Okadaic acid mimics a nuclear component required for cyclin B-cdc2 kinase microinjection to drive starfish oocytes into M phase

G2-arrested oocytes contain cdc2 kinase as an inactive cyclin B-cdc2 complex. When a small amount of highly purified and active cdc2 kinase, prepared from starfish oocytes at first meiotic metaphase, is microinjected into Xenopus oocytes, it induces activation of the inactive endogenous complex and, as a consequence, drives the recipient oocytes into M phase. In contrast, the microinjected kinase undergoes rapid inactivation in starfish oocytes, which remain arrested at G2. Endogenous cdc2 kinase becomes activated in both nucleated and enucleated starfish oocytes injected with cytoplasm taken from maturing oocytes at the time of nuclear envelope breakdown, but only cytoplasm taken from nucleated oocytes becomes able thereafter to release second recipient oocytes from G2 arrest, and thus contains M phase-promoting factor (MPF) activity. Both nucleated and enucleated starfish oocytes produce MPF activity when type 2A phosphatase is blocked by okadaic acid. If type 2A phosphatase is only partially inhibited, neither nucleated nor enucleated oocytes produce MPF activity, although both do so if purified cdc2 kinase is subsequently injected as a primer to activate the endogenous kinase. The nucleus of starfish oocytes contains an inhibitor of type 2A phosphatase, but neither active nor inactive cdc2 kinase. Microinjection of the content of a nucleus into the cytoplasm of G2-arrested starfish oocytes activates endogenous cdc2 kinase, produces MPF activity, and drives the recipient oocytes into M phase. Together, these results show that the MPF amplification loop is controlled, both positively and negatively, by cdc2 kinase and type 2A phosphatase, respectively. Activation of the MPF amplification loop in starfish requires a nuclear component to inhibit type 2A phosphatase in cytoplasm.     

Differential expression of 240 kDa ConA-binding glycoprotein in vitro and in vivo detected by immunochemical methods

The expression of the 240 ConA-binding glycoprotein (240 kDa), a marker of synaptic junctions isolated from the rat cerebellum, was studied by immunocytochemical techniques in forebrain and cerebellum from rat and chicken, and in chick dorsal root ganglia. Parallel studies were carried out either on tissue sections or in dissociated cell cultures. In all cases non neuronal cells were not immunostained. The tissue sections of cerebellum from rat and chick exhibited 240 kDa glycoprotein immunoreactivity, especially in the molecular layer, while the forebrain sections from rat and chick did not show any significant immunostaining. In contrast, in dissociated forebrain cell cultures, all neuronal cells expressed 240 kDa glycoprotein immunoreactivity, while glial cells remained totally unlabelled. In tissue sections of dorsal root ganglion (DRG), sensory neurons expressed the 240 kDa only after the embryonic day (E 10). A large number of small neurons in the dorsomedial part of DRG were immunostained with 240 kDa glycoprotein antiserum, whereas only a small number of neurons in the ventrolateral part of the ganglia displayed 240 kDa immunoreactivity. In dissociated DRG cells cultures (mixed or neuron-enriched DRG cell cultures) all the neuronal perikarya but not their processes were stained. These studies indicate that 240 kDa glycoprotein expression is completely modified in cultures of neurons of CNS or PNS since the antigen becomes synthetized in high amount by all cells independent of synapse formation. This demonstrates that the expression of 240 kDa is controlled by the cell environment.     

BACE: Therapeutic target and potential biomarker for Alzheimer's disease

β-Site APP-cleaving enzyme (BACE) is a membrane-bound aspartyl protease involved in the production of Alzheimer's disease (AD) Aβ amyloid peptides. This enzyme is ubiquitously expressed, with highest levels in the brain and pancreas. Its cellular trafficking is tightly controlled as it recycles between endosomes and trans-Golgi network. BACE expression increases in response to aging and various stress stimuli. It is elevated in the brain cortex of AD sufferers, and increased levels of BACE in the cerebrospinal fluid of patients with mild cognitive impairment may provide an early biomarker of AD. BACE is considered as a rational drug target for AD therapy, and inhibitors are under development. Anomalies in the behaviour and biochemistry of BACE^?/? mice have pointed to the role this enzyme plays in the processing of neuregulin and of voltage-gated sodium channel β-subunit. A full understanding of BACE biology in health and disease is needed to establish a safe AD therapy based on BACE inhibitors.     

