Rheb (Ras homolog enriched in the brain) is a small GTPase protein that plays an important role in cell signaling for development of the neocortex through modulation of mTORC1 (mammalian-target-of-rapamycin-complex-1) activity. mTORC1 is known to control various biological processes including axonal growth in forming complexes at the lysosomal membrane compartment. As such, anchoring of Rheb on the lysosomal membrane via the farnesylation of Rheb at its cysteine residue (C180) is required for its promotion of mTOR activity. To test the significance of Rheb farnesylation, we overexpressed a farnesylation mutant form of Rheb, Rheb C180S, in primary rat hippocampal neurons and also in mouse embryonic neurons using in utero electroporation. Interestingly, we found that Rheb C180S maintained promotional effect of axonal elongation similar to the wild-type Rheb in both test systems. On the other hand, Rheb C180S failed to exhibit the multiple axon-promoting effect which is found in wild-type Rheb. The levels of phospho-4EBP1, a downstream target of mTORC1, were surprisingly increased in Rheb C180S transfected neurons, despite the levels of phosphorylated mTOR being significantly decreased compared to control vector transfectants. A specific mTORC1 inhibitor, rapamycin, also could not completely abolish axon elongation characteristics of Rheb C180S in transfected cells. Our data suggests that Rheb in a non-membrane compartment can promote the axonal elongation via phosphorylation of 4EBP1 and through an mTORC1-independent pathway.
Animals ; Axons ; Biological Processes ; Cysteine ; Electroporation ; GTP Phosphohydrolases ; Hand ; Membranes ; Mice ; Neocortex ; Neurons ; Phosphorylation ; Prenylation ; Protein Prenylation ; Rats ; Sirolimus ; TOR Serine-Threonine Kinases

Sanggenol P (1), a new isoprenylated flavonoid, together with nine known ones, cyclomorusin (2), morusin (3), mulberrofuran G (4), sanggenol A (5), sanggenol L (6), sanggenol N (7), cyclomulberrin (8), cyclocommunol (9) and ursolic acid (10) was isolated from Morus alba L. Sanggenol P (1) was characterized based on extensive IR, UV, 1D and 2D NMR spectroscopic analysis. Compounds 5, 6, 7 and 9 were obtained from this plant for the first time.
Flavonoids ; chemistry ; Molecular Structure ; Morus ; chemistry ; Plant Extracts ; chemistry ; Prenylation

Phytochemical investigation on an EtOH extract of the leaves of Mons yunnanensis led to the isolation and characterization of 11 isoprenylated phenolic compounds, morusyunnansin G (1), morachalcone B (2), cudraxanthone M (3), cudraxanthone D (4), sanggenofuran B(5), moracin D (6), moracin C (7), moracin I (8), demethylmoracin I (9), morachalcone A (10), and isobacachalcone (11). Compound 1 is a new compound and compounds 2-11 were isolated from this plant for the first time.
Molecular Structure ; Morus ; chemistry ; Phenols ; chemistry ; Plant Extracts ; chemistry ; Plant Leaves ; chemistry ; Prenylation

8-prenylnaringenin (8-PN) is a potent estrogen with high medicinal values. It also serves as an important precursor for many prenylated flavonoids. Microbial synthesis of 8-PN is mainly hindered by the low catalytic activity of prenyltransferases (PTS) and insufficient supply of precursors. In this work, a SfN8DT-1 from Sophora flavescens was used to improve the efficiency of (2S)-naringenin prenylation. The predicted structure of SfN8DT-1 showed that its main body is comprised of 9 α-helices and 8 loops, along with a long side chain formed by nearly 120 amino acids. SfN8DT-1 mutants with different side-chain truncated were tested in Saccharomyces cerevisiae. A mutant expressing the truncated enzyme at K62 site, designated as SfND8T-1-t62, produced the highest 8-PN titer. Molecular docking of SfN8DT-1-t62 with (2S)-naringenin and dimethylallyl diphosphate (DMAPP) showed that K185 was a potentially crucial residue. Alanine scanning within a range of 0.5 nm around these two substrates showed that the mutant K185A may decrease its affinity to substrates, which also indicated K185 was a potentially critical residue. Besides, the mutant K185W enhanced the affinity to ligands implied by the simulated saturation mutation, while the saturated mutation of K185 showed a great decrease in 8-PN production, indicating K185 is vital for the activity of SfN8DT-1. Subsequently, overexpressing the key genes of Mevalonate (MVA) pathway further improved the titer of 8-PN to 31.31 mg/L, which indicated that DMAPP supply is also a limiting factor for 8-PN synthesis. Finally, 44.92 mg/L of 8-PN was produced in a 5 L bioreactor after 120 h, which is the highest 8-PN titer reported to date.
Dimethylallyltranstransferase/metabolism* ; Flavonoids/metabolism* ; Molecular Docking Simulation ; Prenylation ; Saccharomyces cerevisiae/metabolism* ; Sophora/metabolism*

