The nucleolus and transcription of ribosomal genes

Ribosome biogenesis is a highly dynamic, steady-state nucleolar process that involves synthesis and maturation of rRNA, its transient interactions with non-ribosomal proteins and RNPs and assembly with ribosomal proteins. In the few years of the 21st century, an exciting progress in the molecular understanding of rRNA and ribosome biogenesis has taken place. In this review, we discuss the recent results on the regulation of rRNA synthesis in relation to the functional organization of the nucleolus, and put an emphasis on the situation encountered in mammalian somatic cells.     

Artificial sowing of endangered dry grassland species into disused basalt quarries

Steppe-like dry (semi) natural grasslands are valuable and endangered habitats in central Europe. In the study area (the ?eské st?edoho?í Hills, Czech Republic), they occur in fragments on southern slopes of volcanic hills, often in the vicinity of quarries, which are numerous there. We performed sowing experiments and observed seedling recruitment of six species, typical for the dry grasslands, in 9 basalt quarries located in 3 different climatic regions over 3 yr. The sowing experiments were established in young successional stages, 5–12 yr after quarrying was stopped. The objectives of the experiments were to evaluate: (i) if seedling recruitment and survival of the target species are possible regarding different climatic regions and (ii) what is the influence of weather fluctuations among years on seedling survival. Seedlings of all studied species were able to recruit and survive in the early successional stages at least in some quarries. The species, except one, showed significant differences in recruitment among the climatic regions, with the best recruitment and survival in the driest and warmest region. Seedlings of two species did not recruit in the wettest and coldest region. All species survived in the driest and warmest region, while only one did so in the coldest and wettest region. Different weather conditions in the studied years significantly influenced recruitment of two species. The number of localities (floristic records) of particular species in the regions was the best predictor of species germination and survival. Thus a traditional floristic survey may help to predict success of species in restoration projects. It emerged that artificial sowing can be considered in restoration programs as a way of contributing to restoration of dry grasslands in disused quarries.     

Ecological Succession of a Relict Central European Peat Bog and Variability of Its Insect Biodiversity

The isolated habitat of the ervené Blato bog (South Bohemia, Czech Republic) and its relict insect fauna have been the subject of long-term monitoring. The species composition and abundance of Lepidoptera (light traps) and Coleoptera (pitfall traps) were monitored for 4 years (1994–1997) simultaneously on two sites – in the edaphic climax pine forest and in wetland successional habitats. The method of statistical evaluation by RDA and CCA ordination, representing the habitat preference of species of Coleoptera (Carabidae only) and Lepidoptera (all nocturnal phototactic taxa) between the edaphic climax forest and succession stages, was used. All categories of the peatland taxa (tyrphobiontic, tyrphophilous and tyrphoneutral species) were analysed. Ten highly stenotopic tyrphobiontic species and 23 tyrphophilous species of Lepidoptera (out of 487) were most characteristic of the bog habitat. Only two tyrphophilous carabid species (out of 20) were characteristic of the bog. The most important relict species (tyrphobionts) of Lepidoptera are most diverse and abundant within the successional habitats and in the open wet forest. The relict fauna of the closed climax pine forest is much less diverse and composed mostly of abundant tyrphophilous and tyrphoneutral forest species. Preservation or restoration of sufficiently constant hydrological conditions, which prevents formation of the closed forest, is the basic management for habitat conservation of all relict tyrphobiontic species of the ervené Blato bog and similar peat land habitat islands. The peat bog is a unified complex system of specific diverse and relict taxa. The most specific taxa are tyrphobiontic Lepidoptera, but a number of other vulnerable tyrphophilous and tyrphoneutral insects are associated with the peat bog as well.     

Rhizobial nod gene-inducing activity in pea nodulation mutants: dissociation of nodulation and flavonoid response

With the characterization of the total genomes of Arabidopsis thaliana and Oryza sativa , several putative plasma membrane components have been identified. However, a lack of knowledge at the protein level, especially for hydrophobic proteins, have hampered analyses of physiological changes. To address whether protein complexes may be present in the native membrane, we subjected plasma membranes isolated from Spinacia oleracea leaves to blue-native polyacrylamide gel electrophoresis (BN-PAGE). BN-PAGE is well established in the separation of functional membrane protein complexes from mitochondria and chloroplasts, but a resolved protein complex pattern from PM of eukaryotic cells has previously not been reported. Using this method, protein complexes from Spinacia oleracea PM could be efficiently solubilized and separated, including the highly hydrophobic aquaporin (apparent molecular mass 230 kDa), a putative tetramer of H⁺-ATPase, and several less abundant complexes with apparent masses around or above 750 kDa. After denaturation and separation of the complexes into their subunits in a second dimension (SDS-PAGE), several of the complexes were identified as hydrophobic membrane proteins. Large amounts of protein (up to 1 mg) can be resolved in each lane, which suggests that the method could be used to study also low-abundance protein complexes, e.g. under different physiological conditions.     

