H3 histamine receptor-mediated activation of protein kinase calpha inhibits the growth of cholangiocarcinoma in vitro and in vivo

Histamine regulates functions via four receptors (HRH1, HRH2, HRH3, and HRH4). The D-myo-inositol 1,4,5-trisphosphate (IP(3))/Ca(2+)/protein kinase C (PKC)/mitogen-activated protein kinase pathway regulates cholangiocarcinoma growth. We evaluated the role of HRH3 in the regulation of cholangiocarcinoma growth. Expression of HRH3 in intrahepatic and extrahepatic cell lines, normal cholangiocytes, and human tissue arrays was measured. In Mz-ChA-1 cells stimulated with (R)-(alpha)-(-)-methylhistamine dihydrobromide (RAMH), we measured (a) cell growth, (b) IP(3) and cyclic AMP levels, and (c) phosphorylation of PKC and mitogen-activated protein kinase isoforms. Localization of PKC alpha was visualized by immunofluorescence in cell smears and immunoblotting for PKC alpha in cytosol and membrane fractions. Following knockdown of PKC alpha, Mz-ChA-1 cells were stimulated with RAMH before evaluating cell growth and extracellular signal-regulated kinase (ERK)-1/2 phosphorylation. In vivo experiments were done in BALB/c nude mice. Mice were treated with saline or RAMH for 44 days and tumor volume was measured. Tumors were excised and evaluated for proliferation, apoptosis, and expression of PKC alpha, vascular endothelial growth factor (VEGF)-A, VEGF-C, VEGF receptor 2, and VEGF receptor 3. HRH3 expression was found in all cells. RAMH inhibited the growth of cholangiocarcinoma cells. RAMH increased IP(3) levels and PKC alpha phosphorylation and decreased ERK1/2 phosphorylation. RAMH induced a shift in the localization of PKC alpha expression from the cytosolic domain into the membrane region of Mz-ChA-1 cells. Silencing of PKC alpha prevented RAMH inhibition of Mz-ChA-1 cell growth and ablated RAMH effects on ERK1/2 phosphorylation. In vivo, RAMH decreased tumor growth and expression of VEGF and its receptors; PKC alpha expression was increased. RAMH inhibits cholangiocarcinoma growth by PKC alpha-dependent ERK1/2 dephosphorylation. Modulation of PKC alpha by histamine receptors may be important in regulating cholangiocarcinoma growth. (Mol Cancer Res 2009;7(10):1704-13

Insights into the regulation of DMSP synthesis in the diatom Thalassiosira pseudonana through APR activity, proteomics and gene expression analyses on cells acclimating to changes in salinity, light and nitrogen

Despite the importance of dimethylsulphoniopropionate (DMSP) in the global sulphur cycle and climate regulation, the biological pathways underpinning its synthesis in marine phytoplankton remain poorly understood. The intracellular concentration of DMSP increases with increased salinity, increased light intensity and nitrogen starvation in the diatom Thalassiosira pseudonana. We used these conditions to investigate DMSP synthesis at the cellular level via analysis of enzyme activity, gene expression and proteome comparison. The activity of the key sulphur assimilatory enzyme, adenosine 5′- phosphosulphate reductase was not coordinated with increasing intracellular DMSP concentration. Under all three treatments coordination in the expression of sulphur assimilation genes was limited to increases in sulphite reductase transcripts. Similarly, proteomic 2D gel analysis only revealed an increase in phosphoenolpyruvate carboxylase following increases in DMSP concentration. Our findings suggest that increased sulphur assimilation might not be required for increased DMSP synthesis, instead the availability of carbon and nitrogen substrates may be important in the regulation of this pathway. This contrasts with the regulation of sulphur metabolism in higher plants, which generally involves upregulation of several sulphur assimilatory enzymes. In T. pseudonana changes relating to sulphur metabolism were specific to the individual treatments and, given that little coordination was seen in transcript and protein responses across the three growth conditions, different patterns of regulation might be responsible for the increase in DMSP concentration seen under each treatment

The origin of secondary heavy rare earth element enrichment in carbonatites: Constraints from the evolution of the Huanglongpu district, China

publisher: Elsevier articletitle: The origin of secondary heavy rare earth element enrichment in carbonatites: Constraints from the evolution of the Huanglongpu district, China journaltitle: Lithos articlelink: http://dx.doi.org/10.1016/j.lithos.2018.02.027 content_type: article copyright: © 2018 The Authors. Published by Elsevier B.V.Copyright: © 2018 Published by Elsevier B.V. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The file attached is the Published/publisher’s pdf version of the articl

