[Show abstract][Hide abstract] ABSTRACT: Ergothioneine is a histidine thiol derivative. Its mycobacterial biosynthetic pathway has five steps (EgtA-E catalysis) with two novel reactions: a mononuclear nonheme iron enzyme (EgtB) catalyzed oxidative C-S bond formation and a PLP-mediated C-S lyase (EgtE) reaction. Our bioinformatic and biochemical analyses indicate that the fungus Neurospora crassa has a more concise ergothioneine biosynthetic pathway because its nonheme iron enzyme, Egt1, makes use of cysteine instead of γ-Glu-Cys as the substrate. Such a change of substrate preference eliminates the competition between ergothioneine and glutathione biosyntheses. In addition, we have identified the N. crassa C-S lyase (NCU11365) and reconstituted its activity in vitro, which makes the future ergothioneine production through metabolic engineering feasible.
[Show abstract][Hide abstract] ABSTRACT: The peanut is a special plant for its aerial flowering but subterranean fructification. The failure of peg penetration into the soil leads to form aerial pod and finally seed abortion. However, the mechanism of seed abortion during aerial pod development remains obscure. Here, a comparative transcriptome analysis between aerial and subterranean pods at different developmental stages was produced using a customized NimbleGen microarray representing 36,158 unigenes. By comparing 4 consecutive time-points, totally 6,203 differentially expressed genes, 4,732 stage-specific expressed genes and 2,401 specific expressed genes only in aerial or subterranean pods were identified in this study. Functional annotation showed their mainly involvement in biosynthesis, metabolism, transcription regulation, transporting, stress response, photosynthesis, signal transduction, cell division, apoptosis, embryonic development, hormone response and light signaling, etc. Emphasis was focused on hormone response, cell apoptosis, embryonic development and light signaling relative genes. These genes might function as potential candidates to provide insights into seed abortion during aerial pod development. Ten candidate genes were validated by Real-time RT-PCR. Additionally, consistent with up-regulation of auxin response relative genes in aerial pods, endogenous IAA content was also significantly increased by HPLC analysis. This study will further provide new molecular insight that auxin and auxin response genes potentially contribute to peanut seed and pod development.
[Show abstract][Hide abstract] ABSTRACT: Given the threat of drug resistance, there is an acute need for new classes of antimalarial agents that act via a unique mechanism of action relative to currently used drugs. We have identified a set of druglike compounds within the Tres Cantos Anti-Malarial Set (TCAMS) which likely act via inhibition of a Plasmodium aspartic protease. Structure− activity relationship analysis and optimization of these aminohydantoins demonstrate that these compounds are potent nanomolar inhibitors of the Plasmodium aspartic proteases PM-II and PM-IV and likely one or more other Plasmodium aspartic proteases. Incorporation of a bulky group, such as a cyclohexyl group, on the aminohydantion N-3 position gives enhanced antimalarial potency while reducing inhibition of human aspartic proteases such as BACE. We have identified compound 8p (CWHM-117) as a promising lead for optimization as an antimalarial drug with a low molecular weight, modest lipophilicity, oral bioavailability, and in vivo antimalarial activity in mice. KEYWORDS: Malaria, antimalarial, aminohydantoin, medicinal chemistry, aspartic protease inhibitors M alaria is a devastating mosquito-borne infectious disease caused by a parasite of the genus Plasmodium, placing over one billion people at high risk for infection. According to the World Health Organization, there were an estimated 225 million cases of malaria in 2010 with 610,000−971,000 deaths. 1 Especially hard hit is sub-Saharan Africa, where 80% of the deaths occur, mostly in children under the age of 5 years old. Although there are a number of drugs used to treat the disease, resistance to most of these drugs is widespread. 2 The introduction of artemisinin and artemisinin combination therapies (ACTs) in 2005 has begun to reverse the trend. While this is a good sign, there have been reports of resistance to artemisinin in Southeast Asia. 3 As a consequence, there is an urgent push for developing antimalarial therapies targeting novel modes of action. Drug discovery efforts in this area have been recently reviewed. 4−6
