The neurotransmitter serotonin (5-HT) plays an important role in the regulation of multiple events in the CNS. We demonstrated recently a coupling between the 5-HT 4 receptor and the heterotrimeric G13-protein resulting in RhoA-dependent neurite retraction and cell rounding (Ponimaskin et al., 2002). In the present study, we identified G12 as an additional G-protein that can be activated by another member of serotonin receptors, the 5-HT 7 receptor. Expression of 5-HT 7 receptor induced constitutive and agonist-dependent activation of a serum response element-mediated gene transcription through G12-mediated activation of small GTPases. In NIH3T3 cells, activation of the 5-HT 7 receptor induced filopodia formation via a Cdc42-mediated pathway correlating with RhoA-dependent cell rounding. In mouse hippocampal neurons, activation of the endogenous 5-HT 7 receptors significantly increased neurite length, whereas stimulation of 5-HT 4 receptors led to a decrease in the length and number of neurites. These data demonstrate distinct roles for 5-HT 7 R/G12 and 5-HT 4 R/G13 signaling pathways in neurite outgrowth and retraction, suggesting that serotonin plays a prominent role in regulating the neuronal cytoarchitecture in addition to its classical role as neurotransmitter.

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A highly efficient microprojectile transformation system for sorghum (Sorghum bicolor L.) has been developed by using immature embryos (IEs) of inbred line Tx430. Co-bombardment was performed with the neomycin phosphotransferase II (nptII) gene and the green fluorescent protein (gfp) gene, both under the control of the maize ubiquitin1 (ubi1) promoter. After optimization of both tissue culture media and parameters of microprojectile transformation, 25 independent transgenic events were obtained from 121 bombarded IEs. The average transformation frequency (the total number of independent transgenic events divided by the total number of bombarded IEs) was 20.7% in three independent experiments. Transgenic events were confirmed by both PCR screening and Southern hybridization of genomic DNA from primary transgenics (T0). More than 90% of transformants were fertile and displayed normal morphology in a containment glasshouse. Co-transformation rate of the nptII and gfp genes was 72% in these experiments. The segregation of nptII and gfp in T1 progenies was observed utilizing fluorescence microscopy and geneticin selection of seedlings indicating both were inherited in the T1 generation. The transformation procedure, from initiating IEs to planting putative transgenic plantlets in the glasshouse, was completed within 11–16 weeks, and was approximately threefold more efficient than the previously reported best sorghum transformation system.

Polymorphonuclear neutrophil (PMN) extravasation requires selectin-mediated tethering, intercellular adhesion molecule-1 (ICAM-1)–dependent firm adhesion, and platelet/endothelial cell adhesion molecule 1 (PECAM-1)–mediated transendothelial migration. An important unanswered question is whether ICAM-1–activated signaling contributes to PMN transmigration mediated by PECAM-1. We tested this concept and the roles of endothelial nitric oxide synthase (eNOS) and Src activated by PMN ligation of ICAM-1 in mediating PECAM-1–dependent PMN transmigration. We observed that lung PMN infiltration in vivo induced in carrageenan-injected WT mice was significantly reduced in ICAM-1−/− and eNOS−/− mice. Crosslinking WT mouse ICAM-1 expressed in human endothelial cells (ECs), but not the phospho-defective Tyr518Phe ICAM-1 mutant, induced SHP-2–dependent Src Tyr530 dephosphorylation that resulted in Src activation. ICAM-1 activation also stimulated phosphorylation of Akt (p-Ser473) and eNOS (p-Ser1177), thereby increasing NO production. PMN migration across EC monolayers was abolished in cells expressing the Tyr518Phe ICAM-1 mutant or by pretreatment with either the Src inhibitor PP2 or eNOS inhibitor L-NAME. Importantly, phospho–ICAM-1 induction of Src signaling induced PECAM-1 Tyr686 phosphorylation and increased EC surface anti–PECAM-1 mAb-binding activity. These results collectively show that ICAM-1–activated Src and eNOS signaling sequentially induce PECAM-1–mediated PMN transendothelial migration. Both Src and eNOS inhibition may be important therapeutic targets to prevent or limit vascular inflammation.

