Different roles for Gi and Go proteins in modulation of adenylyl cyclase type-2 activity

The effect of Gi/o protein-coupled receptors on adenylyl cyclase type 2 (AC2) has been studied in Sf9 insect cells. Stimulation of cells expressing AC2 with the phorbol ester 12-O-tetradecanoyl phorbol-13-acetate (TPA) led to a twofold stimulation of cAMP synthesis that could be blocked with the protein kinase C inhibitor GF109203X. Activation of a coexpressed alpha2A-adrenoceptor or muscarinic M4 receptor inhibited the stimulation by TPA almost completely in a pertussis toxin-sensitive manner. Activation of Gs proteins switched the response of the alpha2A-adrenoceptor to potentiation of prestimulated AC2 activity. The potentiation, but not the inhibition, could be blocked by a Gbetagamma scavenger. A novel methodological approach, whereby signalling through endogenous G proteins was ablated, was used to assess specific G protein species in the signal pathway. Expression of Go proteins (alphao1 + beta1gamma2) restored both the inhibition and the potentiation, whereas expression of Gi proteins (alphai1 + beta1gamma2) resulted in a potentiation of both the TPA- and the Gs-stimulated AC2 activity. The data presented supports the view of AC2 as a molecular switch and implicates this isoform as a target for Go protein-linked signalling.     

Orexin signaling in recombinant neuron-like cells

Recently, we cloned several fluorescent proteins of different colors homologous to Aequorea victoria green fluorescent protein, which have great biotechnological potential as in vivo markers of gene expression. However, later investigations revealed severe drawbacks in the use of novel fluorescent proteins (FPs), in particular, the formation of tetramers (tetramerization) and high molecular weight aggregates (aggregation). In this report, we employ a mutagenic approach to resolve the problem of aggregation. The elimination of basic residues located near the N-termini of FPs results in the generation of non-aggregating versions of several FPs, specifically, drFP583 (DsRed), DsRed-Timer, ds/drFP616, zFP506, zFP538, amFP486, and asFP595.     

Mycorrhizal plants of traditionally managed boreal grasslands in Norway

This paper reports on the mycorrhizal status of 82 plant species growing in traditionally managed grasslands in three different locations in the boreal and boreo-nemoral vegetation zone in the eastern part of Norway. Seventy-four species were found to have arbuscular mycorrhiza (AM). To our knowledge, we report AM for the first time in Achillea ptarmica, Ajuga pyramidalis, Alchemilla glaucescens, Carex brunnescens, Carex pallescens, Crepis praemorsa, Hieracium lactucella, Rumex longifolius, Scorzonera humilis, Trifolium aureum and Trifolium spadiceum. The rare and threatened species Arnica montana, S. humilis, C. praemorsa, Gentianella campestris, Parnassia palustris, T. aureum and T. spadiceum, all confined to grasslands, were found to possess AM fungi.     

Bioengineering of a phytoremediation plant by means of somatic hybridization

Phytoremediation is a technology that exploits a plant's ability to remove contaminants from the environment or render toxic compounds harmless. An efficient metal phytoremediating plant must combine high biomass production and established cultivation methods with high tolerance to a specific contaminant and ability for root uptake, translocation, and compartmentalization of contaminants in the above-ground biomass. Symmetric and asymmetric somatic hybridizations were used to introduce toxic metal-resistant traits from Thlaspi caerulescens into Brassica juncea. B. juncea hypocotyl protoplasts were fused with T. caerulescens mesophyll protoplasts. The hypocotyl protoplasts of B. juncea were stained with CFDA before fusion and thus fluoresced green under UV, whereas the mesophyll protoplasts of T. caerulescens had red autofluorescense. Heteroplasmic fusion products were identified and selected by flow cytometry and cell sorting. All putative hybrids grown in the greenhouse had morphological characteristics of B. juncea. A Thlaspi-specific repetitive sequence was hybridized to total DNA of plants, including the parental species. All plants from both symmetric and asymmetric fusions showed Thlaspi-specific hybridization patterns while B. juncea did not exhibit any hybridization signal. Hybrid plants, produced by asymmetric somatic hybridization between the two species, demonstrated high metal accumulation potential, tolerance to toxic metals, and good biomass production.     

Intravascular plug formation induced by poly-APS is the principal mechanism of the toxin's lethality in rats/rat tissues

Toxic water soluble polymeric 3-alkylpyridinium salts isolated from the sponge Raniera sarai strongly inhibited AChE in vitro. In vivo, experimental animals died due to plugs formed in microcirculation. The mechanism of this plug formation is unknown. In vitro, the toxin did not affect the coagulation rate, but the rate of platelet aggregation was accelerated in a dose-dependent manner. The hemolytic activity of poly-APS was diminished by the addition of serum proteins in a dose-dependent manner. These results support the conclusion that non-specific binding to proteins is the underlying mechanism of the lethality of poly APS.     

Apoptotic pathways: involvement of lysosomal proteases

Apoptosis or programmed cell death is the major mechanism used by multicellular organisms to remove infected, excessive and potentially dangerous cells. Cysteine proteases from the caspase family play a crucial role in the process. However, there is increasing evidence that lysosomal proteases are also involved in apoptosis. In this review various lysosomal proteases and their potential contribution to propagation of apoptosis are discussed.     

Intracerebroventricular interleukin-6 treatment decreases body fat in rats

Recently we found that interleukin-6 (IL-6) knockout mice develop mature-onset obesity and that a single intracerebroventricular (ICV) injection of IL-6 increases energy expenditure. In the present study we investigated if chronic ICV treatment with IL-6 can suppress body fat mass. IL-6 was injected ICV daily for two weeks to rats fed a high-fat diet. IL-6 treatment but not saline treatment decreased body weight by 8.4% and decreased the relative weights of mesenteric and retroperitoneal fat pads. Consistent with this, circulating leptin levels were decreased by 40% after IL-6 treatment but not after saline treatment. Average food intake per day was decreased in the IL-6 treated group compared to the saline treated rats. IL-6 treatment did not change hepatic expression of the acute-phase protein haptoglobin, serum levels of insulin or insulin-like growth factor-I, or the weights of the heart, liver, kidneys, adrenals, and spleen. We conclude that centrally administered IL-6 can decrease body fat in rats without causing acute-phase reaction.     

Pleurotus and Agrocybe hemolysins,new proteins hypothetically involved in fungal fruiting

Novel hemolytic proteins, ostreolysin and aegerolysin, were purified from the fruiting bodies of the edible mushrooms Pleurotus ostreatus and Agrocybe aegerita. Both ostreolysin and aegerolysin have a molecular weight of about 16 kDa, have low isoelectric points of 5.0 and 4.85, are thermolabile, and hemolytic to bovine erythrocytes at nanomolar concentrations. Their activity is impaired by micromolar Hg(2+) but not by membrane lipids and serum low-density lipoproteins (LDL). The sequence of respectively 50 and 10 N-terminal amino acid residues of ostreolysin and aegerolysin has been determined and found to be highly identical with a cDNA-derived amino acid sequence of putative Aa-Pri1 protein from the mushroom A. aegerita, Asp-hemolysin from Aspergillus fumigatus, and two bacterial hemolysin-like proteins expressed during sporulation. We found that ostreolysin is expressed during formation of primordia and fruiting bodies, which is in accord with previous finding that the Aa-Pri1 gene is specifically expressed during fruiting initiation. It is suggestive that the isolated hemolysins play an important role in initial phase of fungal fruiting.