Structure of the RZZ complex and molecular basis of Spindly‐driven corona assembly at human kinetochores

In metazoans, a ≈1 megadalton (MDa) multiprotein complex comprising the dynein–dynactin adaptor Spindly and the ROD–Zwilch–ZW10 (RZZ) complex is the building block of a fibrous biopolymer, the kinetochore fibrous corona. The corona assembles on mitotic kinetochores to promote microtubule capture and spindle assembly checkpoint (SAC) signaling. We report here a high‐resolution cryo‐EM structure that captures the essential features of the RZZ complex, including a farnesyl‐binding site required for Spindly binding. Using a highly predictive in vitro assay, we demonstrate that the SAC kinase MPS1 is necessary and sufficient for corona assembly at supercritical concentrations of the RZZ–Spindly (RZZS) complex, and describe the molecular mechanism of phosphorylation‐dependent filament nucleation. We identify several structural requirements for RZZS polymerization in rings and sheets. Finally, we identify determinants of kinetochore localization and corona assembly of Spindly. Our results describe a framework for the long‐sought‐for molecular basis of corona assembly on metazoan kinetochores.     

Organisation of the Hb 1 genes of the Antarctic skate Bathyraja eatonii: new insights into the evolution of globin genes

An extensive investigation of the organisation of globin genes has greatly contributed to the understanding of universal mechanisms of gene evolution and expression. Cartilaginous fish are the first organisms that have evolved the tetrameric form of hemoglobin (Hb). So far, there has been absolute lack of data about globin genes in chondrichthyans. Bathyraja is the dominant rajid south of 60 degrees S. In the framework of the investigations on globin genes of Antarctic red-blooded and Hb-less fish we obtained the cloning of the alpha- and beta-globin cDNAs of the main Hb (Hb 1) of the skate Bathyraja eatonii. Then, a genomic fragment of 6.2 kb was isolated where the Hb 1 alpha and beta genes are linked in a tail-to-head (3' to 5') orientation. The beta-globin gene promoter region and the chromosomal organisation of the Hb 1 genes of B. eatonii have been compared to their homologues in other vertebrates. The finding of a tail-to-head linkage of the Hb 1 alpha- and beta-globin genes in B. eatonii is the first characterisation of the organisation of globin genes in chondrichthyes; such finding offers a novel contribution to the understanding of the evolution of this class of genes. Moreover, the characterisation of chondrichthyan genes is very important for gaining insight into the ancestral state of vertebrate genomes.     

Systematic Review of the Literature and Evidence-Based Recommendations for Antibiotic Prophylaxis in Trauma: Results from an Italian Consensus of Experts

Background

Antibiotic prophylaxis is frequently administered in severe trauma. However, the risk of selecting resistant bacteria, a major issue especially in critical care environments, has not been sufficiently investigated. The aim of the present study was to provide guidelines for antibiotic prophylaxis for four different trauma-related clinical conditions, taking into account the risks of antibiotic-resistant bacteria selection, thus innovating previous guidelines in the field.

Methods

The MEDLINE database was searched for studies comparing antibiotic prophylaxis to controls (placebo or no antibiotic administration) in four clinical traumatic conditions that were selected on the basis of the traumatic event frequency and/or infection severity. The selected studies focused on the prevention of early ventilator associated pneumonia (VAP) in comatose patients with traumatic brain injury, of meningitis in severe basilar skull fractures, of wound infections in long-bone open fractures. Since no placebo-controlled study was available for deep surgical site-infections prevention in abdominal trauma with enteric contamination, we compared 24-hour and 5-day antibiotic prophylaxis policies. A separate specific research focused on the question of antibiotic-resistant bacteria selection caused by antibiotic prophylaxis, an issue not adequately investigated by the selected studies. Randomised trials, reviews, meta-analyses, observational studies were included. Data extraction was carried out by one author according to a predefined protocol, using an electronic form. The strength of evidence was stratified and recommendations were given according to the Grading of Recommendations Assessment, Development and Evaluation (GRADE) criteria.