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Synthesis and accumulation of the storage lipid triacylglycerol in vegetative plant tissues has emerged as a promising strategy to meet the world's future need for vegetable oil. Sorghum (Sorghum bicolor) is a particularly attractive target crop given its high biomass, drought resistance and C photosynthesis. While oilseed-like triacylglycerol levels have been engineered in the C model plant tobacco, progress in C monocot crops has been lagging behind. In this study, we report the accumulation of triacylglycerol in sorghum leaf tissues to levels between 3 and 8.4% on a dry weight basis depending on leaf and plant developmental stage. This was achieved by the combined overexpression of genes encoding the Zea mays WRI1 transcription factor, Umbelopsis ramanniana UrDGAT2a acyltransferase and Sesamum indicum Oleosin-L oil body protein. Increased oil content was visible as lipid droplets, primarily in the leaf mesophyll cells. A comparison between a constitutive and mesophyll-specific promoter driving WRI1 expression revealed distinct changes in the overall leaf lipidome as well as transitory starch and soluble sugar levels. Metabolome profiling uncovered changes in the abundance of various amino acids and dicarboxylic acids. The results presented here are a first step forward towards the development of sorghum as a dedicated biomass oil crop and provide a basis for further combinatorial metabolic engineering.

Background Retrospective analysis of patients undergoing cancer surgery suggests the use of regional anesthesia may reduce cancer recurrence and improve survival. Amide-linked local anesthetics have anti-inflammatory properties, although the mechanism of action in this regard is unclear. As inflammatory processes involving Src tyrosine protein kinase and intercellular adhesion molecule-1 are important in tumor growth and metastasis, we hypothesized that amide-linked local anesthetics may inhibit inflammatory Src-signaling involved in migration of adenocarcinoma cells. Methods NCI-H838 lung cancer cells were incubated with Tumor Necrosis Factor-α in absence/presence of ropivacaine, lidocaine, or chloroprocaine (1nM-100μM). Cell migration and total cell lysate Src-activation and Intercellular Adhesion Molecule-1 phosphorylation were assessed. The role of voltage-gated sodium-channels in the mechanism of local anesthetic effects was also evaluated. Results Ropivacaine treatment (100μM) of H838 cells for 20 minutes decreased basal Src activity by 62% (p=0.003), and both ropivacaine and lidocaine co-administered with Tumor Necrosis Factor-α statistically significantly decreased Src-activation and Intercellular Adhesion Molecule-1 phosphorylation, whereas chloroprocaine had no such effect. Migration of these cells at 4 hours was inhibited by 26% (p=0.005) in presence of 1μM ropivacaine and 21% by 1μM lidocaine (p=0.004). These effects of ropivacaine and lidocaine were independent of voltage-gated sodium-channel inhibition. Conclusions This study indicates that amide-, but not ester-linked local anesthetics may provide beneficial anti-metastatic effects. The observed inhibition of NCI-H838 cell migration by lidocaine and ropivacaine was associated with the inhibition of Tumor Necrosis Factor-α-induced Src-activation and Intercellular Adhesion Molecule-1 phosphorylation, providing the first evidence of a molecular mechanism which appears to be independent of their known role as sodium-channel blockers.

The unexpected phenomenon and mechanism of the alkali metal involved NH3 selective catalysis are reported. Incorporation of K+ (4.22 wt %) in the tunnels of α‐MnO2 greatly improved its activity at low temperature (50–200 °C, 100 % conversion of NOx vs. 50.6 % conversion over pristine α‐MnO2 at 150 °C). Experiment and theory demonstrated the atomic role of incorporated K+ in α‐MnO2. Results showed that K+ in the tunnels could form a stable coordination with eight nearby Osp3 atoms. The columbic interaction between the trapped K+ and O atoms can rearrange the charge population of nearby Mn and O atoms, thus making the topmost five‐coordinated unsaturated Mn cations (Mn5c, the Lewis acid sites) more positive. Therefore, the more positively charged Mn5c can better chemically adsorb and activate the NH3 molecules compared with its pristine counterpart, which is crucial for subsequent reactions.