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The cuticle constitutes the outermost defensive barrier of most land plants. It comprises a polymeric matrix—cutin, surrounded by soluble waxes. Moreover, the cuticle constitutes the first line of defense against pathogen invasion, while also protecting the plant from many abiotic stresses. Aliphatic monomers in cutin have been suggested to act as immune elicitors in plants. This study analyses the potential of cutin oligomers to activate rapid signaling outputs reminiscent of pattern-triggered immunity in the model plant Arabidopsis. Cutin oligomeric mixtures led to Ca2+ influx and mitogen-activated protein kinase activation. Comparable responses were measured for cutin, which was also able to induce a reactive oxygen species burst. Furthermore, cutin oligomer treatment resulted in a unique transcriptional reprogramming profile, having many archetypal features of pattern-triggered immunity. Targeted spectroscopic and spectrometric analyses of the cutin oligomers suggest that the elicitor compounds consist mostly of two up to three 10,16-dihydroxyhexadecanoic acid monomers linked together through ester bonds. This study demonstrates that cutin breakdown products can act as inducers of early plant immune responses. Further investigation is needed to understand how cutin breakdowns are perceived and to explore their potential use in agriculture. [ABSTRACT FROM AUTHOR]

Fresh horticultural products are economically significant foods that are highly demanded by consumers worldwide; however, they are highly perishable and susceptible to deterioration by fungi, which contribute to their short shelf-life and cause significant post-harvest losses. Among the alternatives suggested for fungal control in plants is the elicitation of the innate plant defense mechanism, which may be activated when specific molecules of the phytopathogen, such as chitin, are recognized. Chitin is a long-chain polymer of N-acetyl-α-D-glucosamine of the fungal cell wall; it possesses biological activity by eliciting the plant immune response. This molecule and its oligosaccharides are recognized through transmembrane receptors known as receptor-like kinases (RLKs) and receptor-like proteins (RLPs). Mediated by receptor-like cytoplasmic kinases (RLCKs), which bind to the intracellular domain of these receptors, they initiate intracellular signal transduction via MAP kinases, triggering the plant defense response. In model plants, such as Oryza sativa (rice) and Arabidopsis thaliana, the set of RLK/RLP-RLCK-MAP kinases is involved in plant immunity triggered by chitin. Furthermore, in horticultural products, research into the molecular events between these three elements has suggested that similar processes occur. However, little is known about these molecular events in fruits. Against this background, the present review provides the most recent and relevant findings on the molecular associations of these three elements in the response to fungal chitin in plants and outlines which elements could participate in this signaling process in horticultural crops. [ABSTRACT FROM AUTHOR]

Background: Oocytes of large animal species isolated from small ovarian follicles (< 2 mm) are less competent to support early embryonic development after in vitro maturation and fertilization than their counterparts isolated from medium-sized and preovulatory follicles. This study aimed to assess the effect of a new maturation medium containing FGF2, LIF, and IGF1 (FLI medium) on the meiotic and developmental competence of pig cumulus-oocytes complexes (COCs) derived from the small and medium-sized follicles. Methods: The growing oocytes were isolated from 1 to 2 (small follicle; SF) and the fully-grown ones from 3 to 6 (large follicle; LF) mm follicles and matured in a control M199 medium with gonadotropins and EGF and the FLI medium enriched by the triplet of growth factors. The matured oocytes were parthenogenetically activated and cultured to the blastocyst stage. Chromatin configuration before and during the culture and MAP kinase activity were assessed in the oocytes. Finally, the expression of cumulus cell genes previously identified as markers of oocyte quality was assessed. Results: The maturation and blastocyst rates of oocytes gained from LF were significantly higher than that from SF in the control medium. In contrast, similar proportions of oocytes from LF and SF completed meiosis and developed to blastocysts when cultured in FLI. Most of the oocytes freshly isolated from SF possessed germinal vesicles with fine filaments of chromatin (GV0) or chromatin surrounding the nucleolus (GVI; 30%); the oocytes from LF were mainly in GVI (or GVII) exhibiting a few small lumps of chromatin beneath the nuclear membrane. When cultured in the FLI medium for 16 h, an acceleration of the course of maturation in oocytes both from SF and LF compared to the control medium was observed and a remarkable synchrony in the course of chromatin remodeling was noticed in oocytes from SF and LF. Conclusions: This work demonstrates that the enrichment of culture medium by FGF2, LIF, and IGF1 can enhance the meiotic and developmental competence of not only fully-grown, but also growing pig oocytes and significantly thus expanding the number of oocytes available for various assisted reproductive technology applications. [ABSTRACT FROM AUTHOR]