Results

Uncertain evidence deserving further studies was found for two-dose antibiotic prophylaxis for early VAP prevention in comatose patients. In the other cases the risk of resistant-bacteria selection caused by antibiotic administration for 48 hours or more, outweighed potential benefits.

Conclusions

When accounting for antibiotic-resistant bacteria selection we found no evidence in favour of antibiotic prophylaxis lasting two or more days in the studied clinical conditions.     

The animat: new frontiers in whole brain modeling

The researchers at Boston University (BU)'s Neuromorphics Laboratory, part of the National Science Foundation (NSF)-sponsored Center of Excellence for Learning in Education, Science, and Technology (CELEST), are working in collaboration with the engineers and scientists at Hewlett-Packard (HP) to implement neural models of intelligent processes for the next generation of dense, low-power, computer hardware that will use memristive technology to bring data closer to the processor where computation occurs. The HP and BU teams are jointly designing an optimal infrastructure, simulation, and software platform to build an artificial brain. The resulting Cog Ex Machina (Cog) software platform has been successfully used to implement a large-scale, multicomponent brain system that is able to simulate some key rat behavioral results in a virtual environment and has been applied to control robotic platforms as they learn to interact with their environment.     

Conformational transitions of the Spindly adaptor underlie its interaction with Dynein and Dynactin

Cytoplasmic Dynein 1, or Dynein, is a microtubule minus end–directed motor. Dynein motility requires Dynactin and a family of activating adaptors that stabilize the Dynein–Dynactin complex and promote regulated interactions with cargo in space and time. How activating adaptors limit Dynein activation to specialized subcellular locales is unclear. Here, we reveal that Spindly, a mitotic Dynein adaptor at the kinetochore corona, exists natively in a closed conformation that occludes binding of Dynein–Dynactin to its CC1 box and Spindly motif. A structure-based analysis identified various mutations promoting an open conformation of Spindly that binds Dynein–Dynactin. A region of Spindly downstream from the Spindly motif and not required for cargo binding faces the CC1 box and stabilizes the intramolecular closed conformation. This region is also required for robust kinetochore localization of Spindly, suggesting that kinetochores promote Spindly activation to recruit Dynein. Thus, our work illustrates how specific Dynein activation at a defined cellular locale may require multiple factors.     

Functional distinction between serotonin uptake and serotonin-induced shape change receptors in rat platelets

Tyramine and dopamine are taken up by rat platelets through the serotonin uptake mechanism while phenethylamine is not taken up. This indicates that an aromatic hydroxyl group is a structural requirement for the uptake of phenethylamine derivatives by rat platelets. Although none of these phenethylamine derivatives induce platelet shape change, they inhibit serotonin-induced shape change and serotonin uptake with the same relative potency ($\text{tyramine}\text{>}\text{phenethylamine}\text{?}\text{dopamine}$). This suggests that the receptors controlling serotonin uptake and serotonin-induced shape change have a common structural component that binds phenethylamine derivatives. However, the fact that phenethylamine derivatives activate the serotonin uptake mechanism but do not induce platelet shape change suggests that serotonin uptake and serotonin-induced shape change are mediated by two distinct activation sites of serotonin receptors.     

The localization of acetylcholinesterase activity in the spinal ganglia of the adult fowl studied by electron microscope histochemistry

Summary AChE activity was localized in spinal ganglia of adult fowls at the electron microscope level using Karnovsky’s method. Controls with BW 284 C 51 were carried out. In the neuronal bodies, AChE activity was evident within the rough-surfaced cisternae of the endoplasmic reticulum, including the perinuclear cisterna and the subsurface cisternae, and sometimes in the innermost cisternae of the Golgi complex. AChE activity was also demonstrated along the axolemma and associated with smooth-surfaced vesicles and tubules in the initial segment of the axon, in all the ganglionic myelinated fibers examined by serial section analysis and in more than half of the ganglionic unmyelinated fibers examined by this method. In the myelinated fibers the reaction product appeared more abundant at the level of the nodes of Ranvier than in the internodal segments. Both in the myelinated and unmyelinated fibers a considerable quantitative variability of reaction product was observed among the various sections of the same fiber. These results were compared with those previously obtained in the spinal ganglia of the chick embryo using the same histochemical method. This research was supported by a grant of the National Research Council (C.N.R.), Italy, and of the Deutsche Forschungsgemeinschaft (DFG), Federal Republic of Germany. 1 AChE=acetylcholinesterase (=true or specific cholinesterase); ACh=acetylcholine; ChAc=choline acetylase (=choline acetyltransferase).     

