雄性弯角长角羚和普通大羚羊激素分泌模式与社会等级和季节的关系

为了确定羚羊类固醇激素分泌模式与行为和社群等级的关系,我们检测了成年雄性弯角长角羚(n=15)和普通大羚羊(n=11)的粪样皮质酮和睾酮浓度.采用随意取样和目标取样法记录了共650 h的行为.结果发现,两种羚羊的睾酮水平在雌性的发情时段里都是最高的.两种羚羊皮质激素水平有季节性变化,表现为雨季皮质酮水平高于干旱季节.优势胁迫作用明显存在于普通大羚羊,而在弯角长角羚不明显.没有证据表明从属个体压力的存在,一只弯角长角羚曾经是优势个体,但是后来有两年不是优势个体,与其他雄羚相比,这只雄羚在雌性发情期的睾酮和皮质酮水平有3个交迭出现的峰值.     

Parallel topology of genetically fused EmrE homodimers

EmrE is a small H+-coupled multidrug transporter in Escherichia coli. Claims have been made for an antiparallel topology of this homodimeric protein. However, our own biochemical studies performed with detergent-solubilized purified protein support a parallel topology of the protomers. We developed an alternative approach to constrain the relative topology of the protomers within the dimer so that their activity can be assayed also in vivo before biochemical handling. Tandem EmrE was built with two identical monomers genetically fused tail to head (C-terminus of the first to N-terminus of the second monomer) with hydrophilic linkers of varying length. All the constructs conferred resistance to ethidium by actively removing it from the cytoplasm. The purified proteins bound substrate and transported methyl viologen into proteoliposomes by a proton-dependent mechanism. A tandem where one of the essential glutamates was replaced with glutamine transported only monovalent substrates and displayed a modified stoichiometry. The results support a parallel topology of the protomers in the functional dimer. The implications regarding insertion and evolution of membrane proteins are discussed.     

Identification of extracellular signal-regulated kinase 1/2 and p38 MAPK as regulators of human sperm motility and acrosome reaction and as predictors of poor spermatozoan quality

Mature spermatozoa acquire progressive motility only after ejaculation. Their journey in the female reproductive tract also includes suppression of progressive motility, reactivation, capacitation, and hyperactivation of motility (whiplash), the mechanisms of which are obscure. MAPKs are key regulatory enzymes in cell signaling, participating in diverse cellular functions such as growth, differentiation, stress, and apoptosis. Here we report that ERK1/2 and p38 MAPK are primarily localized to the tail of mature human spermatozoa. Surprisingly, c-Jun N-terminal kinase 1/2, which is thought to be ubiquitously expressed, could not be detected in mature human spermatozoa. ERK1/2 stimulation is downstream to protein kinase C (PKC) activation, which is also present in the human sperm tail (PKCbetaI and PKCepsilon). ERK1/2 stimulates and p38 inhibits forward and hyperactivated motility, respectively. Both ERK1/2 and p38 MAPK are involved in the acrosome reaction. Using a proteomic approach, we identified ARHGAP6, a RhoGAP, as an ERK substrate in PMA-stimulated human spermatozoa. Inverse correlation was obtained between the relative expression level of ERK1 or the relative activation level of p38 and sperm motility, forward progression motility, sperm morphology, and viability. Therefore, increased expression of ERK1 and activated p38 can predict poor human sperm quality.     

Cytosolic phospholipase A2alpha is targeted to the p47phox-PX domain of the assembled NADPH oxidase via a novel binding site in its C2 domain