Genetic regulation of glucoraphanin accumulation in Beneforté® broccoli

Diets rich in broccoli (Brassica oleracea var italica) have been associated with maintenance of cardiovascular health and reduction in risk of cancer. These health benefits have been attributed to glucoraphanin that specifically accumulates in broccoli. The development of broccoli with enhanced concentrations of glucoraphanin may deliver greater health benefits. Three high-glucoraphanin F1 broccoli hybrids were developed in independent programmes through genome introgression from the wild species Brassica villosa. Glucoraphanin and other metabolites were quantified in experimental field trials. Global SNP analyses quantified the differential extent of B. villosa introgression The high-glucoraphanin broccoli hybrids contained 2.5–3 times the glucoraphanin content of standard hybrids due to enhanced sulphate assimilation and modifications in sulphur partitioning between sulphur-containing metabolites. All of the high-glucoraphanin hybrids possessed an introgressed B. villosa segment which contained a B. villosa Myb28 allele. Myb28 expression was increased in all of the high-glucoraphanin hybrids. Two high-glucoraphanin hybrids have been commercialised as Beneforte broccoli. The study illustrates the translation of research on glucosinolate genetics from Arabidopsis to broccoli, the use of wild Brassica species to develop cultivars with potential consumer benefits, and the development of cultivars with contrasting concentrations of glucoraphanin for use in blinded human intervention studie

The sulfur pathway and diagnosis of sulfate depletion in grapevine

Sulfur is an essential nutrient to all plant species. Plants assimilate sulfur in a well-described pathway, which has been taken up by roots. Regulatory mech- anism has been the subject of many research papers. However, recent studies highlighted differences between crop plants and the model plant Arabidopsis thaliana. Our work focuses on the identification of genes involved in the sulfur metabolism in the Vitis vinifera genome, and their response to sulfur deficiency and other abiotic stress endured by grapevine in the field, namely water stress. Here, we describe the identification and brief characterization of the first assimilation enzymes involved in the sulfur pathway, the enzyme responsible for sulfur activa- tion, ATP sulfurylase (ATPS), and the two enzymes that reduce sulfate to sulfide, Adenosine 50-phosphosulate reductase (APR) and Sulfite reductase (SiR). A reduc- tion was observed in the number of ATPS and APR isoforms identified in V. vinifera genome when compared to A. thaliana or Glycine max genomes. Two ATPS isoforms were present in the Vitis genome, of which only ATPS1 transcript was detected in the tested tissues, and one APR isoform, suggesting an absence of redundancy in the role of both enzymes. ATPS1, APR and SiR transcript level was up-regulated in response to 2 days exposure to sulfur deficiency in V. vinifera cell cultures, which was completely reversed by the addition of GSH to the culture medium. Apparently, oxidative stress triggered GSH has a pivotal role in the regulation of ATPS1, APR and SiR transcription level, since their up-regulation was observed in mRNA from field grapevine berries under water stress, which is known to induce oxidative stress.info:eu-repo/semantics/publishedVersio

Chloroplasts lacking class I glutaredoxins are functional but show a delayed recovery of protein cysteinyl redox state after oxidative challenge

Redox status of protein cysteinyl residues is mediated via glutathione (GSH)/glutaredoxin (GRX) and thioredoxin (TRX)-dependent redox cascades. An oxidative challenge can induce post-translational protein modifications on thiols, such as protein S-glutathionylation. Class I GRX are small thiol-disulfide oxidoreductases that reversibly catalyse S-glutathionylation and protein disulfide formation. TRX and GSH/GRX redox systems can provide partial backup for each other in several subcellular compartments, but not in the plastid stroma where TRX/light-dependent redox regulation of primary metabolism takes place. While the stromal TRX system has been studied at detail, the role of class I GRX on plastid redox processes is still unknown. We generate knockout lines of GRXC5 as the only chloroplast class I GRX of the moss Physcomitrium patens. While we find that PpGRXC5 has high activities in GSH-dependent oxidoreductase assays using hydroxyethyl disulfide or redox-sensitive GFP2 as substrates in vitro, Δgrxc5 plants show no detectable growth defect or stress sensitivity, in contrast to mutants with a less negative stromal EGSH (Δgr1). Using stroma-targeted roGFP2, we show increased protein Cys steady state oxidation and decreased reduction rates after oxidative challenge in Δgrxc5 plants in vivo, indicating kinetic uncoupling of the protein Cys redox state from EGSH. Compared to wildtype, protein Cys disulfide formation rates and S-glutathionylation levels after H2O2 treatment remained unchanged. Lack of class I GRX function in the stroma did not result in impaired carbon fixation. Our observations suggest specific roles for GRXC5 in the efficient transfer of electrons from GSH to target protein Cys as well as negligible cross-talk with metabolic regulation via the TRX system. We propose a model for stromal class I GRX function in efficient catalysis of protein dithiol/disulfide equilibria upon redox steady state alterations affecting stromal EGSH and highlight the importance of identifying in vivo target proteins of GRXC5