[Show abstract][Hide abstract] ABSTRACT: The peanut plant produces flowers aerially, while develops the fruits and seeds underground. Pod swelling is a vital process of peanut pod and seed development only occurring after the gynophore carrying the ovule into the soil. The failure of gynophore penetration into the soil leads to suppression of pod swelling initiation. However, the molecular mechanism underlying the process remains unknown. A comparative proteome analysis between developing aerial and subterranean pods at various developmental stages was performed using 2-DE approach. 47 significantly differentially expressed spots were selected to further identification by MALDI-TOF-TOF MS. They were corresponded to 31 distinct proteins, suggesting that many identified spots were modified in post-translation. Functional annotation revealed their involvement in twelve important biological processes, such as photosynthesis, oxidative stress response, lignin synthesis, fatty acid biosynthesis, glycolysis, protein catabolic process, cellular metabolic process, and regulation process, etc. Furthermore, 10 identified proteins were validated by Real-time RT-PCR analysis. Several photosynthesis and oxidative stress proteins displayed elevated expression levels in aerial pods. Otherwise, enzymes in lignin synthesis and ubiquitin proteasome system were down-accumulation in subterranean pods. These enzymes might function as potential candidate proteins and play critical roles to regulate pods swelling and development.
Pod swelling plays a crucial role in peanut fruit and seed development. However, a large number of aerial pods can't form normal pods due to suppression of swelling initiation by the failure of penetration into the soil, thereby causing to seed yield loss. Limited knowledge is available underlying molecular mechanism regulating initiation of swelling in peg tips and pod development. The results generated in this study may provide evidence for some functional proteins as potential candidates to pod swelling and new molecular insights to improve our understanding of pod development under light and darkness conditions, which may contribute valuable information to high yield breeding in future.
Journal of proteomics 07/2013; · 5.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Peanut (Arachis hypogaea L.) is one of the most susceptible host crops to Aspergillus flavus invasion and subsequent aflatoxin contamination. In this report, a new member of PR10 family putative resistant gene (designated as ARAhPR10, No. EU661964.1) encoding a PR10 protein was isolated and characterized. Analysis of qRT-PCR showed that the ex- pression of ARAhPR10 was induced by pre-harvested A. flavus infection, but no significant difference was observed between resistant genotype “GT-C20” and susceptible genotype “Yueyou 7”. Seven transgenic peanut lines expressing the ARAhPR10 gene under the control of 35S promoter were obtained using the Agrobacterium tumefaciens-mediated method. Real time RT-PCR results showed that the expression level of the ARAhPR10 was significantly higher and the A. flavus infection and aflatoxin content were significantly lower in seeds of transgenic lines than that of the wild type. A significant negative correlation between ARAhPR10 expression at transcript level and seeds aflatoxin production was observed. Combining the previous results, it is suggested that ARAhPR10 expression play an important role in peanut host resistance to A. flavus infection and aflatoxin producing.
[Show abstract][Hide abstract] ABSTRACT: Low genetic diversity makes peanut (Arachis hypogaea L.) very vulnerable to plant pathogens, causing severe yield loss and reduced seed quality. Several hundred partial genomic DNA sequences as nucleotide-binding-site leucine-rich repeat (NBS-LRR) resistance genes (R) have been identified, but a small portion with expressed transcripts has been found. We aimed to identify resistance gene analogs (RGAs) from peanut expressed sequence tags (ESTs) and to develop polymorphic markers. The protein sequences of 54 known R genes were used to identify homologs from peanut ESTs from public databases. A total of 1,053 ESTs corresponding to six different classes of known R genes were recovered, and assembled 156 contigs and 229 singletons as peanut-expressed RGAs. There were 69 that encoded for NBS-LRR proteins, 191 that encoded for protein kinases, 82 that encoded for LRR-PK/transmembrane proteins, 28 that encoded for Toxin reductases, 11 that encoded for LRR-domain containing proteins and 4 that encoded for TM-domain containing proteins. Twenty-eight simple sequence repeats (SSRs) were identified from 25 peanut expressed RGAs. One SSR polymorphic marker (RGA121) was identified. Two PCR-based markers (Ahsw-1 and Ahsw-2) developed from RGA013 were homologous to the Tomato Spotted Wilt Virus (TSWV) resistance gene. All three markers were mapped on the same linkage group AhIV. These expressed RGAs are the source for RGA-tagged marker development and identification of peanut resistance genes.