We have previously demonstrated a physical interaction between cytosolic phospholipase A2alpha (cPLA2) and the assembled NADPH oxidase on plasma membranes following neutrophil stimulation. The aim of the present study was to define the exact binding sites between these two enzymes. Here we show, based on blot overlay experiments, F?rster resonance energy transfer analysis and studies in neutrophils from patients with chronic granulomatous disease deficient in p67phox or p47phox, that cPLA2 specifically binds to p47phox and that p47phox is sufficient to anchor cPLA2 to the assembled oxidase on the plasma membranes upon stimulation. Blot overlay and affinity binding experiments using subfragments of cPLA2 and p47phox demonstrated that the cPLA2-C2 domain and the p47phox-PX domain interact to form a complex that is resistant to high salt. Computational docking was used to identify hydrophobic peptides within these two domains that inhibited the association between the two enzymes and NADPH oxidase activity in electro-permeabilized neutrophils. These results were used in new docking computations that produced an interaction model. Based on this model, cPLA2-C2 domain mutations were designed to explore its interaction p47phox in neutrophil lysates. The triple mutant F35A/M38A/L39A of the cPLA2-C2 domain caused a slight inhibition of the affinity binding to p47phox, whereas the single mutant I67A was highly effective. The double mutant M59A/H115A of the p47phox-PX domain caused a significant inhibition of the affinity binding to cPLA2. Thus, Ile67 of the cPLA2-C2 domain is identified as a critical, centrally positioned residue in a hydrophobic interaction in the p47phox-PX domain.     

Multi-annual changes in the parasite communities of rabbitfish Siganus rivulatus (Siganidae) in the Gulf of Aqaba, Red Sea

The parasite communities of the rabbitfish, Siganus rivulatus, were used to track multi-annual changes in the northern Gulf of Aqaba, Red Sea, in an environment subjected to ongoing anthropogenic impact. Parasitological data from these fish were collected from 1998 to 2000, with spring and fall samplings at three locations: at a coral reef (OBS), at a sandy beach area (NB) and at a mariculture cage farm (FF). These data were compared with data from 1995–1997 as well as data collected during 1981–1985 at the coral reef sampling site. The data analyses indicate that the ratio between heteroxenous and monoxenous parasite species declined significantly at all sites between 1995–1997 and 1998–2000. During the same period, the species richness of monoxenous parasites increased significantly at all sites. The species richness of heteroxenous parasites decreased significantly at the coral reef site, but remained steady at the other two sites. This coincided with a significant increase in the prevalence of monogeneans at the OBS and FF sites and a significant decrease in the prevalence of digeneans at the FF and NB sites. The decline in the abundance of the latter, specifically of Opisthogonoporoides sp. and Gyliauchen sp., was even more significant when compared with the 1981–1985 data. The prevalence of other gut helminths, namely the digenean Hexangium sigani and the nematodes Cucullanus sigani and Procamallanus elatensis, however, showed a significant increase over the same period. Analysis of the species richness and diversity indices of the parasite communities did not reveal conspicuous differences. These, however, did become apparent when heteroxenous and monoxenous members of particular taxa were analyzed separately. Therefore, when using parasite assemblages to detect ecological changes, it is essential to analyze not only at the community level, but also to consider separate components of particular parasitic groups. Communicated by H. von Westernhagen, A. Diamant     

Vaccination with empty plasmid DNA or CpG oligonucleotide inhibits diabetes in nonobese diabetic mice: modulation of spontaneous 60-kDa heat shock protein autoimmunity

Nonobese diabetic (NOD) mice develop insulitis and diabetes through a process involving autoimmunity to the 60-kDa heat shock protein (HSP60). Treatment of NOD mice with HSP60 or with peptides derived from HSP60 inhibits this diabetogenic process. We now report that NOD diabetes can be inhibited by vaccination with a DNA construct encoding human HSP60, with the pcDNA3 empty vector, or with an oligonucleotide containing the CpG motif. Prevention of diabetes was associated with a decrease in the degree of insulitis and with down-regulation of spontaneous proliferative T cell responses to HSP60 and its peptide p277. Moreover, both the pcDNA3 vector and the CpG oligonucleotide induced specific Abs, primarily of the IgG2b isotype, to HSP60 and p277, and not to other islet Ags (glutamic acid decarboxylase or insulin) or to an unrelated recombinant Ag expressed in bacteria (GST). The IgG2b isotype of the specific Abs together with the decrease in T cell proliferative responses indicate a shift of the autoimmune process to a Th2 type in treated mice. These results suggest that immunostimulation by bacterial DNA motifs can modulate spontaneous HSP60 autoimmunity and inhibit NOD diabetes.