Farnesyl transferase inhibitor(FTI) disrupt farnesylation of ras protein and thus, suppress tumor growth in vivo. The ras oncogene is thought to contribute to tumor development directly by promoting tumor cell proliferation and indirectly by stimulating vascular endothelial growth from Cinnamomum Cassia Blume(CB2`-ph) interferes with angiogenesis, we studied the effect of CB2`-ph on rabbit corneal angiogenesis induced by VEGF. A hygrogel disk containing 250ng of VEGF was implated intrastromally in the superior cornea of 16 NZW rabbit eyes. All eyes received a second intrastromal disk, randomized to contain either 40 micro gram of CB2`-ph(n=8) or phosphate-buffered saline(PBS)(n=8). Both disks were positioned side-by-side, 1.2mm from the superior limbus. Each eye was examined daily by two masked observers and assigned an angiogenesis score based on number and length of new blood vessels. At 3, 5 and 7 days postimplantation of VEGF disks, eyes treated with CB2`=ph showed mean angiogenesis score of 2.0 +/- 1.7, 13.4 +/- 6.6 and 23.8 +/- 11.3, respectively, while PBS=treated controls scored 8.6 +/- 4.4, 30.3 +/- 20.8 and 52.2 +/- 26.9, respectively(p<0.05, Wilcoxon signed rank test). In a rabbit corneal pocket assay, CB2`-ph appears to be effective on VEGF-induced corneal angiogenesusis in the model.
Blood Vessels ; Cell Proliferation ; Cinnamomum aromaticum ; Cornea ; Corneal Neovascularization* ; Genes, ras ; Masks ; Prenylation ; Rabbits ; Transferases* ; Vascular Endothelial Growth Factor A

BACKGROUND: Although so many experimental trials have been done to improve the redifferentiation and responsiveness of radioiodide therapy, they have not yet yielded any satisfactory results. As statins inhibit both farnesylation and geranylgeranylation, they have been reported to have an antineoplastic and redifferentiation effect in experimental and clinical studies. In this study, we investigated the relationship between statins and the alteration of the NIS expression and, TPC-1 cell apotosis to evaluate the possibility of using statins as adjuvant therapeutic agents for papillary thyroid cancer. METHODS: We used the TPC-1 cell lines for our experiments. Cell viabilities were measured by CCK-8. The degrees of apoptosis and, the expressions of NIS mRNA and NIS protein were measured by flow cytometry, semi quantitative RT-PCR and Western blot assay. RESULTS: Increased levels of NIS mRNA and NIS protein were observed under therapeutic blood concentrations (concentrations of simvastatin: 20, 50, 80 nM, concentrations of atorvastatin: 50, 80,110 nM), but the dose-response relationship was only manifested within simvastatin. The TPC-1 cells showed a concentration dependent decrease of viability and an increase of apoptosis not under therapeutic blood concentrations, but under excessively high concentrations (after treatment with 10-50 microM of atorvastatin and with 1-10 microM of simvastatin). CONCLUSION: The results of this study show that effective therapeutic blood concentrations of simvastatin and atorvastatin can give a favorable effect on the NIS expression under effective therapeutic blood concentrations. Therefore, we demonstrated the possibility that simvastatin and atorvastatin might have an important role as adjuvant therapeutic agents to improve the responsiveness of radioiodide therapy for papillary thyroid cancer. Further studies are needed to clarify this issue.
Apoptosis ; Blotting, Western ; Cell Line ; Cell Survival ; Flow Cytometry ; Heptanoic Acids ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; Prenylation ; Pyrroles ; RNA, Messenger ; Simvastatin ; Sincalide ; Symporters ; Thyroid Neoplasms ; Atorvastatin Calcium