Journal of Integrative Plant Biology 02/2013; · 3.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Germin-like superfamily members are ubiquitously expressed in various plant species and play important roles in plant development and defense. Although several GLPs have been identified in peanut (Arachis hypogaea L.), their roles in development and defense remain unknown. In this research, we study the spatiotemporal expression of AhGLPs in peanut and their functions in plant defense.
We have identified three new AhGLP members (AhGLP3b, AhGLP5b and AhGLP7b) that have distinct but very closely related DNA sequences. The spatial and temporal expression profiles revealed that each peanut GLP gene has its distinct expression pattern in various tissues and developmental stages. This suggests that these genes all have their distinct roles in peanut development. Subcellular location analysis demonstrated that AhGLP2 and 5 undergo a protein transport process after synthesis. The expression of all AhGLPs increased in responding to Aspergillus flavus infection, suggesting AhGLPs' ubiquitous roles in defense to A. flavus. Each AhGLP gene had its unique response to various abiotic stresses (including salt, H2O2 stress and wound), biotic stresses (including leaf spot, mosaic and rust) and plant hormone stimulations (including SA and ABA treatments). These results indicate that AhGLPs have their distinct roles in plant defense. Moreover, in vivo study of AhGLP transgenic Arabidopsis showed that both AhGLP2 and 3 had salt tolerance, which made transgenic Arabidopsis grow well under 100 mM NaCl stress.
For the first time, our study analyzes the AhGLP gene expression profiles in peanut and reveals their roles under various stresses. These results provide an insight into the developmental and defensive roles of GLP gene family in peanut.
PLoS ONE 01/2013; 8(4):e61722. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background
Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) have been successfully used to knock out endogenous genes in stem cell research. However, the deficiencies of current gene-based delivery systems may hamper the clinical application of these nucleases. A new delivery method that can improve the utility of these nucleases is needed.
In this study, we utilized a cell-penetrating peptide-based system for ZFN and TALEN delivery. Functional TAT-ZFN and TAT-TALEN proteins were generated by fusing the cell-penetrating TAT peptide to ZFN and TALEN, respectively. However, TAT-ZFN was difficult to purify in quantities sufficient for analysis in cell culture. Purified TAT-TALEN was able to penetrate cells and disrupt the gene encoding endogenous human chemokine (C-C motif) receptor 5 (CCR5, a co-receptor for HIV-1 entry into cells). Hypothermic treatment greatly enhanced the TAT-TALEN-mediated gene disruption efficiency. A 5% modification rate was observed in human induced pluripotent stem cells (hiPSCs) treated with TAT-TALEN as measured by the Surveyor assay.
TAT-TALEN protein-mediated gene disruption was applicable in hiPSCs and represents a promising technique for gene knockout in stem cells. This new technique may advance the clinical application of TALEN technology.
[Show abstract][Hide abstract] ABSTRACT: Legumes are important food crops worldwide, contributing to more than 33% of human dietary protein. The production of crop legumes is frequently impacted by abiotic and biotic stresses. It is therefore important to identify genes conferring resistance to biotic stresses and tolerance to abiotic stresses that can be used to both understand molecular
mechanisms of plant response to the environment and to accelerate crop improvement. Recent advances in genomics offer a range of approaches such as the sequencing of genomes and transcriptomes, gene expression microarray as well as RNA-seq
based gene expression profiling, and map-based cloning for the identification and isolation of biotic and abiotic stress responsive genes in several crop legumes. These candidate stress associated genes should provide insights into the molecular
mechanisms of stress tolerance and ultimately help to develop legume varieties with improved stress tolerance and productivity under adverse conditions. This review provides an overview on recent advances in the functional genomics of crop legumes that includes the discovery as well as validation of candidate genes.
