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Biomedical subjects

Li Zheng

Publications and source records attributed to Li Zheng.

At least 19 recordsLinked to original sources

Chromosomal-level genome assembly of minute pirate bug Orius nagaii Yasunaga, 1993 (Hemiptera: Anthocoridae).

Species of the genus Orius, diminutive predatory insects that act as natural enemies of other arthropods, are frequently employed in agricultural pest management for controlling various pests, such as thrips, mites, aphids, whiteflies, etc. However, the scarcity of high-quality genomic resources for these predators hinders our comprehension of their population evolution and predation ecology. Consequently, we assembled and annotated a chromosomal-scale genome of Orius nagaii by collating PacBio and Illumina sequencing and Hi-C genomic analysis techniques. The final genome assembly size 152.62 Mb, with scaffold and contig N50 lengths of 11.53 and 2.39 Mb, respectively. It is organized into 12 pairs of autosomes and a pair of XY sex chromosomes. The quality assessment of the genomic data with BUSCO revealed a completeness of 98.5% (n = 1,367). Also, 11,917 protein-coding genes were discovered, with 94.28% of them having functional annotations. The high-quality genome of O. nagaii produced serves as a valuable resource for comprehending the interactions between predatory natural enemies and hosts, along with their evolutionary trajectories.

Animals↗

Okazaki fragment maturation involves α-segment error editing by the mammalian FEN1/MutSα functional complex.

During nuclear DNA replication, proofreading-deficient DNA polymerase α (Pol α) initiates Okazaki fragment synthesis with lower fidelity than bulk replication by proofreading-proficient Pol δ or Pol ε. Here, we provide evidence that the exonuclease activity of mammalian flap endonuclease (FEN1) excises Pol α replication errors in a MutSα-dependent, MutLα-independent mismatch repair process we call Pol α-segment error editing (AEE). We show that MSH2 interacts with FEN1 and facilitates its nuclease activity to remove mismatches near the 5' ends of DNA substrates. Mouse cells and mice encoding FEN1 mutations display AEE deficiency, a strong mutator phenotype, enhanced cellular transformation, and increased cancer susceptibility. The results identify a novel role for FEN1 in a specialized mismatch repair pathway and a new cancer etiological mechanism.

Animals↗

The TIGR Rice Genome Annotation Resource: improvements and new features.

In The Institute for Genomic Research Rice Genome Annotation project (http://rice.tigr.org), we have continued to update the rice genome sequence with new data and improve the quality of the annotation. In our current release of annotation (Release 4.0; January 12, 2006), we have identified 42,653 non-transposable element-related genes encoding 49,472 gene models as a result of the detection of alternative splicing. We have refined our identification methods for transposable element-related genes resulting in 13,237 genes that are related to transposable elements. Through incorporation of multiple transcript and proteomic expression data sets, we have been able to annotate 24 799 genes (31,739 gene models), representing approximately 50% of the total gene models, as expressed in the rice genome. All structural and functional annotation is viewable through our Rice Genome Browser which currently supports 59 tracks. Enhanced data access is available through web interfaces, FTP downloads and a Data Extractor tool developed in order to support discrete dataset downloads.

DNA Transposable Elements↗

Concerted action of exonuclease and Gap-dependent endonuclease activities of FEN-1 contributes to the resolution of triplet repeat sequences (CTG)n- and (GAA)n-derived secondary structures formed during maturation of Okazaki fragments.

