The G-protein-coupled receptor phosphatase: a protein phosphatase type 2A with a distinct subcellular distribution and substrate specificity.

Phosphorylation of G-protein-coupled receptors plays an important role in regulating their function. In this study the G-protein-coupled receptor phosphatase (GRP) capable of dephosphorylating G-protein-coupled receptor kinase-phosphorylated receptors is described. The GRP activity of bovine brain is a latent oligomeric form of protein phosphatase type 2A (PP-2A) exclusively associated with the particulate fraction. GRP activity is observed only when assayed in the presence of protamine or when phosphatase-containing fractions are subjected to freeze/thaw treatment under reducing conditions. Consistent with its identification as a member of the PP-2A family, the GRP is potently inhibited by okadaic acid but not by I-2, the specific inhibitor of protein phosphatase type 1. Solubilization of the membrane-associated GRP followed by gel filtration in the absence of detergent yields a 150-kDa peak of latent receptor phosphatase activity. Western blot analysis of this phosphatase reveals a likely subunit composition of AB alpha C. PP-2A of this subunit composition has previously been characterized as a soluble enzyme, yet negligible soluble GRP activity was observed. The subcellular distribution and substrate specificity of the GRP suggests significant differences between it and previously characterized forms of PP-2A.

Differential subcellular mRNA targeting: deletion of a single nucleotide prevents the transport to axons but not to dendrites of rat hypothalamic magnocellular neurons.

It has previously been shown that mRNA encoding the arginine vasopressin (AVP) precursor is targeted to axons of rat magnocellular neurons of the hypothalamo-neurohypophyseal tract. In the homozygous Brattle-boro rat, which has a G nucleotide deletion in the coding region of the AVP gene, no such targeting is observed although the gene is transcribed. RNase protection and heteroduplex analyses demonstrate that, in heterozygous animals, which express both alleles of the AVP gene, the wild-type but not the mutant transcript is subject to axonal compartmentation. In contrast, wild-type and mutant AVP mRNAs are present in dendrites. These data suggest the existence of different mechanisms for mRNA targeting to the two subcellular compartments. Axonal mRNA localization appears to take place after protein synthesis; the mutant transcript is not available for axonal targeting because it lacks a stop codon preventing its release from ribosomes. Dendritic compartmentation, on the other hand, is likely to precede translation and, thus, would be unable to discriminate between the two mRNAs.

Cell cycle-related shifts in subcellular localization of BCR: association with mitotic chromosomes and with heterochromatin.

The disruption of the BCR gene and its juxtaposition to and consequent activation of the ABL gene has been implicated as the critical molecular defect in Philadelphia chromosome-positive leukemias. The normal BCR protein is a multifunctional molecule with domains that suggest its participation in phosphokinase and GTP-binding pathways. Taken together with its localization to the cytoplasm of uncycled cells, it is therefore presumed to be involved in cytoplasmic signaling. By performing a double aphidicolin block for cell cycle synchronization, we currently demonstrate that the subcellular localization of BCR shifts from being largely cytoplasmic in interphase cells to being predominantly perichromosomal in mitosis. Furthermore, with the use of immunogold labeling and electron microscopy, association of BCR with DNA, in particular heterochromatin, can be demonstrated even in quiescent cells. Results were similar in cell lines of lymphoid or myeloid origin. These observations suggest a role for BCR in the phosphokinase interactions linked to condensed chromatin, a network previously implicated in cell cycle regulation.

Studying Coxiella burnetii Type IV Substrates in the Yeast Saccharomyces cerevisiae: Focus on Subcellular Localization and Protein Aggregation.