We recently reported an RAF rearrangement without NRAS or BRAF mutations in lesions from Giant Congenital Melanocytic Nevi (CMN). The new gene fusion involves the 5′‐end of the promoter‐containing N terminus of the SOX5 gene fused to exons 7–16 of the 3′‐end of RAF1 gene leading to a SOX5‐RAF1 fusion transcript which loses the auto‐inhibitory CR1 domain but retains the complete in‐frame coding sequence for the C‐Terminal kinase domain of the RAF1. Stable expression of SOX5‐RAF1 fusion induced growth factor‐independent cell growth in murine hematopoietic Ba/F3 cells and melan‐a immortalized melanocytes. Besides, it led to the transformation of both Ba/F3 and NIH 3T3 cells as revealed by colony formation assays. Furthermore, its expression results in MAPK activation assessed by increased levels of p‐ERK protein in the cytosol of transduced cells. Treatment with Sorafenib and UO126 inhibited proliferation of Ba/F3‐SOX5‐RAF1 cells in the absence of IL3 but not the PLX 4720, a specific inhibitor of BRAF. Moreover, the tumorigenic and metastatic capacities of SOX5‐RAF1 were assessed in vivo. These results indicate that SOX5‐RAF1, a driver event for CMN development, has oncogenic capacity. Thus, sequencing of CMN transcriptomes may lead to the identification of this druggable fusion and interfere with the progression toward melanoma. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is an opportunistic pathogen and is the major cause of corneal infection worldwide that secret several virulent toxins through its type III secretion system (T3SS). In defense against pathogenic insults, epithelial cells and macrophages express antimicrobial peptides (AMPs) that are essential components of host immune response. In this study, we have determined the expression of several AMPs in patients with P. aeruginosa corneal infection. We also used an in vitro model of infection using human corneal epithelial cells and macrophages to determine the gene expression of AMPs and cellular response to wild-type and T3SS mutant P. aeruginosa. We found differential expression of several AMPs in patient samples and also found that P. aeruginosa repress AMP expression in both epithelial cells and macrophages by its T3SS in vitro. It dampens AMP expression by causing delay in NF-κB, p38 and ERK activation and inhibits reactive oxygen species generation in these cells by its T3SS. Our study show the profile of AMPs expressed during P. aeruginosa keratitis and suggest the pivotal role of the T3SS in epithelial cells and macrophages during P. aeruginosa infection. [ABSTRACT FROM AUTHOR]

Vertebrate β-catenin has two functions, as a structural component of the adherens junction in cell adhesion and as the T-cell factor (TCF) transcriptional coactivator in canonical Wnt (wingless-related integration site) signaling. These two functions are split between three of the four β-catenin-related proteins present in the round worm Caenorhabditis elegans. The fourth β-catenin-related protein, WRM-1, exhibits neither of these functions. Instead, WRM-1 binds the MAPK loss of intestine 1 (LIT-1), and these two proteins have been shown to be essential for the transcription ofWnt target genes by phosphorylating and regulating the nuclear level of the sole worm TCF protein.We showed previously thatWRM-1 binds to worm TCF and functions as the substrate-binding subunit for LIT-1. In this study, we show that phosphorylation of T220 in the activation loop is essential for LIT-1 kinase activity in vivo and in vitro. T220 can be phosphorylated either through LIT-1 autophosphorylation or directly by the upstream MAP3K MOM-4. Our data support a model in which WRM-1, which can undergo homotypic interaction, binds LIT-1 and thereby generates a kinase complex in which LIT-1 molecules are situated in a conformation enabling autophosphorylation as well as promoting phosphorylation of the T220 residue by MOM-4. In addition, we show that WRM-1 is essential for the translocation of the LIT-1 kinase complex to the nucleus, the site of its TCF substrate. To our knowledge, this is the first report of a MAP3K directly activating a MAPK by phosphorylation within the activation loop. This study should help uncover novel and as yet underappreciated functions of vertebrate β-catenin. [ABSTRACT FROM AUTHOR]

