Leucine-derived cyano glucosides in barley

Barley (Hordeum vulgare) seedlings contain five cyano glucosides derived from the amino acid L-leucine (Leu). The chemical structure and the relative abundance of the cyano glucosides were investigated by liquid chromatography-mass spectrometry and nuclear magnetic resonance analyses using spring barley cultivars with high, medium, and low cyanide potential. The barley cultivars showed a 10-fold difference in their cyano glucoside content, but the relative content of the individual cyano glucosides remained constant. Epiheterodendrin, the only cyanogenic glucoside present, comprised 12% to 18% of the total content of cyano glucosides. It is proposed that the aglycones of all five cyano glucosides are formed by the initial action of a cytochrome P450 enzyme of the CYP79 family converting L-Leu into Z-3-methylbutanal oxime and subsequent action of a less specific CYP71E enzyme converting the oxime into 3-methylbutyro nitrile and mediating subsequent hydroxylations at the alpha-, as well as beta- and gamma-, carbon atoms. Presence of cyano glucosides in the barley seedlings was restricted to leaf tissue, with 99% confined to the epidermis cell layers of the leaf blade. Microsomal preparations from epidermal cells were not able to convert L-[(14)C]Leu into the biosynthetic intermediate, Z-3-methylbutanal-oxime. This was only achieved using microsomal preparations from other cell types in the basal leaf segment, demonstrating translocation of the cyano glucosides to the epidermal cell layers after biosynthesis. A beta-glucosidase able to degrade epiheterodendrin was detected exclusively in yet a third compartment, the endosperm of the germinating seed. Therefore, in barley, a putative function of cyano glucosides in plant defense is not linked to cyanide release.     

Responses of domestic chicks (Gallus gallus domesticus) to multimodal aposematic signals

Many aposematic prey combine their visual warning signals withadditional signals. Together, these signals constitute a multimodalor multicomponent warning display. The additional signals arethought to increase the effects of the visual signals on predators.Olfactory signals are much emphasized, but later studies haveshown that also auditory signals like the buzzing of certaininsects might have multimodal effects. The wasp displays typicalvisual aposematic signals, black and yellow stripes, but doesalso emit a characteristic buzzing. We wanted to test if, andin what way, the visual and acoustic display of the wasp hasan aversive function on the predators. We therefore conducteda 12-trial discrimination-learning task on inexperienced chicksto study whether there are innate biases toward these signalsand how they affect the speed of avoidance learning. We alsoperformed three extinction-learning trials to study how memorablethe signals were to the chicks. We show that the visual signalsin the display of the wasp contribute to the protection frompredators but in different ways; the yellow color had an aversiveeffect on inexperienced predators, while the striped patternimproved the aversion learning. The sound did not enhance theinnate aversions but increased the aversion learning of stripesin green prey.     

Ketogenic diet is antiepileptogenic in pentylenetetrazole kindled mice and decrease levels of N-acylethanolamines in hippocampus

The ketogenic diet (KD) is used for the treatment of refractory epilepsy in children, however, the mechanism(s) remains largely unknown. Also, the antiepileptogenic potential in animal models of epilepsy has been poorly addressed. Activation of cannabinoid type-1 receptor (CB₁-R) upon seizure activity may mediate neuroprotection as may several N-acylethanolamines. It is unknown how the KD interfere with the endocannabinoid system.We investigated the antiepileptogenic potential of the KD in the pentylenetetrazole kindling model in young mice and measured the hippocampal levels of CB₁-R by Western blot and of endocannabinoids and N-acylethanolamines by mass spectrometry.The KD significantly decreased incidence and severity of seizures, and significantly increased the latency to clonic convulsions. There were no changes in levels of endocannabinoids or CB₁-R expression by either seizure activity or type of diet. The level of oleoylethanolamide as well as the sum of N-acylethanolamines were significantly decreased by the KD, but were unaffected by seizure activity.The study shows that the KD had clear antiepileptogenic properties in the pentylenetetrazole kindling model and does not support a role of endocannabinoids in this model. The significance of the decreased hippocampal level of oleoylethanolamide awaits further studies.     

Phylogeny of Nerillidae (Polychaeta, Annelida) as inferred from combined 18S rDNA and morphological data

A phylogeny of the meiofaunal polychaete family Nerillidae based on morphological, molecular and combined data is presented here. The data sets comprise nearly complete sequences of 18S rDNA and 40 morphological characters of 17 taxa. Sequences were analyzed simultaneously with the morphological data by direct optimization in the program POY, with a variety of parameter sets (costs of gaps: transversions: transitions). Three outgroups were selected from the major polychaete group Aciculata and one from Scolecida. The 13 nerillid species from 11 genera were monophyletic in all analyses with very high support, and three new apomorphies for Nerillidae are identified. The topology of the ingroup varied according to the various parameter settings. Reducing the number of outgroups to one decreased the variance among the phylogenetic hypotheses. The congruence among these was tested and a parameter set, with equal weights (222) and extension gap weighted 1, yielded minimum incongruence (ILD). Several terminal clades of the combined analysis were highly supported, as well as the position of Leptonerilla prospera as sister terminal to the other nerillids. The evolution of morphological characters such as segment numbers, chaetae, appendages and ciliation are traced and discussed. A regressive pathway within Nerillidae is indicated for several characters, however, generally implying several convergent losses. Numerous genera are shown to require revision. © The Willi Hennig Society 2005.     

