Synthesis and primary cytotoxicity evaluation of arylmethylenenaphthofuranones derivatives

New series of 2(or 3)-arylmethylenenaphtho[2,1-b]furan-3(or 2)-ones were synthesized, characterized and tested for anticancer properties in vitro. The target compounds were prepared by Knoevenagel coupling between the naphthofuranones 3, 28-30 and formyl derivatives. 2-(4-Oxo-1-benzopyran-3-ylmethylene)naphtho[2,1-b]furan-3-one 36 was the most active compound (IC50 (L1210) = 1.6 microM). These compounds were also evaluated, in an independent manner, as inhibitors of Src protein tyrosine kinase, but only minor activity was observed.     

Proteins associated with the exon junction complex also control the alternative splicing of apoptotic regulators

Several apoptotic regulators, including Bcl-x, are alternatively spliced to produce isoforms with opposite functions. We have used an RNA interference strategy to map the regulatory landscape controlling the expression of the Bcl-x splice variants in human cells. Depleting proteins known as core (Y14 and eIF4A3) or auxiliary (RNPS1, Acinus, and SAP18) components of the exon junction complex (EJC) improved the production of the proapoptotic Bcl-x(S) splice variant. This effect was not seen when we depleted EJC proteins that typically participate in mRNA export (UAP56, Aly/Ref, and TAP) or that associate with the EJC to enforce nonsense-mediated RNA decay (MNL51, Upf1, Upf2, and Upf3b). Core and auxiliary EJC components modulated Bcl-x splicing through different cis-acting elements, further suggesting that this activity is distinct from the established EJC function. In support of a direct role in splicing control, recombinant eIF4A3, Y14, and Magoh proteins associated preferentially with the endogenous Bcl-x pre-mRNA, interacted with a model Bcl-x pre-mRNA in early splicing complexes, and specifically shifted Bcl-x alternative splicing in nuclear extracts. Finally, the depletion of Y14, eIF4A3, RNPS1, SAP18, and Acinus also encouraged the production of other proapoptotic splice variants, suggesting that EJC-associated components are important regulators of apoptosis acting at the alternative splicing level.     

Jelly-falls historic and recent observations: a review to drive future research directions

The biological pump describes the transport of particulate matter from the sea surface to the ocean’s interior including the seabed. The contribution by gelatinous zooplankton bodies as particulate organic matter (POM) vectors (“jelly-falls”) has been neglected owing to technical and spatiotemporal sampling limitations. Here, we assess the existing evidence on jelly-falls from early ocean observations to present times. The seasonality of jelly-falls indicates that they mostly occur after periods of strong upwelling and/or spring blooms in temperate/subpolar zones and during late spring/early summer. A conceptual model helps to define a jelly-fall based on empirical and field observations of biogeochemical and ecological processes. We then compile and discuss existing strategic and observational oceanographic techniques that could be implemented to further jelly-falls research. Seabed video- and photography-based studies deliver the best results, and the correct use of fishing techniques, such as trawling, could provide comprehensive regional datasets. We conclude by considering the possibility of increased gelatinous biomasses in the future ocean induced by upper ocean processes favouring their populations, thus increasing jelly-POM downward transport. We suggest that this could provide a “natural compensation” for predicted losses in pelagic POM with respect to fuelling benthic ecosystems.     

Peripheral regulation by ecdysteroids of olfactory responsiveness in male Egyptian cotton leaf worms, Spodoptera littoralis

Physiological and behavioral plasticity allows animals to adapt to changes in external (environmental) and internal (physiological) factors. In insects, the physiological state modulates adult behavior in response to different odorant stimuli. Hormones have the potential to play a major role in the plasticity of the olfactory responses. To explore if peripheral olfactory processing could be regulated by steroid hormones, we characterized the molecular, electrophysiological, and behavioral response to changes in endogenous hormone levels in adult male Spodoptera littoralis. The expression of the receptor complex (EcR/USP) was localized by in situ hybridization in the olfactory sensilla of antennae. Injections of 20-hydroxyecdysone (20E) induced an ecdysteroid signaling pathway in antennae and increased expression of the nuclear receptors EcR, USP and E75. Diacylglycerol kinase (DGK) and CaM expression were also up-regulated by 20E. Taken together, these molecular, electrophysiological, and behavioral results suggest a hormonal regulation of the peripheral olfactory processing in S. littoralis.     