The nature of "toxic" tau in Alzheimer's disease (AD) has been unclear. During pathogenesis, the importance of tau oligomerization vs. tau phosphorylation is controversial and the investigation of both remains critical toward defining the "toxicity" of tau. The phosphorylation of tau on serines and/or threonines occurs early in the disease course and altering phosphorylation has been shown to disrupt neuropathogenesis.We have recently reported that in PC12-derived cells, tau had a role in signal transduction processes activated by NGF. By depleting tau, NGF-induced MAPK activation was attenuated and by restoring tau, MAPK activation was restored. Furthermore, the phosphorylation of tau on Thr231 was required for tau to potentiate MAPK activation. Here we report the effects of additional disease-related tau phosphorylation sites and tau isoform on the ability of tau to potentiate MAPK activation. Our findings, which tested three other sites of phosphorylation, showed that phosphorylation at these other sites mainly lessened MAPK activation; none potentiated MAPK activation. In comparing 0N3R tau to the other five brain tau isoforms, most showed a trend toward less MAPK activation, with only 2N4R tau showing significantly less activation. Since MAPK activation has been reported in AD brain and is characteristic of cell proliferation mechanisms, tau phosphorylation that promotes MAPK activation could promote cell cycle activation mechanisms. In neurons, the activation of the cell cycle leads to cell death, suggesting that abnormally phosphorylated tau can be a toxic species. The relationship between tau oligomerization and its ability to potentiate MAPK activation needs to be determined. [ABSTRACT FROM AUTHOR]

Receptor activator of NF-κ?B ligand (RANKL)-induced osteoclastogenesis is accompanied by intracellular Ca2+ mobilization in a form of oscillations, which plays essential roles by activating sequentially Ca2+ /calmodulin-dependent protein kinase, calcineurin and NFATc1, necessary in the osteoclast differentiation. However, it is not known whether Ca2+ mobilization which is evoked in RANKL-independent way induces to differentiate into osteoclasts. In present study, we investigated Ca2+ mobilization induced by aluminum fluoride (AlF4 -), a G-protein activator, with or without RANKL and the effects of AlF4 - on the osteoclastogenesis in primary cultured mouse bone marrow-derived macrophages (BMMs). We show here that AlF4 - induces intracellular Ca2+ concentration ([Ca2+]i) oscillations, which is dependent on extracellular Ca2+]i oscillations and MAPK activation which lead to enhancement of RANKL-mediated osteoclastogenesis. [ABSTRACT FROM AUTHOR]4 - with RANKL resulted in enhanced NFATc1 expression and formation of tartrate-resistant acid phosphatase (TRAP) positive multinucleated cells. Additionally, we confirmed that mitogen-activated protein kinase (MAPK) is also activated by AlF4 -. Taken together, these results demonstrate that G-protein would be a novel modulator responsible for [Ca2+]i oscillations and MAPK activation which lead to enhancement of RANKL-mediated osteoclastogenesis. [ABSTRACT FROM AUTHOR]