Smaller stomata require less severe leaf drying to close: A case study in Rosa hydrida

Stomata formed at high relative air humidity (RH) close less as leaf dries; an effect that varies depending on the genotype. We here quantified the contribution of each stomatal response characteristic to the higher water loss of high RH-grown plants, and assessed the relationship between response characteristics and intraspecific variation in stomatal size. Stomatal size (length multiplied by width), density and responsiveness to desiccation, as well as pore dimensions were analyzed in ten rose cultivars grown at moderate (60%) or high (85%) RH. Leaf morphological components and transpiration at growth conditions were also assessed. High growth RH resulted in thinner (11%) leaves with larger area. A strong positive genetic correlation of daytime and nighttime transpiration at either RH was observed. Stomatal size determined pore area (r = 0.7) and varied by a factor of two, as a result of proportional changes in length and width. Size and density of stomata were not related. Following desiccation, high RH resulted in a significantly lower (6–19%) decline of transpiration in three cultivars, whereas the relative water content (RWC) of high RH-expanded leaflets was lower (29–297%) in seven cultivars. The lower RWC of these leaflets was caused by (a) higher (33–72%) stable transpiration and/or (b) lower (12–143%) RWC at which this stable transpiration occurred, depending on the cultivar. Stomatal size was significantly correlated with both characteristics (r = 0.5 and -0.7, respectively). These results indicate that stomatal size explains much of the intraspecific variation in the regulation of transpiration upon water deprivation on rose.     

Claudin-15 and -25b expression in the intestinal tract of Atlantic salmon in response to seawater acclimation,smoltification and hormone treatment

In seawater fishes, osmotic homeostasis requires uptake of ions and water in the intestine and these processes are governed by the combined trans- and paracellular pathways. The current study examined mRNA expression of two tight junction proteins (claudin-15 and -25b) predominantly expressed in the intestine of Atlantic salmon. We examined the response in pyloric caecae, middle and posterior intestine to seawater challenge, during smoltification and after injection with osmoregulatory hormones. Seawater (SW) transfer elevated levels of claudin-15 and -25b while no change was induced throughout the smolt stage. In freshwater, cortisol and growth hormone inhibited claudin-15 expression in the two anterior segments. Claudin-25b was elevated in all intestinal segments by growth hormone, while cortisol had an inhibitory effect. In seawater, prolactin and cortisol inhibited claudin expression. The data suggest that claudin expression is involved in the reorganisation of intestinal epithelium and possibly change paracellular permeability during SW acclimation. The lack of preparatory change during smoltification suggests that this process is not completed during smolt development. The effects of the tested hormones cannot explain the sum of changes induced by salinity, which, like the smoltification data, suggests the importance of additional factors and possibly contact with the imbibed SW per se.     

A systematic study of site-specific GalNAc-type O-glycosylation modulating proprotein convertase processing

Site-specific GalNAc-type O-glycosylation is emerging as an important co-regulator of proprotein convertase (PC) processing of proteins. PC processing is crucial in regulating many fundamental biological pathways and O-glycans in or immediately adjacent to processing sites may affect recognition and function of PCs. Thus, we previously demonstrated that deficiency in site-specific O-glycosylation in a PC site of the fibroblast growth factor, FGF23, resulted in marked reduction in secretion of active unprocessed FGF23, which cause familial tumoral calcinosis and hyperostosis hyperphosphatemia. GalNAc-type O-glycosylation is found on serine and threonine amino acids and up to 20 distinct polypeptide GalNAc transferases catalyze the first addition of GalNAc to proteins making this step the most complex and differentially regulated steps in protein glycosylation. There is no reliable prediction model for O-glycosylation especially of isolated sites, but serine and to a lesser extent threonine residues are frequently found adjacent to PC processing sites. In the present study we used in vitro enzyme assays and ex vivo cell models to systematically address the boundaries of the region within site-specific O-glycosylation affect PC processing. The results demonstrate that O-glycans within at least ±3 residues of the RXXR furin cleavage site may affect PC processing suggesting that site-specific O-glycosylation is a major co-regulator of PC processing.     

Phylogeny and systematics of Aphroditiformia

Aphroditiformia represents one of the most successful radiations of annelids, and is therefore an interesting model to understand morphological and functional evolution. Previous phylogenetic analyses yielded most families as monophyletic but excluded anchialine and interstitial species while failing to recover relationships within Sigalionidae. Here we address these shortcomings through the analysis of four molecular markers and 87 morphological characters sampled across 127 species under the assumptions of parsimony and model‐based methods. Of the 34 newly sequenced taxa, five anchialine and 24 interstitial species were included, with increased representation of Sigalionidae. An additional 28 elusive Sigalionidae taxa were included, represented only by morphological partitions. Molecular and morphological partitions were evaluated under exhaustive sensitivity analyses, testing the effects of alignment algorithms and optimization criteria on tree topologies. Our trees congruently recovered six clades corresponding to the families within Aphroditiformia: Acoetidae, Aphroditidae, Eulepethidae, Iphionidae, Polynoidae and Sigalionidae, respectively. An anchialine polynoid lineage was nested among strictly deep sea species, and interstitial pisionids and pholoids formed two independent clades nested within Sigalionidae. Additionally, Sigalionidae resulted in four clades, defined by combinations of apomorphies, and hereby we propose the subfamilies Pelogeniinae, Pholoinae, Pisioninae, Sthenelanellinae, as well as the provisionally included polyphyletic Sigalioninae.