Lycium barbarum polysaccharides: Protective effects against heat-induced damage of rat testes and H2O2-induced DNA damage in mouse testicular cells and beneficial effect on sexual behavior and reproductive function of hemicastrated rats

Lycium barbarum, a famous Chinese medicinal herb, has a long history of use as a traditional remedy for male infertility. Polysaccharides are the most important functional constituent in L. barbarum fruits. We systematically investigated the effect of L. barbarum polysaccharides (LBP) on rat testis damage induced by a physical factor (43 degrees C heat exposure), on DNA damage of mouse testicular cells induced by a chemical factor (H2O2), and on sexual behavior and reproductive function of hemicastrated male rats. The results showed that LBP provided a protective effect against the testicular tissue damage induced by heat exposure. When compared with negative control, a suitable concentration of LBP significantly increased testis and epididymis weights, improved superoxide dismutase (SOD) activity, and raised sexual hormone levels in the damaged rat testes. LBP had a dose-dependent protective effect against DNA oxidative damage of mouse testicular cells induced by H2O2. LBP improved the copulatory performance and reproductive function of hemicastrated male rats, such as shortened penis erection latency and mount latency, regulated secretion of sexual hormones and increased hormone levels, raised accessory sexual organ weights, and improved sperm quantity and quality. The present findings support the folk reputation of L. barbarum fruits as an aphrodisiac and fertility-facilitating agent, and provide scientific evidence for a basis for the extensive use of L. barbarum fruits as a traditional remedy for male infertility in China.     

Synthesis, characterization, cytotoxic activities, and DNA-binding properties of the La(III) complex with Naringenin Schiff-base

A new Naringenin Schiff-base ligand (H3L) and its complex, [La(H2L)2(NO3).3H2O], have been synthesized and characterized on the basis of elemental analyses, molar conductivities, mass spectra, 1H NMR, thermogravimetry/differential thermal analysis (TG-DTA), UV spectra, and IR spectra. Spectrometric titrations, ethidium bromide displacement experiments, and viscosity measurements indicate that the two compounds, especially the La(III) complex, strongly bind with calf-thymus DNA, presumably via an intercalation mechanism. The intrinsic binding constants of the La(III) complex and ligand with DNA were 1.83 x 10(7) and 9.46 x 10(5) M(-1), respectively. Comparative cytotoxic activities of the La(III) complex and ligand were also determined by MTT [3-(4,5-dimethyl-2-thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide] and SRB (sulforhodamine B) methods. The results showed that the La(III) complex had significant cytotoxic activity against the tested cells.     

The proteolytic function of the Arabidopsis 26S proteasome is required for specifying leaf adaxial identity

Polarity formation is central to leaf morphogenesis, and several key genes that function in adaxial-abaxial polarity establishment have been identified and characterized extensively. We previously reported that Arabidopsis thaliana ASYMMERTIC LEAVES1 (AS1) and AS2 are important in promoting leaf adaxial fates. We obtained an as2 enhancer mutant, asymmetric leaves enhancer3 (ae3), which demonstrated pleiotropic plant phenotypes, including a defective adaxial identity in some leaves. The ae3 as2 double mutant displayed severely abaxialized leaves, which were accompanied by elevated levels of leaf abaxial promoting genes FILAMENTOUS FLOWER, YABBY3, KANADI1 (KAN1), and KAN2 and a reduced level of the adaxial promoting gene REVOLUTA. We identified AE3, which encodes a putative 26S proteasome subunit RPN8a. Furthermore, double mutant combinations of as2 with other 26S subunit mutations, including rpt2a, rpt4a, rpt5a, rpn1a, rpn9a, pad1, and pbe1, all displayed comparable phenotypes with those of ae3 as2, albeit with varying phenotypic severity. Since these mutated genes encode subunits that are located in different parts of the 26S proteasome, it is possible that the proteolytic function of the 26S holoenzyme is involved in leaf polarity formation. Together, our findings reveal that posttranslational regulation is essential in proper leaf patterning.     

Characterization of the structures involved in localization of the SUN proteins to the nuclear envelope and the centrosome

The nuclear envelope forms a selective barrier that separates the cytoplasm from the nucleus. During mitosis the nuclear envelope breaks down so that the microtubule network can form contacts with the kinetochore and guide chromosome segregation. Previous studies have suggested a model in which the centrosome and the microtubule network may play a role in nuclear envelope breakdown through as yet unidentified interactions with proteins localized to the nuclear envelope. In the current study we characterized a nuclear envelope protein SUN2 and identified a substructure involved in its localization to the nuclear envelope. We found that a structurally related protein, SUN1, may be localized to the nuclear envelope through a different mechanism. Furthermore, the SUN2 protein can form different assemblies, including homodimers and heterodimers with SUN1. Finally, we provide evidence indicating that SUN1 and SUN2 may form a physical interaction between the nuclear envelope and the centrosome.     