Estrogen-like effects of cadmium (Cd) have been reported in several animal studies, and recent epidemiological findings suggest increased risk of hormone-dependent cancers after Cd exposure. The mechanisms underlying these effects are still under investigation. Our aim was to study the effects of Cd on cellular signaling pathways in vivo with special focus on estrogen signaling and to perform benchmark dose analysis on the effects. Transgenic adult ERE-luciferase male mice were exposed subcutaneously to 0.5–500 μg CdCl2 per kg body weight (bw) or 17α-ethinylestradiol (EE2) for 3 days. These doses had no effects on organ and bw or testicular histology, indicating subtoxic exposure levels. The transgene luciferase, reporting genomic estrogen response, was significantly increased by EE2 but not by Cd. However, Cd significantly affected kinase phosphorylation and endogenous gene expression. Interestingly, gene expression changes displayed a traditional dose-response relationship, with benchmark dose levels for the expression of Mt1, Mt2, p53, c-fos, and Mdm2 being 92.9, 19.9, 7.6, 259, and 25.9 μg/kg bw, respectively, but changes in kinase phosphorylation were only detected at low exposure levels. Phosphorylation of Erk1/2 was significantly increased even in the lowest dose group, 0.5 μg/kg bw, rendering pErk1/2 a more sensitive sensor of exposure than changes in gene expression. Collectively, our data suggest that the effects triggered by Cd in vivo are markedly concentration dependent. Furthermore, we conclude that the estrogen-like effects of Cd are likely to result from a mechanism different from steroidal estrogens. [ABSTRACT FROM PUBLISHER]

Advanced ovarian cancer (OC) is not curable by surgery alone and chemotherapy is essential for its treatment. Isothiocyanates have been shown to inhibit carcinogen-induced tumorigenesis in animal models, yet no efforts have been made to determine their therapeutic potential in OC. In the present study, we investigated the mechanism of the antiproliferative and apoptotic activity of benzyl isothiocyanate (BITC) in OC. BITC inhibited the proliferation of OC cells and induced apoptosis in OC cells. Apoptosis was induced by a strong activation of caspase-3 and -9, and cleavage of PARP-1. However, caspase-8 was not activated by BITC. Cytotoxic effects of BITC were reversed by the inhibition caspase-3 and -9 specific inhibitors. BITC showed a concentration dependent decrease in the levels of Bcl-2 with a concomitant increase in Bax levels. In addition, BITC activated proapoptotic signaling by phosphorylation JNK1/2 and p38 while simultaneously inhibiting survival signaling mediated by ERK1/2 and Akt phosphorylation in a dose-dependent manner. While JNK inhibitor SP600125 and p38 inhibitor SB203580, abolished the cytotoxic effect of BITC, MEK inhibitor, PD98059 and PI3 kinase inhibitor, LY294002 failed to show such reversal indicating a critical role played by JNK1/2 and p38 signaling in apoptosis induced by BITC. In summary, our studies demonstrate that BITC inhibits proliferation of OC cells and induces apoptosis via caspase-9 and -3 pathways. BITC inhibits ERK1/2 and Akt survival signaling while simultaneously activating pro-apoptotic p38 and JNK1/2. Therefore, BITC can be potentially developed as a therapeutic agent to treat OC. [ABSTRACT FROM AUTHOR]

Abstract Recent evidence suggest that many G protein-coupled receptors (GPCR) and signalling molecules localize in microdomains of the plasma membrane. In this study, flotation gradient analysis in the absence of detergents demonstrated the presence of the metabotropic glutamate receptor type 1α (mGlu1α) in low-density caveolin-enriched membrane fractions (CEMF) in permanently transfected BHK cells. BHK-1α cells exhibit a similar pattern of staining for caveolin-1 and caveolin-2, and these two proteins show a high degree of co-localization with mGlu1α receptor as demonstrated by immunogold and confocal laser microscopy. The presence of mGlu1α in CEMF was also demonstrated by co-immunoprecipitation of mGlu1α receptor using antibodies against caveolin proteins. Activation of the mGlu1α receptor by agonist increased extracellular signal-regulated kinases phosphorylation in CEMF and not in high-density membrane fractions (HDMF), suggesting that mGlu1α receptor-mediated signal transduction could occur in caveolae-like domains. Overall, these results clearly show a molecular and functional association of mGlu1α receptor with caveolins. [ABSTRACT FROM AUTHOR]