Ecological role of physical dormancy in seeds of Oxytropis racemosa in a semiarid sandland with unpredictable rainfall

Aims Seed dormancy and the soil seed bank are crucial to plant regeneration strategy, especially in semiarid ecosystems with unpredictable precipitation. The aim of this study was to investigate how seed dormancy is controlled by environmental factors and how it is correlated with the soil seed bank and regeneration of the perennial legume Oxytropis racemosa, a dominant perennial herb in Mu Us Sandland of semiarid China.Methods Germination and imbibition experiments on fresh intact and scarified seeds of O. racemosa were used to identify physical dormancy (PY) in seeds of this species. Soil seed bank dynamics, timing of seedling emergence and the fate of buried seeds in the natural habitat were investigated.Important findings PY was broken by mechanical scarification or wet heat/ice water cycles but not solely by dry heat or wet heat treatment. The soil seed bank exhibited seasonal changes in the number of seeds, which was highest in September and lowest in July. Seeds buried at different sand depths gradually lost dormancy; 20–42% of the seeds remained dormant after 20 months of burial. Dormancy break occurs gradually throughout the year. Our results indicate that O. racemosa exhibits hardcoatedness heterogeneity that spreads germination of a seed cohort between seasons and years in the semiarid environment, where the amount of precipitation during the growing season is highly variable.     

Mutations in a Novel CLN6-Encoded Transmembrane Protein Cause Variant Neuronal Ceroid Lipofuscinosis in Man and Mouse

The CLN6 gene that causes variant late-infantile neuronal ceroid lipofuscinosis (vLINCL), a recessively inherited neurodegenerative disease that features blindness, seizures, and cognitive decline, maps to 15q21-23. We have used multiallele markers spanning this approximately 4-Mb candidate interval to reveal a core haplotype, shared in Costa Rican families with vLINCL but not in a Venezuelan kindred, that highlighted a region likely to contain the CLN6 defect. Systematic comparison of genes from the minimal region uncovered a novel candidate, FLJ20561, that exhibited DNA sequence changes specific to the different disease chromosomes: a G-->T transversion in exon 3, introducing a stop codon on the Costa Rican haplotype, and a codon deletion in exon 5, eliminating a conserved tyrosine residue on the Venezuelan chromosome. Furthermore, sequencing of the murine homologue in the nclf mouse, which manifests recessive NCL-like disease, disclosed a third lesion-an extra base pair in exon 4, producing a frameshift truncation on the nclf chromosome. Thus, the novel approximately 36-kD CLN6-gene product augments an intriguing set of unrelated membrane-spanning proteins, whose deficiency causes NCL in mouse and man.     

Lateral clustering of platelet GP Ib-IX complexes leads to up-regulation of the adhesive function of integrin alpha IIbbeta 3.

Binding of von Willebrand factor (VWF) to GP Ib-IX mediates initial platelet adhesion and increases the subsequent adhesive function of alpha(IIb)beta(3). Because these responses are promoted most effectively by large VWF multimers, we hypothesized that receptor clustering modulates GP Ib-IX function. To test this, GP IX was fused at its cytoplasmic tail to tandem repeats of FKBP, and GP Ib-IX(FKBP)(2) and alpha(IIb)beta(3) were expressed in Chinese hamster ovary cells. Under flow conditions at wall shear rates of up to 2000 s(-1), GP Ib-IX(FKBP)(2) mediated cell tethering to immobilized VWF, just as in platelets. Conditional oligomerization of GP Ib-IX(FKBP)(2) by AP20187, a cell-permeable FKBP dimerizer, caused a decrease in cell translocation velocities on VWF (p < 0.001). Moreover, clustering of GP Ib-IX(FKBP)(2) by AP20187 led to an increase in alpha(IIb)beta(3) function, manifested under static conditions by increased cell adhesion to fibrinogen (p < 0.01) and under flow by increased stable cell adhesion to VWF (p < 0.04). Clustering of GP Ib-IX(FKBP)(2) also stimulated rapid tyrosine phosphorylation of ectopically expressed Syk, a putative downstream effector of GP Ib-IX in platelets. These studies establish that GP Ib-IX oligomerization, per se, affects the interaction of this receptor with VWF and its ability to influence the adhesive function of alpha(IIb)beta(3). By extrapolation, GP Ib-IX clustering in platelets may promote thrombus formation.     