A Qualitative Study of Vaccine Acceptability and Decision Making among Pregnant Women in Morocco during the A (H1N1) pdm09 Pandemic

Vaccination uptake of pregnant women in Morocco during the A (H1N1) pdm09 pandemic was lower than expected. A qualitative study using open-ended questions was developed to explore the main determinants of acceptance and non-acceptance of the monovalent A (H1N1) pdm09 vaccine among pregnant women in Morocco and to identify information sources that influenced their decision-making process. The study sample included 123 vaccinated and unvaccinated pregnant women who were in their second or third trimester between December 2009 and March 2010. They took part in 14 focus group discussions and eight in-depth interviews in the districts of Casablanca and Kenitra. Thematic qualitative analysis identified reasons for vaccine non-acceptance: (1) fear of the monovalent A (H1N1) pdm09 vaccine, (2) belief in an A (H1N1) pdm09 pandemic conspiracy, (3) belief in the inapplicability of the monovalent A (H1N1) pdm09 vaccine to Moroccans, (4) lack of knowledge of the monovalent A (H1N1) pdm09 vaccine, and (5) challenges of vaccination services/logistics. Reasons for vaccine acceptance included: (1) perceived benefits and (2) modeling. Decision-making was strongly influenced by family, community, mass media, religious leaders and health providers suggesting that broad communication efforts should also be used to advocate for vaccination. Meaningful communication for future vaccine campaigns must consider these context-specific findings. As cultural and religious values are shared across many Arab countries, these findings may also provide valuable insights for seasonal influenza vaccine planning in the Middle East and North Africa region at large.     

Carbonic anhydrase activity in primary sensory neurons

Neuronal subpopulations of dorsal root ganglion (DRG) cells in the chicken exhibit carbonic anhydrase (CA) activity. To determine whether CA activity is expressed by DRG cells maintained in in vitro cultures, dissociated DRG cells from 10-day-old chick embryos were cultured on a collagen substrate. The influence exerted by environmental factors on the enzyme expression was tested under various conditions of culture. Neuron-enriched cell cultures and mixed DRG-cell cultures (including numerous non-neuronal cells) were performed either in a defined medium or in a horse serum-supplemented medium. In all the tested conditions, subpopulations of cultured sensory neurons expressed CA activity in their cell bodies, while their neurites were rarely stained; in each case, the percentage of CA-positive neurons declined with the age of the cultures. The number and the persistence of neurons possessing CA activity as well as the intensity of the reaction were enhanced by addition of horse serum. In contrast, the expression of the neuronal CA activity was not affected by the presence of non-neuronal cells or by the rise of CO2 concentration. Thus, the appearance and disappearance of neuronal subpopulations expressing CA activity may be decisively influenced by factors contained in the horse serum. The loss of CA-positive neurons with time could result from a cell selection or from genetic repression. Analysis of the time curves does not support a preferential cell death of CA-positive neurons but suggests that the eventual conversion of CA-positive neurons into CA-negative neurons results from a loss of the enzyme activity. These results indicate that the phenotypic expression of cultured sensory neurons is dependent on defined environmental factors.     

X-changing information on X inactivation

In female somatic cells of mammalian species one X chromosome is inactivated to ensure dosage equality of X-encoded genes between females and males, during development and adulthood. X chromosome inactivation (XCI) involves various epigenetic mechanisms, including RNA mediated gene silencing in cis, DNA methylation, and changes in chromatin modifications and composition. XCI therefore provides an attractive paradigm to study epigenetic gene regulation in a more general context. The XCI process starts with counting of the number of X chromosomes present in a nucleus, and initiation of XCI follows if this number exceeds one per diploid genome. Recently, X-encoded RNF12 has been identified as a dose-dependent activator of XCI. In addition, other factors, including the pluripotency factors OCT4, SOX2 and Nanog, have been implicated to play a role in suppression of initiation of XCI. In this review, we highlight and explain these new and old findings in the context of a stochastic model for X chromosome counting and XCI initiation.     

