Fingerprinting the junctions of RNA structure by an open-paddlewheel diruthenium compound

RNA function is determined by its structural organization. The RNA structure consists of the combination of distinct secondary structure motifs connected by junctions that play an essential role in RNA folding. Selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE) probing is an established methodology to analyze the secondary structure of long RNA molecules in solution, which provides accurate data about unpaired nucleotides. However, the residues located at the junctions of RNA structures usually remain undetected. Here we report an RNA probing method based on the use of a novel open-paddlewheel diruthenium (OPW-Ru) compound [Ru₂Cl₂(µ-DPhF)₃(DMSO)] (DPhF = N,N′-diphenylformamidinate). This compound has four potential coordination sites in a singular disposition to establish covalent bonds with substrates. As a proof of concept, we have analyzed the reactivity of OPW-Ru toward RNA using two viral internal ribosome entry site (IRES) elements whose function depends on the structural organization of the molecule. Our study suggests that the compound OPW-Ru preferentially attacks at positions located one or two nucleotides away from junctions or bulges of the RNA structure. The OPW-Ru fingerprinting data differ from that obtained by other chemical reagents and provides new information about RNA structure features.     

Serotonin Depletion Does not Modify the Short-Term Brain Hypometabolism and Hippocampal Neurodegeneration Induced by the Lithium–Pilocarpine Model of Status Epilepticus in Rats

It has been reported that fluoxetine, a selective serotonin (5-hydroxytryptamine; 5-HT) reuptake inhibitor, has neuroprotective properties in the lithium–pilocarpine model of status epilepticus (SE) in rats. The aim of the present study was to investigate the effect of 5-HT depletion by short-term administration of p-chlorophenylalanine (PCPA), a specific tryptophan hydroxylase inhibitor, on the brain hypometabolism and neurodegeneration induced in the acute phase of this SE model. Our results show that 5-HT depletion did modify neither the brain basal metabolic activity nor the lithium–pilocarpine-induced hypometabolism when evaluated 3 days after the insult. In addition, hippocampal neurodegeneration and astrogliosis triggered by lithium–pilocarpine were not exacerbated by PCPA treatment. These findings point out that in the early latent phase of epileptogenesis, non-5-HT-mediated actions may contribute, at least in some extent, to the neuroprotective effects of fluoxetine in this model of SE.     

Invasion of amphisteginid foraminifera in the Adriatic Sea

Amphisteginid foraminifera are larger symbiont-bearing foraminifera that are abundant in tropical and subtropical reef and shelf regions of the world’s oceans. There is now unequivocal evidence that climate change has led to an expansion of tropical belts. As temperatures rise, surface isotherms are shifting poleward and habitat ranges are moving towards higher latitudes. Fueled by substantial warming, the Mediterranean has been identified as one of the most severely affected areas. This study documents for the first time that amphisteginid foraminifera have now crossed the Strait of Otranto and invaded the eastern Adriatic coast along southern Albania. Sampling on the opposite side along the southern part of the Italian boot shows, however, that amphisteginid foraminifera have not yet colonized the western coast of the Adriatic. The asymmetric invasion of the Adriatic displays spatial heterogeneity, where the progressive colonization follows the major surface currents with a northerly flow along the eastern coast and a southerly return along the western coastline. Previous sampling along Ionian Sea sample sites provides a baseline chronology allowing computations of range expansion rates. The observed recent rate of range expansion in Amphistegina is computed between 4.0 and 10 km/year^?1 and provides strong support for current species model projections. Based on the new data, the range boundary shift is projected to lead to a total northward range expansion of 5.2° latitude in the year 2100. The latitudinal range extension computed for the Mediterranean is thus almost twice as large as those computed for the southern hemisphere. The ongoing range extension into new areas of the Mediterranean Sea is shown to trigger changes in community structures with potential consequences for ecosystem functioning. Numerical abundances and impacts of invasive amphisteginids are greatest at shallow depths <20 m. Evidence is now accumulating that the invasion of amphisteginid foraminifera results in (1) a loss of benthic foraminiferal biodiversity, (2) alterations of foraminiferal community structures and (3) shifts in abundances of functional groups of foraminifera.     

Differential Targeting of SLC30A10/ZnT10 Heterodimers to Endolysosomal Compartments Modulates EGF‐Induced MEK/ERK1/2 Activity

The solute carrier 30A (SLC30A) family of zinc exporters transports zinc into the lumen of intracellular organelles in order to prevent zinc toxicity. We reported that formation of tyrosine dimers is required for ZnT3 (zinc transporter 3) zinc transport activity and targeting to synaptic‐like microvesicles (SLMVs) in PC12 cells and the formation of ZnT3/ZnT10 heterodimers. Here, we focused on ZnT10 to determine the role of heterodimerization in the sorting of ZnTs in the endolysosomal pathway. Using cell fractionation, immunoprecipitation and immunofluorescence approaches, we found that ZnT10 resides in transferrin receptor and Rab5‐positive endosomes and forms covalent heterodimers and oligomers with ZnT2, ZnT3 and ZnT4. The interaction of ZnT10 with ZnT3, mediated by dityrosine bonds, was unable to target ZnT10 into SLMVs in vitro or into synaptic vesicles isolated from mouse brain in vivo. However, ZnT3/ZnT10 heterodimers regulate epidermal growth factor receptor (EGF‐R) signaling by increasing the phosphorylation of mitogen‐activated protein kinase kinase (MEK) and extracellular signal‐regulated kinase (ERK1/2), but not EGF‐R, C‐Raf or Akt phosphorylation in response to EGF. Further, mutation of tyrosine 4 in ZnT10 reduced ZnT3/ZnT10 dityrosine‐mediated heterodimerization and zinc transport, as well as MEK and ERK1/2 phosphorylation, which were also reduced by the zinc chelator TPEN. Phosphorylation of these kinases is likely to occur in the cytosol as no differences in phosphorylation were observed in membrane fractions of control and ZnT3/ZnT10‐expressing cells. We propose that ZnT10 plays a role in signal transduction, which is mediated by homo and heterodimerization with other ZnTs.      

A role for galectin-3 in CD13-mediated homotypic aggregation of monocytes

We recently reported that anti-CD13 mAbs induce homotypic aggregation of monocytic cells. This phenomenon is signal transduction dependent and does not require CD13 aminopeptidase activity. Since CD13 is heavily glycosylated and a member of the galectin family (galectin-4) has been shown to associate with CD13 in the intestinal epithelium, we hypothesized that CD13-mediated aggregation might proceed through a carbohydrate-dependent mechanism involving galectin-3, the most highly expressed galectin on monocytes. We report here that lactose and anti-galectin-3 antibodies completely abrogate homotypic aggregation induced by anti-CD13 antibodies. Furthermore, galectin-3 co-immunoprecipitates with CD13 from resting U-937 cells and this association decreases during the aggregation process, a phenomenon that may have functional implications. Together, the results presented here point to a key role for galectin-3 in CD13-mediated homotypic aggregation of monocytic cells.