Scent Marking in Voles: A Reassessment of Over Marking, Counter Marking, and Self-Advertisement

We conducted a series of experiments to discern among the counter-marking, over-marking, and self-advertisement hypotheses for secondary marking in male prairie voles, Microtus ochrogaster , and meadow voles, M. pennsylvanicus . Secondary scent marks (those placed in an area that has already been marked by a conspecific) were not significantly greater than initial marks placed on clean substrate (a substrate without any previous scent marks) for either species and thus did not support a counter-marking hypothesis. Similarly, overlapping of initial scent marks with secondary marks occurred less often than expected by chance and did not support an over-marking hypothesis. Secondary marks tended to avoid overlap with scent marks previously deposited by a potential competitor. Our results suggest that secondary scent marking functions to self-advertise by maximizing individual identity and avoiding masking or blending with previous donors. Future studies on secondary marking should be designed to quantify the observed and expected frequency and placement of original and secondary marks to discern among alternative hypotheses for the adaptive significance of secondary marking.     

The protein SET binds the neuronal Cdk5 activator p35nck5a and modulates Cdk5/p35nck5a activity.

The neuronal Cdk5 kinase is composed of the catalytic subunit Cdk5 and the activator protein p35(nck5a) or its isoform, p39(nck5ai). To identify novel p35(nck5a)- and p39(nck5ai)-binding proteins, fragments of p35(nck5a) and p39(nck5ai) were utilized in affinity isolation of binding proteins from rat brain homogenates, and the isolated proteins were identified using mass spectrometry. With this approach, the nuclear protein SET was shown to interact with the N-terminal regions of p35(nck5a) and p39(nck5ai). Our detailed characterization showed that the SET protein formed a complex with Cdk5/p35(nck5a) through its binding to p35(nck5a). The p35(nck5a)-interacting region was mapped to a predicted alpha-helix in SET. When cotransfected into COS-7 cells, SET and p35(nck5a) displayed overlapping intracellular distribution in the nucleus. The nuclear co-localization was corroborated by immunostaining data of endogenous SET and Cdk5/p35(nck5a) from cultured cortical neurons. Finally, we demonstrated that the activity of Cdk5/p35(nck5a), but not that of Cdk5/p25(nck5a), was enhanced upon binding to the SET protein. The tail region of SET, which is rich in acidic residues, is required for the stimulatory effect on Cdk5/p35(nck5a).     

Attenuation of baroreflex sensitivity after domoic acid lesion of the nucleus ambiguus of rats.

The nucleus ambiguus (NA) and the dorsal motor nucleus of the vagus (DmnX) innervate distinct populations of cardiac ganglionic principal neurons. This anatomic evidence suggests that these two nuclei play different roles (Cheng Z and Powley TL, Soc Neurosci Abstr 26: 1189, 2000). However, lesion of the DmnX does not attenuate baroreflex sensitivity (Cheng Z, Guo SZ, Lipton AJ, and Gozal D, J Neurosci 22: 3215-3226, 2002). The present study tested the functional role of the NA in baroreflex control of heart rate (HR). Domoic acid (DA) was injected into the left NA of Sprague-Dawley rats to lesion the NA. The neuronal loss was assessed using retrograde labeling and confocal microscopy. HR changes induced by phenylephrine and sodium nitroprusside administration and after electrical stimulation of the left vagal trunk were measured at 15 days, and HR responses to left NA microinjection of L-glutamate were determined at 180 days postlesion. Compared with vehicle injections, DA lesions significantly reduced the population of NA motor neurons by approximately 68% (P < 0.01) and attenuated baroreflex sensitivity by approximately 83% (P < 0.01) at 15 days. Similarly, electrical stimulation of the vagal trunk of DA-lesioned animals led to attenuated decreases in HR responses. NA neuronal counts were reduced by approximately 81% (P < 0.01) and mean HR responses to l-glutamate injection into the lesioned NA were attenuated by approximately 65% (P < 0.01) at 180 days. Therefore, the NA plays a major role in baroreflex control of HR, and the integrity of the NA is critically important for the normal baroreflex control. In addition, NA lesions produce long-term anatomic and functional dysfunction of the nucleus, and thus it may provide an useful model for functional assessment of respective roles of the NA and DmnX.     

Molecular genetic linkage maps for allotetraploid Leymus wildryes (Gramineae: Triticeae).

