the same, indicating that reduced food intake occurs upstream and independently of the biological clock. Thus we surmise that CR could lead to pronounced and synchronized biological rhythms. Because the biological clock controls mitochondrial, hormonal, and physiological parameters, system synchronicity could lead to extended life span.     

Caveolin-1 and dynamin-2 are essential for removal of the complement C5b-9 complex via endocytosis

The complement system, an important element of both innate and adaptive immunity, is executing complement-dependent cytotoxicity (CDC) with its C5b-9 protein complex that is assembled on cell surfaces and transmits to the cell death signals. In turn, cells, and in particular cancer cells, protect themselves from CDC in various ways. Thus, cells actively remove the C5b-9 complexes from their plasma membrane by endocytosis. Inhibition of clathrin by transfection with shRNA or of EPS-15 with a dominant negative plasmid had no effect on C5b-9 endocytosis and on cell death. In contrast, inhibition of caveolin-1 (Cav-1) by transfection with an shRNA or a dominant negative plasmid sensitized cells to CDC and inhibited C5b-9 endocytosis. Similarly, both inhibition of dynamin-2 by transfection with a dominant negative plasmid or by treatment with Dynasore reduced C5b-9 endocytosis and enhanced CDC. C5b-9 endocytosis was also disrupted by pretreatment of the cells with methyl-β-cyclodextrin or Filipin III, hence implicating membrane cholesterol in the process. Analyses by confocal microscopy demonstrated co-localization of Cav-1-EGFP with C5b-9 at the plasma membrane, in early endosomes, at the endocytic recycling compartment and in secreted vesicles. Further investigation of the process of C5b-9 removal by exo-vesiculation demonstrated that inhibition of Cav-1 and cholesterol depletion abrogated C5b-9 exo-vesiculation, whereas, over-expression of Cav-1 increased C5b-9 exo-vesiculation. Our results show that Cav-1 and dynamin-2 (but not clathrin) support cell resistance to CDC, probably by facilitating purging of the C5b-9 complexes by endocytosis and exo-vesiculation.