Coxiella burnetii is a Gram-negative obligate parasitic bacterium that causes the disease Q-fever in humans. To establish its intracellular niche, it utilizes the Icm/Dot type IVB secretion system (T4BSS) to inject protein effectors into the host cell cytoplasm. The host targets of most cognate and candidate T4BSS-translocated effectors remain obscure. We used the yeast Saccharomyces cerevisiae as a model to express and study six C. burnetii effectors, namely AnkA, AnkB, AnkF, CBU0077, CaeA and CaeB, in search for clues about their role in C. burnetii virulence. When ectopically expressed in HeLa cells, these effectors displayed distinct subcellular localizations. Accordingly, GFP fusions of these proteins produced in yeast also decorated distinct compartments, and most of them altered cell growth. CaeA was ubiquitinated both in yeast and mammalian cells and, in S. cerevisiae, accumulated at juxtanuclear quality-control compartments (JUNQs) and insoluble protein deposits (IPODs), characteristic of aggregative or misfolded proteins. AnkA, which was not ubiquitinated, accumulated exclusively at the IPOD. CaeA, but not AnkA or the other effectors, caused oxidative damage in yeast. We discuss that CaeA and AnkA behavior in yeast may rather reflect misfolding than recognition of conserved targets in the heterologous system. In contrast, CBU0077 accumulated at vacuolar membranes and abnormal ER extensions, suggesting that it interferes with vesicular traffic, whereas AnkB associated with the yeast nucleolus. Both effectors shared common localization features in HeLa and yeast cells. Our results support the idea that C. burnetii T4BSS effectors manipulate multiple host cell targets, which can be conserved in higher and lower eukaryotic cells. However, the behavior of CaeA and AnkA prompt us to conclude that heterologous protein aggregation and proteostatic stress can be a limitation to be considered when using the yeast model to assess the function of bacterial effectors.

Sediment and organic matter properties, and molecular formula from sampling locations in the Northern Black Sea

Remineralization of organic matter in reactive marine sediments releases nutrients and dissolved organic matter (DOM) into the ocean. Here we focused on the molecular-level characterization of DOM by high-resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) in sediment pore waters and bottom waters from contrasting redox regimes in the northern Black Sea with particular emphasis on nitrogen-bearing compounds to derive an improved understanding of the molecular transformations involved in nitrogen release. The number of nitrogen-bearing molecules is generally higher in pore waters than in bottom waters. This suggests intensified degradation of nitrogen-bearing precursor molecules such as proteins in anoxic sediments: No significant difference was observed between sediments deposited under oxic vs anoxic conditions (average O/C ratios of 0.55) suggesting that the different organic matter quality induced by contrasting redox conditions does not impact protein diagenesis in the subseafloor. Compounds in the pore waters were on average larger, less oxygenated, and had a higher number of unsaturations. Applying a mathematical model, we could show that the assemblages of nitrogen-bearing molecular formulas are potential products of proteinaceous material that was transformed by the following reactions: (a) hydrolysis and deamination, both reducing the molecular size and nitrogen content of the products and intermediates; (b) oxidation and hydration of the intermediates; and (c) methylation and dehydration.

(Tables 1,2) Viability of Poa trivialis seeds transported to Antarctica from different locations

Antarctic ecosystems are at risk from the introduction of invasive species. The first step in the process of invasion is the transportation of alien species to Antarctic in a viable state. However, the effect of long-distance human-mediated dispersal, over different time-scales, on propagule viability is not well known. We assessed the viability of Poa trivialis seeds transported to Antarctica from the UK, South Africa and Australia by ship or by ship and aircraft. Following transportation to the Antarctic Treaty area, no reduction in seed viability was found, despite journey times lasting up to 284 days and seeds experiencing temperatures as low as -1.5°C. This work confirms that human-mediated transport may overcome the dispersal barrier for some propagules, and highlights the need for effective pre-departure biosecurity measures.

Raised pavement markers at hazardous locations. Final report.

Mode of access: Internet.

Airline airports : locations designated for airports for air carrier scheduled operations and related information.

"April 1958."

Market locations for recovered materials /

Mode of access: Internet.

Optimal detector locations for HOV lane operations.

Texas Department of Transportation, Austin