Electrotonic Coupling between Pyramidal Neurons in the Neocortex

Electrotonic couplings (i.e., electrical synapses or gap junctions) are fundamental to neuronal synchronization, and thus essential for many physiological functions and pathological disorders. Interneuron electrical synapses have been studied intensively. Although studies on electrotonic couplings between pyramidal cells (PCs) are emerging, particularly in the hippocampus, evidence is still rare in the neocortex. The electrotonic coupling of PCs in the neocortex is therefore largely unknown in terms of electrophysiological, anatomical and synaptological properties. Using multiple patch-clamp recording with differential interference contrast infrared videomicroscopy (IR-DIC) visualization, histochemical staining, and 3D-computer reconstruction, electrotonic coupling was recorded between close PCs, mainly in the medial prefrontal cortex as well as in the visual cortical regions of ferrets and rats. Compared with interneuron gap junctions, these electrotonic couplings were characterized by several special features. The recording probability of an electrotonic coupling between PCs is extremely low; but the junctional conductance is notably high, permitting the direct transmission of action potentials (APs) and even tonic firing between coupled neurons. AP firing is therefore perfectly synchronized between coupled PCs; Postjunctional APs and spikelets alternate following slight changes of membrane potentials; Postjunctional spikelets, especially at high frequencies, are summated and ultimately reach AP-threshold to fire. These properties of pyramidal electrotonic couplings largely fill the needs, as predicted by simulation studies, for the synchronization of a neuronal assembly. It is therefore suggested that the electrotonic coupling of PCs plays a unique role in the generation of neuronal synchronization in the neocortex.     

The desert of Tataouine: an extreme environment that hosts a wide diversity of microorganisms and radiotolerant bacteria

The phylogenetic diversity of prokaryotic communities exposed to arid conditions in the hot desert of Tataouine (south Tunisia) was estimated with a combination of a culture and - molecular-based analysis. Thirty-one isolates, representative of each dominant morphotypes, were affiliated to Actinobacteria, Firmicutes, Proteobacteria and the CFB group while none related to Archaea. Analysis of 16S rRNA gene libraries revealed the presence of species related to Bacteria and Archaea. Sequences related to Archaea were all affiliated to the non-thermophilic Crenarchaeota subgroup. Bacterial sequences were dominated by Proteobacteria, Actinobacteria and Acidobacteria; a few sequences were distributed among eight others phyla, including Thermus/Deinococcus relatives. A correlation between tolerance to desiccation and to radiation has been demonstrated for the radiotolerant bacteria Deinococcus radiodurans. Because bacteria living in the hot desert of Tataouine are one way or another tolerant to desiccation, we investigate whether they could also be tolerant to radiation. Exposition of soil samples to intense gamma radiation yields Bacillus, Thermus/Deinococcus and alpha-Proteobacteria relatives. Four of these strains correspond to radiotolerant species as revealed by evaluation of the resistance levels of the individual cultures. A detailed analysis of the resistance levels for two Thermus/Deinococcus and two alpha-Proteobacteria relatives revealed that they correspond to new radiotolerant species.     

Flow-activated ion channels in vascular endothelium

The ability of vascular endothelial, cells (ECs) to respond to fluid mechanical forces associated with blood flow is essential for flow-mediated vasoregulation and arterial wall remodeling. Abnormalities in endothelial responses to flow also play a role in the development of atherosclerosis. Although our understanding of the endothelial signaling pathways stimulated by flow has greatly increased over the past two decades, the mechanisms by which ECs sense flow remain largely unknown. Activation of flow-sensitive ion channels is among the fastest known endothelial responses to flow; therefore, these ion channels have been proposed as candidate flow sensors. This review focuses on: 1) describing the various types of flow-sensitive ion channels that have been reported in ECs, 2) discussing the implications of activation of these ion channels for endothelial function, and 3) proposing candidate mechanisms for activation of flow-sensitive ion channels.