S-locus F-Box Protein as Pollen S determinant Targets Non-self S-RNase underlying Self-Incompatibility in Citrus.

Self-incompatibility (SI) is a crucial mechanism that prevents self-fertilization and inbreeding in flowering plants. Citrus exhibits SI regulated by a polymorphic S-locus containing an S-RNase gene and multiple S-locus F-box (SLF) genes. It has been documented that S-RNase functions as the pistil S determinant, but there is no direct evidence that the SLFs closely linked with S-RNase function as pollen S determinants in Citrus. This study assembled the genomes of two pummelo (Citrus grandis) plants and obtained three novel complete and well-annotated S-haplotypes and isolated 36 SLF or SLF-like alleles on the S-loci. Phylogenetic analysis of 138 SLFs revealed that the SLFs were classified into 12 types, including six types with divergent or missing alleles. Furthermore, transformation experiments verified that the conserved S6-SLF7a protein can lead the transition of SI to self-compatibility (SC) by recognizing non-self S8-RNase in 'Mini-Citrus' plants (S7S8 and S8S29, Fortunella hindsii), a model plant for citrus gene function studies. In vitro assays demonstrated interactions between SLFs of different S haplotypes and the Skp1-Cullin1-F-box subunit CgSSK1 protein. This study provides direct evidence that SLF controls the pollen function in Citrus, demonstrating its role in the 'non-self-recognition' SI system.

Low and high doses of oral maslinic acid protect against Parkinson's disease via distinct gut microbiota-related mechanisms.

The use of oral agents that can modify the gut microbiota (GM) could be a novel preventative or therapeutic option for Parkinson's disease (PD). Maslinic acid (MA), a pentacyclic triterpene acid with GM-dependent biological activities when it is taken orally, has not yet been reported to be effective against PD. The present study found both low and high dose MA treatment significantly prevented dopaminergic neuronal loss in a classical chronic PD mouse model by ameliorating motor functions and improving tyrosine hydroxylase expressions in the substantia nigra pars compacta (SNpc) and increasing dopamine and its metabolite homovanillic acid levels in the striatum. However, the effects of MA in PD mice were not dose-responsive, since similar beneficial effects for low and high doses of MA were observed. Further mechanism studies indicated that low dose MA administration favored probiotic bacterial growth in PD mice, which helped to increase striatal serotonin, 5-hydroxyindole acetic acid, and γ-aminobutyric acid levels. High dose MA treatment did not influence GM composition in PD mice but significantly inhibited neuroinflammation as indicated by reduced levels of tumor necrosis factor alpha and interleukin 1ß in the SNpc; moreover, these effects were mainly mediated by microbially-derived acetic acid in the colon. In conclusion, oral MA at different doses protected against PD via distinct mechanisms related to GM. Nevertheless, our study lacked in-depth investigations of the underlying mechanisms involved; future studies will be designed to further delineate the signaling pathways involved in the interactive actions between different doses of MA and GM.

Tegillarca granosa extract Haishengsu inhibits tumor activity via a mitochondrial­mediated apoptotic pathway.

Haishengsu (HSS) is an active natural extract isolated from Tegillarca granosa, which has previously been demonstrated to inhibit the proliferation of several types of cancer cells in vitro. Our previous study indicated that HSS may induce apoptosis to suppress growth of human hepatocellular carcinoma BEL­7402 cells by activating Fas pathway. The present study demonstrated that HSS treatment induces the in vitro apoptosis of BEL­7402 cells via the mitochondrial­mediated apoptotic pathway detected by DNA fragmentation assay, caspase activity assay and transmission electron microscopy assay, and inhibits tumor xenograft growth in vivo. Alterations in apoptotic regulatory proteins were detected, including decreased expression of B­cell lymphoma2 (Bcl­2), upregulation of Bcl­2­associated X protein and mitochondrial cytochrome c release, and downstream activation of apoptotic signaling. Furthermore, apoptotic induction was caspase­dependent, as indicated by cleavage of the caspase substrate, poly (ADP­ribose) polymerase. Oral administration of 62.5­250 mg/kg HSS markedly educed the growth of hepatocellular carcinoma tumor xenografts in nude mice. In addition, immunohistochemical staining for caspase­3 protein and transmission electron microscopy further indicated the induction of apoptosis in these tumor tissues. Taken together, the present study demonstrated that HSS may effectively induce apoptosis to suppress the growth of BEL­7402 cells in vitro and in vivo, and therefore may hold promise for further development as a novel cancer therapy.