Defective dendritic cell migration and activation of adaptive immunity in PI3Kgamma-deficient mice

Gene-targeted mice were used to evaluate the role of the gamma isoform of phosphoinositide 3-kinase (PI3Kgamma) in dendritic cell (DC) migration and induction of specific T-cell-mediated immune responses. DC obtained from PI3Kgamma-/- mice showed a reduced ability to respond to chemokines in vitro and ex vivo and to travel to draining lymph nodes under inflammatory conditions. PI3Kgamma-/- mice had a selective defect in the number of skin Langerhans cells and in lymph node CD8alpha- DC. Furthermore, PI3Kgamma-/- mice showed a defective capacity to mount contact hypersensitivity and delayed-type hypersensitivity reactions. This defect was directly related to the reduced ability of antigen-loaded DC to migrate from the periphery to draining lymph nodes. Thus, PI3Kgamma plays a nonredundant role in DC trafficking and in the activation of specific immunity. Therefore, PI3Kgamma may be considered a new target to control exaggerated immune reactions.     

SPOC: A widely distributed domain associated with cancer,apoptosis and transcription

Background  

The Split ends (Spen) family are large proteins characterised by N-terminal RNA recognition motifs (RRMs) and a conserved SPOC (Spen paralog and ortholog C-terminal) domain. The aim of this study is to characterize the family at the sequence level.     

Identification of the N-glycosylation sites on glutamate carboxypeptidase II necessary for proteolytic activity

Glutamate carboxypeptidase II (GCPII) is a membrane peptidase expressed in the prostate, central and peripheral nervous system, kidney, small intestine, and tumor-associated neovasculature. The GCPII form expressed in the central nervous system, termed NAALADase, is responsible for the cleavage of N-acetyl-L-aspartyl-L-glutamate (NAAG) yielding free glutamate in the synaptic cleft, and is implicated in various pathologic conditions associated with glutamate excitotoxicity. The prostate form of GCPII, termed prostate-specific membrane antigen (PSMA), is up-regulated in cancer and used as an effective prostate cancer marker. Little is known about the structure of this important pharmaceutical target. As a type II membrane protein, GCPII is heavily glycosylated. In this paper we show that N-glycosylation is vital for proper folding and subsequent secretion of human GCPII. Analysis of the predicted N-glycosylation sites also provides evidence that these sites are critical for GCPII carboxypeptidase activity. We confirm that all predicted N-glycosylation sites are occupied by an oligosaccharide moiety and show that glycosylation at sites distant from the putative catalytic domain is critical for the NAAG-hydrolyzing activity of GCPII calling the validity of previously described structural models of GCPII into question.     

Stress kinase phosphorylation is increased in pacing-induced heart failure in rabbits

In hearts with chronic left ventricular (LV) systolic dysfunction secondary to hypertension or myocardial infarction, MAPK phosphorylation and/or activity are increased. Whether other settings of LV dysfunction not associated with ischemia-reperfusion are also characterized by increased MAPK phosphorylation or activity is unknown. After 3 wk of rapid LV pacing (400 beats/min), eight rabbits displayed clinical signs of heart failure (HF), and echocardiography revealed an increase in LV end-diastolic diameter from 15.6 +/- 0.7 (means +/- SE) to 18.8 +/- 0.7 mm and a reduced shortening fraction from 31 +/- 1to10 +/- 2% (both P < 0.05). Morphological alterations in HF included increased numbers of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cardiomyocytes, extent of fibrosis, and cross-sectional cardiomyocyte area. Total p38 MAPK did not differ between failing and normal hearts (n = 8). However, p38 MAPK phosphorylation [164,488 +/- 29,323 vs. 43,565 +/- 14,817 arbitrary units (AU), P < 0.05, densitometry] and the activities of p38 MAPK-alpha and -beta were increased in failing compared with normal hearts (149,441 +/- 38,381 and 170,430 +/- 32,952 vs. 68,815 +/- 28,984 and 81,788 +/- 22,774 AU, respectively, both P < 0.05). In failing compared with normal hearts, total and phosphorylated JNK46 and JNK54 MAPK were increased, whereas total and phosphorylated ERK MAPK remained unchanged. In pacing-induced HF, p38 and JNK MAPK phosphorylation as well as p38 MAPK activity was increased. Further studies will have to define whether or not chronic specific blockade of MAPK activity can interfere with apoptosis/fibrosis and thereby attenuate the progression of HF.     

Cutting edge: scavenging of inflammatory CC chemokines by the promiscuous putatively silent chemokine receptor D6

In an effort to define the actual function of the promiscuous putatively silent chemokine receptor D6, transfectants were generated in different cell types. Engagement of D6 by inflammatory CC chemokines elicited no calcium response nor chemotaxis, but resulted in efficient agonist internalization and degradation. Also in lymphatic endothelium, where this receptor is expressed in vivo, D6 did not elicit cellular responses other than ligand internalization and degradation. In particular, no evidence was obtained for D6-mediated transcytosis of chemokines in the apical-to-basal or basal-to-apical directions. These results indicate that D6 acts as an inflammatory chemokine scavenging nonactivatory decoy receptors and suggest that in lymphatic vessels D6 may function as a gatekeeper for inflammatory CC chemokines, by clearing them and preventing excessive diffusion via afferent lymphatics to lymph nodes.