[Show abstract][Hide abstract] ABSTRACT: The failure of peg penetration into the soil leads to seed abortion in peanut. Knowledge of genes involved in these processes is comparatively deficient. Here, we used RNA-seq to gain insights into transcriptomes of aerial and subterranean pods. More than 2 million transcript reads with an average length of 396 bp were generated from one aerial (AP) and two subterranean (SP1 and SP2) pod libraries using pyrosequencing technology. After assembly, sets of 49 632, 49 952 and 50 494 from a total of 74 974 transcript assembly contigs (TACs) were identified in AP, SP1 and SP2, respectively. A clear linear relationship in the gene expression level was observed between these data sets. In brief, 2194 differentially expressed TACs with a 99.0% true-positive rate were identified, among which 859 and 1068 TACs were up-regulated in aerial and subterranean pods, respectively. Functional analysis showed that putative function based on similarity with proteins catalogued in UniProt and gene ontology term classification could be determined for 59 342 (79.2%) and 42 955 (57.3%) TACs, respectively. A total of 2968 TACs were mapped to 174 KEGG pathways, of which 168 were shared by aerial and subterranean transcriptomes. TACs involved in photosynthesis were significantly up-regulated and enriched in the aerial pod. In addition, two senescence-associated genes were identified as significantly up-regulated in the aerial pod, which potentially contribute to embryo abortion in aerial pods, and in turn, to cessation of swelling. The data set generated in this study provides evidence for some functional genes as robust candidates underlying aerial and subterranean pod development and contributes to an elucidation of the evolutionary implications resulting from fruit development under light and dark conditions.
[Show abstract][Hide abstract] ABSTRACT: Over the past five decades, cultivated peanut in China has been
subjected to strong artificial selection in breeding programmes. To
investigate the impact of artificial selection on expression diversity, we
compared gene expression profiles in pod and leaf of five widespread
cultivars in Southern China. In terms of tissues, hierarchical clustering
analysis revealed that expression data of pod and leaf generated
different dendrograms owing to artificial selection. K-means analysis
also showed that there were 16 gene expression patterns in leaf, while
only eight in pod. In considering cultivars, a cultivar specificity index
(s) was employed to characterize expression patterns, which suggested
that genes having 0.15 < s < 0.85 constituted >80% of all expression
patterns. Additionally, the diversity of gene expression in pod
among cultivars of the �YY7� pedigree decreased from 23.8% to 3.9%.
Taken together, nucleotide polymorphisms in regulatory elements
owing to artificial selection led to low-expression polymorphisms in
both tissues and cultivars, contributed to the narrow genetic diversity
and might be a driving force behind the breeding of cultivated peanut
[Show abstract][Hide abstract] ABSTRACT: C-C chemokine receptor type 5 (CCR5) is a major co-receptor for the entry of human immunodeficiency virus type-1 (HIV-1) into target cells. Human hematopoietic stem cells (hHSCs) with naturally occurring CCR5 deletions (Δ32) or artificially disrupted CCR5 have shown potential for curing acquired immunodeficiency syndrome (AIDS). However, Δ32 donors are scarce, heterologous bone marrow transplantation is not exempt of risks, and genetic engineering of autologous hHSCs is not trivial. Here, we have disrupted the CCR5 locus of human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) using specific zinc finger nucleases (ZFNs) combined with homologous recombination. The modified hESCs and hiPSCs retained pluripotent characteristics and could be differentiated in vitro into CD34(+) cells that formed all types of hematopoietic colonies. Our results suggest the potential of using patient-specific hHSCs derived from ZFN-modified hiPSCs for treating AIDS.