High identification rates of endogenous neuropeptides from mouse brain

Mass spectrometry-based neuropeptidomics is one of the most powerful approaches for identification of endogenous neuropeptides in the brain. Until now, however, the identification rate of neuropeptides in neuropeptidomics is relatively low and this severely restricts insights into their biological function. In the present study, we developed a high accuracy mass spectrometry-based approach to enhance the identification rates of neuropeptides from brain tissue. Our integrated approach used mixing on column for loading aqueous and organic extracts to reduce the loss of peptides during sample treatment and used charge state-directed tandem mass spectrometry to increase the number of peptides subjected to high mass accuracy fragmentation. This approach allowed 206 peptides on average to be identified from a single mouse brain sample that was prepared using 15 μL of solutions per 1 mg of tissue. In total, we identified more than 500 endogenous peptides from mouse hypothalamus and whole brain samples. Our identification rate is about two to four times higher compared to previously reported studies conducted on mice or other species. The hydrophobic peptides, such as neuropeptide Y and galanin, could be presented and detected with hydrophilic peptides in the same LC-MS run, allowing a high coverage of peptide characterization over an organism. This will advance our understanding of the roles of diverse peptides and their links in the brain functions.     

Extensive characterization of Tupaia belangeri neuropeptidome using an integrated mass spectrometric approach

Neuropeptidomics is used to characterize endogenous peptides in the brain of tree shrews (Tupaia belangeri). Tree shrews are small animals similar to rodents in size but close relatives of primates, and are excellent models for brain research. Currently, tree shrews have no complete proteome information available on which direct database search can be allowed for neuropeptide identification. To increase the capability in the identification of neuropeptides in tree shrews, we developed an integrated mass spectrometry (MS)-based approach that combines methods including data-dependent, directed, and targeted liquid chromatography (LC)-Fourier transform (FT)-tandem MS (MS/MS) analysis, database construction, de novo sequencing, precursor protein search, and homology analysis. Using this integrated approach, we identified 107 endogenous peptides that have sequences identical or similar to those from other mammalian species. High accuracy MS and tandem MS information, with BLAST analysis and chromatographic characteristics were used to confirm the sequences of all the identified peptides. Interestingly, further sequence homology analysis demonstrated that tree shrew peptides have a significantly higher degree of homology to equivalent sequences in humans than those in mice or rats, consistent with the close phylogenetic relationship between tree shrews and primates. Our results provide the first extensive characterization of the peptidome in tree shrews, which now permits characterization of their function in nervous and endocrine system. As the approach developed fully used the conservative properties of neuropeptides in evolution and the advantage of high accuracy MS, it can be portable for identification of neuropeptides in other species for which the fully sequenced genomes or proteomes are not available.     

Immunological tolerance to muscle autoantigens involves peripheral deletion of autoreactive CD8+ T cells

Muscle potentially represents the most abundant source of autoantigens of the body and can be targeted by a variety of severe autoimmune diseases. Yet, the mechanisms of immunological tolerance toward muscle autoantigens remain mostly unknown. We investigated this issue in transgenic SM-Ova mice that express an ovalbumin (Ova) neo-autoantigen specifically in skeletal muscle. We previously reported that antigen specific CD4(+) T cell are immunologically ignorant to endogenous Ova in this model but can be stimulated upon immunization. In contrast, Ova-specific CD8(+) T cells were suspected to be either unresponsive to Ova challenge or functionally defective. We now extend our investigations on the mechanisms governing CD8(+) tolerance in SM-Ova mice. We show herein that Ova-specific CD8(+) T cells are not detected upon challenge with strongly immunogenic Ova vaccines even after depletion of regulatory T cells. Ova-specific CD8(+) T cells from OT-I mice adoptively transferred to SM-Ova mice started to proliferate in vivo, acquired CD69 and PD-1 but subsequently down-regulated Bcl-2 and disappeared from the periphery, suggesting a mechanism of peripheral deletion. Peripheral deletion of endogenous Ova-specific cells was formally demonstrated in chimeric SM-Ova mice engrafted with bone marrow cells containing T cell precursors from OT-I TCR-transgenic mice. Thus, the present findings demonstrate that immunological tolerance to muscle autoantigens involves peripheral deletion of autoreactive CD8(+) T cells.     

The identification of cutin synthase: formation of the plant polyester cutin

A hydrophobic cuticle consisting of waxes and the polyester cutin covers the aerial epidermis of all land plants, providing essential protection from desiccation and other stresses. We have determined the enzymatic basis of cutin polymerization through characterization of a tomato extracellular acyltransferase, CD1, and its substrate, 2-mono(10,16-dihydroxyhexadecanoyl)glycerol. CD1 has in vitro polyester synthesis activity and is required for cutin accumulation in vivo, indicating that it is a cutin synthase.