Molecular genetic maps were constructed for two full-sib populations, TTC1 and TTC2, derived from two Leymus triticoides x Leymus cinereus hybrids and one common Leymus triticoides tester. Informative DNA markers were detected using 21 EcoRI-MseI and 17 PstI-MseI AFLP primer combinations, 36 anchored SSR or STS primer pairs, and 9 anchored RFLP probes. The 164-sib TTC1 map includes 1069 AFLP markers and 38 anchor loci in 14 linkage groups spanning 2001 cM. The 170-sib TTC2 map contains 1002 AFLP markers and 36 anchor loci in 14 linkage groups spanning 2066 cM. Some 488 homologous AFLP loci and 24 anchor markers detected in both populations showed similar map order. Thus, 1583 AFLP markers and 50 anchor loci were mapped into 14 linkage groups, which evidently correspond to the 14 chromosomes of allotetraploid Leymus (2n = 4x = 28). Synteny of two or more anchor markers from each of the seven homoeologous wheat and barley chromosomes was detected for 12 of the 14 Leymus linkage groups. Moreover, two distinct sets of genome-specific STS markers were identified in these allotetraploid Leymus species. These Leymus genetic maps and populations will provide a useful system to evaluate the inheritance of functionally important traits of two divergent perennial grass species.     

Nitrogen oxide emissions following wetting of dry soils in forest and pastures in Rondônia, Brazil

Rains at the end of the dry season can trigger increases in emissions of nitric oxide (NO) and nitrous oxide from forest and pasture soils in the Amazon Basin. The relative importance of the rain-stimulated emissions in the seasonal and annual budgets of these nitrogen gases for forests and pastures in the western Amazon is not well established. We measured soil emissions of NO and N₂O from a forest and two pastures, 11 and 26 years old, after a simulated rain event. Wetting the soil resulted in very small pulses of NO or N₂O from forest soils and no significant NO or N₂O pulses from the pastures. We estimated that in the forest, the amounts of each gas emitted from pulses during the dry to wet transition period represented 3.4% of the NO and 1.8% of the N₂O dry-season emissions, but amounted to less than 2% of the annual emissions of either gas. Total N oxide emissions of 5.6 kg N/ha/yr from the forest were nearly evenly divided between NO (42%) and N₂O (58%). The emissions of NO were evenly distributed over the wet and dry seasons, while over 84% N₂O fluxes occurred during the wet season.     

An insertion sequence unique to Frankia strain ArI5

At the genetic level, understanding of symbiotic nitrogen fixation by Frankia is limited to nif functions that are highly conserved among all organisms. The genetics and biochemistry of nodulation are largely unexplored because of a complete lack of genetic tools. In other bacteria, mobile genetic elements such as insertion sequences (IS) and transposons are commonly used to create mutations and insert new genetic material. We have characterized a 4 kbp segment of DNA from Frankia strain ArI5 that has the hallmarks of a mobile genetic element, inverted repeats flanking a gene encoding a transposase. There are at least six copies of this element in strain ArI5 but none in either strain CcI3 or CpI1. The inverted repeats are 17 nt long and separated by 2156 bp. Within that region are two, overlapping ORFs that each encode a transposase. RT-PCR analysis of RNA from Frankia ArI5 cells conclusively demonstrates the expression of one transposase gene and suggests that both may be transcribed. Numerous attempts to clone the intact IS in E. coli were unsuccessful suggesting that the element may be unstable in this context. A clone containing the complete IS was constructed in E. coli then modified by insertion of the kanamycin (KAN) resistance gene from Tn5. A fragment of DNA including the inverted repeats, transposase genes and KAN gene, was transferred to the suicide vector pJBSD1. The construct, pFRISK, was transformed into E. coli to search for transposition events.     

Profiling with structural elucidation of the neutral and anionic O-linked oligosaccharides in the egg jelly coat of Xenopus laevis by Fourier transform mass spectrometry

A strategic method with high speed and sensitivity is outlined for the analysis of mucin-type oligosaccharide from the jelly coat of Xenopus laevis. The method relies primarily on mass spectrometric techniques, in this case matrix-assisted laser desorption/ionization Fourier-transform mass spectrometry (MALDI-FTMS) and collision-induced dissociation (CID). Separation with isolation of the oligosaccharides was streamlined to couple well with mass spectrometry allowing the rapid determination of all detectable components from both neutral and anionic species. Partial structures of anionic components, composed primarily of sulfate esters, were obtained with CID. For neutral species, a method that allowed the complete structural determination using mass spectrometry was used. The method builds on the structure of small number of known compounds to determine unknown structures from the same biological source. In this example, a small number of oligosaccharides, elucidated previously by NMR, were used to develop a set of substructural motifs that were characterized by CID. The presence of the motifs in the CID spectra were then used to determine the structures of unknown compounds that were in abundances too small for NMR analysis.