Human gene therapy 02/2012; 23(2):238-42. · 4.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: With 9 figures and 1 table AbstractOver the past five decades, cultivated peanut in China has been subjected to strong artificial selection in breeding programmes. To investigate the impact of artificial selection on expression diversity, we compared gene expression profiles in pod and leaf of five widespread cultivars in Southern China. In terms of tissues, hierarchical clustering analysis revealed that expression data of pod and leaf generated different dendrograms owing to artificial selection. K‐means analysis also showed that there were 16 gene expression patterns in leaf, while only eight in pod. In considering cultivars, a cultivar specificity index (τ) was employed to characterize expression patterns, which suggested that genes having 0.15 80% of all expression patterns. Additionally, the diversity of gene expression in pod among cultivars of the ‘YY7’ pedigree decreased from 23.8% to 3.9%. Taken together, nucleotide polymorphisms in regulatory elements owing to artificial selection led to low‐expression polymorphisms in both tissues and cultivars, contributed to the narrow genetic diversity and might be a driving force behind the breeding of cultivated peanut.
[Show abstract][Hide abstract] ABSTRACT: Only a few genetic maps based on recombinant inbred line (RIL) and backcross (BC) populations have been developed for tetraploid groundnut. The marker density, however, is not very satisfactory especially in the context of large genome size (2800 Mb/1C) and 20 linkage groups (LGs). Therefore, using marker segregation data for 10 RILs and one BC population from the international groundnut community, with the help of common markers across different populations, a reference consensus genetic map has been developed. This map is comprised of 897 marker loci including 895 simple sequence repeat (SSR) and 2 cleaved amplified polymorphic sequence (CAPS) loci distributed on 20 LGs (a01-a10 and b01-b10) spanning a map distance of 3, 863.6 cM with an average map density of 4.4 cM. The highest numbers of markers (70) were integrated on a01 and the least number of markers (21) on b09. The marker density, however, was lowest (6.4 cM) on a08 and highest (2.5 cM) on a01. The reference consensus map has been divided into 20 cM long 203 BINs. These BINs carry 1 (a10_02, a10_08 and a10_09) to 20 (a10_04) loci with an average of 4 marker loci per BIN. Although the polymorphism information content (PIC) value was available for 526 markers in 190 BINs, 36 and 111 BINs have at least one marker with >0.70 and >0.50 PIC values, respectively. This information will be useful for selecting highly informative and uniformly distributed markers for developing new genetic maps, background selection and diversity analysis. Most importantly, this reference consensus map will serve as a reliable reference for aligning new genetic and physical maps, performing QTL analysis in a multi-populations design, evaluating the genetic background effect on QTL expression, and serving other genetic and molecular breeding activities in groundnut.
PLoS ONE 01/2012; 7(7):e41213. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Four new β-carboline alkaloids, designated marinacarbolines A-D (1-4), two new indolactam alkaloids, 13-N-demethyl-methylpendolmycin (5) and methylpendolmycin-14-O-α-glucoside (6), and the three known compounds 1-acetyl-β-carboline (7), methylpendolmycin (8), and pendolmycin (9) were obtained from the fermentation broth of Marinactinospora thermotolerans SCSIO 00652, a new actinomycete belonging to the family Nocardiopsaceae. Their structures were elucidated by extensive MS and 1D and 2D NMR spectroscopic data analyses. The structure of compound 1 was further confirmed by single-crystal X-ray crystallography. The new compounds 1-6 were inactive against a panel of eight tumor cell lines (IC50>50 μM) but exhibited antiplasmodial activities against Plasmodium falciparum lines 3D7 and Dd2, with IC50 values ranging from 1.92 to 36.03 μM.