Genomic approaches to drug discovery

Considerable progress has been made in exploiting the enormous amount of genomic and genetic information for the identification of potential targets for drug discovery and development. New tools that incorporate pathway information have been developed for gene expression data mining to reflect differences in pathways in normal and disease states. In addition, forward and reverse genetics used in a high-throughput mode with full-length cDNA and RNAi libraries enable the direct identification of components of signaling pathways. The discovery of the regulatory function of microRNAs highlights the importance of continuing the investigation of the genome with sophisticated tools. Furthermore, epigenetic information including DNA methylation and histone modifications that mediate important biological processes add to the possibilities to identify novel drug targets and patient populations that will benefit from new therapies.     

Array-based comparative genomic hybridization and copy number variation in cancer research

Array-based comparative genomic hybridization (aCGH) is a molecular cytogenetic technique used in detecting and mapping DNA copy number alterations. aCGH is able to interrogate the entire genome at a previously unattainable, high resolution and has directly led to the recent appreciation of a novel class of genomic variation: copy number variation (CNV) in mammalian genomes. All forms of DNA variation/polymorphism are important for studying the basis of phenotypic diversity among individuals. CNV research is still at its infancy, requiring careful collation and annotation of accumulating CNV data that will undoubtedly be useful for accurate interpretation of genomic imbalances identified during cancer research.     

The oligomeric stromal proteome of Arabidopsis thaliana chloroplasts

This study presents an analysis of the stromal proteome in its oligomeric state extracted from highly purified chloroplasts of Arabidopsis thaliana. 241 proteins (88% with predicted cTP), mostly assembled in oligomeric complexes, were identified by mass spectrometry with emphasis on distinguishing between paralogues. This is critical because different paralogues in a gene family often have different subcellular localizations and/or different expression patterns and functions. The native protein masses were determined for all identified proteins. Comparison with the few well characterized stromal complexes from A. thaliana confirmed the accuracy of the native mass determination, and by extension, the usefulness of the native mass data for future in-depth protein interaction studies. Resolved protein interactions are discussed and compared with an extensive collection of native mass data of orthologues in other plants and bacteria. Relative protein expression levels were estimated from spot intensities and also provided estimates of relative concentrations of individual proteins. No such quantification has been reported so far. Surprisingly proteins dedicated to chloroplast protein synthesis, biogenesis, and fate represented nearly 10% of the total stroma protein mass. Oxidative pentose phosphate pathway, glycolysis, and Calvin cycle represented together about 75%, nitrogen assimilation represented 5-7%, and all other pathways such as biosynthesis of e.g. fatty acids, amino acids, nucleotides, tetrapyrroles, and vitamins B(1) and B(2) each represented less than 1% of total protein mass. Several proteins with diverse functions outside primary carbon metabolism, such as the isomerase ROC4, lipoxygenase 2 involved in jasmonic acid biosynthesis, and a carbonic anhydrase (CA1), were surprisingly abundant in the range of 0.75-1.5% of the total stromal mass. Native images with associated information are available via the Plastid Proteome Database.     

Distribution, levels and phosphorylation of Raf-1 in Alzheimer's disease

Extracellular signal-regulated kinase (ERK), a member of the mitogen-activated protein kinase pathway, has been increasingly implicated in the pathogenesis of Alzheimer's disease due to its critical role in brain function. While we previously demonstrated that ERK is activated in Alzheimer's disease, the upstream cascade leading to its activation had not been fully examined. In this study, we focused on Raf-1, one of the physiological activators of the ERK pathway. Raf-1 is activated by phosphorylation at Ser338 and Tyr340/341 and inhibited by phosphorylation at Ser259. Interestingly, phosphorylation at all three sites on Raf-1 was increased as evidenced by both immunocytochemistry and immunoblot analysis in Alzheimer's disease brains compared to age-matched controls. Both phospho-Raf-1 (Ser259) and phospho-Raf-1 (Ser338) were localized to intracytoplasmic granular structures, whereas phospho-Raf-1 (Tyr340/341) was localized to neurofibrillary tangles and granules in pyramidal neurons in Alzheimer's disease hippocampus. There is extensive overlap between phospho-Raf-1 (Ser338) and phospho-Mek1/2, the downstream effector of Raf-1, suggestive of a mechanistic link. Additionally, increased levels of Raf-1 are associated with Ras and MEK1 in Alzheimer's disease as evidenced by its coimmunoprecipitation with Ras and Mek1, respectively. Based on these findings, we speculate that Raf-1 is activated to effectively mediate Ras-dependent signals in Alzheimer's disease.