Physcion inhibits the metastatic potential of human colorectal cancer SW620 cells in vitro by suppressing the transcription factor SOX2.

AIM: Physcion, an anthraquinone derivative, exhibits hepatoprotective, anti-inflammatory, anti-microbial and anti-cancer activities. In this study we examined whether and how physcion inhibited metastatic potential of human colorectal cancer cells in vitro. METHODS: Human colorectal cancer cell line SW620 was tested. Cell migration and invasion were assessed using a wound healing and Transwell assay, respectively. The expression levels of transcription factor SOX2 in the cells were modulated with shRNA targeting SOX2 and SOX2 overexpressing plasmid. The expression of target molecules involved in epithelial-mesenchymal transition (EMT) process and the signaling pathways was determined with Western blots or qRT-PCR. ROS levels were measured using DCF-DA. RESULTS: Physcion (2.5, 5 mol/L) did not affect the cell viability, but dose-dependently inhibited the cell adhesion, migration and invasion. Physcion also inhibited the EMT process in the cells, as evidenced by the increased epithelial marker E-cadherin expression, and by decreased expression of mesenchymal markers N-cadherin, vimentin, fibronectin and α-SMA, as well as transcriptional repressors Snail, Slug and Twist. Physcion suppressed the expression of SOX2, whereas overexpression of SOX2 abrogated the inhibition of physcion on metastatic behaviors. Physcion markedly increased ROS production and phosphorylation of AMPK and GSK3ß in the cells, whereas the AMPK inhibitor compound C or the ROS inhibitor NAC abolished the inhibition of physcion on metastatic behaviors. CONCLUSION: Physcion inhibits the metastatic potential of human colorectal cancer cells in vitro via activating ROS/AMPK/GSK3ß signaling pathways and suppressing SOX2.

Comprehensive analysis of expressed sequence tags from the pulp of the red mutant 'Cara Cara' navel orange (Citrus sinensis Osbeck).

Expressed sequence tag (EST) analysis of the pulp of the red-fleshed mutant 'Cara Cara' navel orange provided a starting point for gene discovery and transcriptome survey during citrus fruit maturation. Interpretation of the EST datasets revealed that the mutant pulp transcriptome held a high section of stress responses related genes, such as the type III metallothionein-like gene (6.0%), heat shock protein (2.8%), Cu/Zn superoxide dismutase (0.8%), late embryogenesis abundant protein 5 (0.8%), etc. 133 transcripts were detected to be differentially expressed between the red mutant and its orange-color wild genotype 'Washington' via digital expression analysis. Among them, genes involved in metabolism, defense/stress and signal transduction were statistical overrepresented. Fifteen transcription factors, composed of NAM, ATAF, and CUC transcription factor (NAC); myeloblastosis (MYB); myelocytomatosis (MYC); basic helix-loop-helix (bHLH); basic leucine zipper (bZIP) domain members, were also included. The data reflected the distinct expression profile and the unique regulatory module associated with these two genotypes. Eight differently expressed genes analyzed in digital were validated by quantitative real-time polymerase chain reaction. For structural polymorphism, both simple sequence repeats and single nucleotide polymorphisms (SNP) loci were surveyed; dinucleotide presentation revealed a bias toward AG/GA/TC/CT repeats (52.5%), against GC/CG repeats (0%). SNPs analysis found that transitions (73%) outnumbered transversions (27%). Seventeen potential cultivar-specific and 387 heterozygous SNP loci were detected from 'Cara Cara' and 'Washington' EST pool.

Olfactory ensheathing cells genetically modified to secrete GDNF to promote spinal cord repair.

Olfactory ensheathing cell (OEC) transplantation has emerged as a very promising therapy for spinal cord repair. In this study, we tested the ability of genetically modified OECs to secrete high levels of glial cell line-derived neurotrophic factor (GDNF) to promote spinal cord repair. The GDNF gene was transduced into OECs using a retroviral-based system. The engineered OECs were first characterized by their ability to express and secrete biologically active GDNF in vitro. After implantation into the spinal cord of adult rats with complete spinal cord transection, OEC survival and GDNF production were examined. The locomotor functions of animals were assessed and axon regeneration was evaluated at the morphological level. To our knowledge, we report for the first time that the genetically modified OECs are capable of producing GDNF in vivo to significantly improve recovery after spinal cord injury (SCI). This work combined the outgrowth-promoting property of OECs with the neuroprotective effects of the additionally overexpressed neurotrophic factors and opens new avenues for the treatment of SCI.