Journal of Natural Products 10/2011; 74(10):2122-7. · 3.29 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: T lymphocytes play a vital role in antimalaria immunity, but there is little information about the role of T cells in malaria infection. In order to explore the profile of T cells in malaria immunity, we infected Chinese rhesus macaques with the malaria parasite (Plasmodium cynomolgi) and examined the dynamics of T cell subsets. Both repeated and long-term infections were involved. Our results showed that the monkeys in the repeated infection group acquired protective immunity through primary infection, which was evidenced by a much lower parasitemia, milder anemia, and milder fever during reinfection; the monkeys in the long-term infection group also developed protective immunity, but this was not sufficient to eliminate the parasite. The total counts of leukocytes, neutrophils, CD3+ T cells, CD4+ or CD8+ T cells, and naïve and memory CD4+ and CD8+ T cells declined during the acute phase of malaria but increased after the parasite was controlled. The total number of activated CD4+ T cells significantly increased during malaria in animals with a long-term infection, which remained at least 3 months after the termination of malaria. However, the activated CD4+ T cells decreased during the acute phase of infection in the repeated infection group and converted to preinfection levels after malaria was cured. Regulatory CD4+ T cells continued to increase during the malaria infections and quickly reverted to preinfection levels after the parasite was controlled. Our study provides a systematic analysis of the kinetic profiles of T lymphocyte subsets during malaria infections and provides some experimental insight into malaria immunology.
Parasitology Research 08/2011; 110(2):961-9. · 2.85 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Tumor-associated macrophages (TAMs) are the most abundant immune cells within the tumor stroma and play a crucial role in tumor development. Although clinical investigations indicate that high levels of macrophage (MΦ) infiltration into tumors are associated with a poor prognosis, the exact role played by TAMs during tumor development remains unclear. The present study aimed to investigate dynamic changes in TAM major histocompatibility complex (MHC) class II expression levels and to assess the effects of these changes on tumor progression.
Significant inhibition of tumor growth in the murine hepatocellular carcinoma Hepa1-6 model was closely associated with partial TAM depletion. Strikingly, two distinct TAM subsets were found to coexist within the tumor microenvironment during Hepa1-6 tumor development. An MHC class II(hi) TAM population appeared during the early phase of tumor development and was associated with tumor suppression; however, an MHC class II(low) TAM population became increasingly predominant as the tumor progressed.
Tumor progression was positively correlated with increasing infiltration of the tumor tissues by MHC class II(low) TAMs. Thus, targeting the transition of MΦ may be a novel strategy for drug development and immunotherapy.
[Show abstract][Hide abstract] ABSTRACT: Aspergillus flavus and A. parasiticus infect peanut seeds and produce aflatoxins, which are associated with various diseases in domestic animals and humans throughout the world. The most cost-effective strategy to minimize aflatoxin contamination involves the development of peanut cultivars that are resistant to fungal infection and/or aflatoxin production. To identify peanut Aspergillus-interactive and peanut Aspergillus-resistance genes, we carried out a large scale peanut Expressed Sequence Tag (EST) project which we used to construct a peanut glass slide oligonucleotide microarray. The fabricated microarray represents over 40% of the protein coding genes in the peanut genome. For expression profiling, resistant and susceptible peanut cultivars were infected with a mixture of Aspergillusflavus and parasiticus spores. The subsequent microarray analysis identified 62 genes in resistant cultivars that were up-expressed in response to Aspergillus infection. In addition, we identified 22 putative Aspergillus-resistance genes that were constitutively up-expressed in the resistant cultivar in comparison to the susceptible cultivar. Some of these genes were homologous to peanut, corn, and soybean genes that were previously shown to confer resistance to fungal infection. This study is a first step towards a comprehensive genome-scale platform for developing Aspergillus-resistant peanut cultivars through targeted marker-assisted breeding and genetic engineering.
[Show abstract][Hide abstract] ABSTRACT: The antimalarial activity of the human immunodeficiency virus protease inhibitors indinavir and saquinavir was evaluated in rhesus macaques for the first time. Indinavir effectively suppressed the growth of Plasmodium cynomolgi and Plasmodium knowlesi in vivo after a 7- or 3-day treatment, respectively, with clinically relevant doses, whereas saquinavir showed only weak activity against P. cynomolgi.
Antimicrobial Agents and Chemotherapy 06/2011; 55(6):3039-42. · 4.57 Impact Factor