Selective stimulation of serotonin2c receptors blocks the enhancement of striatal and accumbal dopamine release induced by nicotine administration

The effects of acute and repeated nicotine administration on the extracellular levels of dopamine (DA) in the corpus striatum and the nucleus accumbens were studied in conscious, freely moving rats by in vivo microdialysis. Acute intraperitoneal (i.p.) injection of nicotine (1 mg/kg) increased DA outflow both in the corpus striatum and the nucleus accumbens. Repeated daily injection of nicotine (1 mg/kg, i.p.) for 10 consecutive days caused a significant increase in basal DA outflow both in the corpus striatum and the nucleus accumbens. Acute challenge with nicotine (1 mg/kg, i.p.) in animals treated repeatedly with this drug enhanced DA extracellular levels in both brain areas. However, the effect of nicotine was potentiated in the nucleus accumbens, but not in the corpus striatum. To test the hypothesis that stimulation of 5-HT (5-hydroxytryptamine, serotonin)(2C) receptors could affect nicotine-induced DA release, the selective 5-HT(2C) receptor agonist RO 60-0175 was used. Pretreatment with RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently prevented the enhancement in DA release elicited by acute nicotine in the corpus striatum, but was devoid of any significant effect in the nucleus accumbens. RO 60-0175 (1 and 3 mg/kg, i.p.) dose-dependently reduced the stimulatory effect on striatal and accumbal DA release induced by an acute challenge with nicotine (1 mg/kg, i.p.) in rats treated repeatedly with this alkaloid. However, only the effect of 3 mg/kg RO 60-0175 reached statistical significance. The inhibitory effect of RO 60-0175 on DA release induced by nicotine in the corpus striatum and the nucleus accumbens was completely prevented by SB 242084 (0.5 mg/kg, i.p.) and SB 243213 (0.5 mg/kg, i.p.), two selective antagonists of 5-HT(2C) receptors. It is concluded that selective activation of 5-HT(2C) receptors can block the stimulatory action of nicotine on central DA function, an effect that might be relevant for the reported antiaddictive properties of RO 60-0175.     

Non-steroidal anti-inflammatory drugs (NSAIDs) in Alzheimer's disease: old and new mechanisms of action

Alzheimer's disease (AD) is characterized by cerebral deposits of beta-amyloid (A beta) peptides and neurofibrillary tangles (NFT) which are surrounded by inflammatory cells. Epidemiological studies have shown that prolonged use of non-steroidal anti-inflammatory drugs (NSAIDs) reduces the risk of developing AD and delays the onset of the disease. It has been postulated that some NSAIDs target pathological hallmarks of AD by interacting with several pathways, including the inhibition of cyclooxygenases (COX) and activation of the peroxisome proliferator-activated receptor gamma. A variety of experimental studies indicate that a subset of NSAIDs such as ibuprofen, flurbiprofen, indomethacin and sulindac also possess A beta-lowering properties in both AD transgenic mice and cell cultures of peripheral, glial and neuronal origin. While COX inhibition occurs at low concentrations in vitro (nM-low microm range), the A beta-lowering activity is observed at high concentrations (< or = 50 microm). Nonetheless, studies with flurbiprofen or ibuprofen in AD transgenic mice show that the effects on A beta levels or deposition are attained at plasma levels similar to those achieved in humans at therapeutic dosage. Still, it remains to be assessed whether adequate concentrations are reached in the brain. This is a crucial aspect that will allow defining the dose-window and the length of treatment in future clinical trials. Here, we will discuss how the combination of anti-amyloidogenic and anti-inflammatory activities of certain NSAIDs may produce a profile potentially relevant to their clinical use as disease-modifying agents for the treatment of AD.