There is much evidence to indicate that FEN-1 efficiently cleaves single-stranded DNA flaps but is unable to process double-stranded flaps or flaps adopting secondary structures. However, the absence of Fen1 in yeast results in a significant increase in trinucleotide repeat (TNR) expansion. There are then two possibilities. One is that TNRs do not always form stable secondary structures or that FEN-1 has an alternative approach to resolve the secondary structures. In the present study, we test the hypothesis that concerted action of exonuclease and gap-dependent endonuclease activities of FEN-1 play a role in the resolution of secondary structures formed by (CTG)n and (GAA)n repeats. Employing a yeast FEN-1 mutant, E176A, which is deficient in exonuclease (EXO) and gap endonuclease (GEN) activities but retains almost all of its flap endonuclease (FEN) activity, we show severe defects in the cleavage of various TNR intermediate substrates. Precise knock-in of this point mutation causes an increase in both the expansion and fragility of a (CTG)n tract in vivo. Taken together, our biochemical and genetic analyses suggest that although FEN activity is important for single-stranded flap processing, EXO and GEN activities may contribute to the resolution of structured flaps. A model is presented to explain how the concerted action of EXO and GEN activities may contribute to resolving structured flaps, thereby preventing their expansion in the genome.

Alanine↗

Analysis of bacteria degradation products of methyl parathion by liquid chromatography/electrospray time-of-flight mass spectrometry and gas chromatography/mass spectrometry.

The biodegradation of the organophosphorus insecticide methyl parathion (MP) in aqueous environment by bacteria isolated from river sediment has been studied. Two species of bacteria which show strong MP degradation ability are identified as Shewanella and Vibrio parahaemolyticus. The biodegradation of MP proceeded rapidly with the formation of a series of intermediate products, which were analyzed using a combination of GC/MS and HPLC/ESI-TOFMS techniques. The major products tentatively identified include a series of reduced products of MP. Results demonstrate that the coupling of TOFMS to HPLC enhances further the capability of LC-MS in the identification of polar organic species in complex environmental samples. Degradation pathways leading to the formation of these products are proposed which involves first the reduction of nitro to amino group in MP, followed by combination with some intrinsic matters of bacteria. The mechanism and products from biodegradation are quite different from those of photocatalytic process for which the main intermediates included methyl paraoxon and 4-nitrophenol.

Biotransformation↗

Plasmodium yoelii: distinct CD4(+)CD25(+) regulatory T cell responses during the early stages of infection in susceptible and resistant mice.

The outcome of experimental murine infection with different strains of malaria parasites, ranging from spontaneous cure to death, depends largely on the establishment of effective Th1 responses during the early stages of infection. Here we describe the disparity in CD4(+)CD25(+) regulatory T cell (Treg) responses during the early stages of infection with the highly virulent Plasmodium yoelii 17XL strain in susceptible (BALB/c) and resistant (DBA/2) mice. An increased proportion of Tregs 3-4 days post inoculation, co-occurring with elevated IL-10 levels, is observed in BALB/c but not in DBA/2 mice. These findings suggest that Treg proliferation might be causally associated with the suppression of Th1 responses during early malaria infection, leading to increase parasitemia and mortality in BALB/c mice, possibly in an IL-10-dependent manner.

Animals↗

Expressed sequence tags from loblolly pine embryos reveal similarities with angiosperm embryogenesis.

The process of embryogenesis in gymnosperms differs in significant ways from the more widely studied process in angiosperms. To further our understanding of embryogenesis in gymnosperms, we have generated Expressed Sequence Tags (ESTs) from four cDNA libraries constructed from un-normalized, normalized, and subtracted RNA populations of zygotic and somatic embryos of loblolly pine (Pinus taeda L.). A total of 68,721 ESTs were generated from 68,131 cDNA clones. Following clustering and assembly, these sequences collapsed into 5,274 contigs and 6,880 singleton sequences for a total of 12,154 non-redundant sequences. Searches of a non-identical amino acid database revealed a putative homolog for 9,189 sequences, leaving 2,965 sequences with no known function. More extensive searches of additional plant sequence data sets revealed a putative homolog for all but 1,388 (11.4%) of the sequences. Using gene ontologies, a known function could be assigned for 5,495 of the 12,154 total non-redundant sequences with 13,633 associations in total assigned. When compared to approximately 72,000 sequences in a collated P. taeda transcript assembly derived from >245,000 ESTs derived from root, xylem, stem, needles, pollen cone, and shoot ESTs, 3,458 (28.5%) of the non-redundant embryo sequences were unique and thereby provide a valuable addition to development of a complete loblolly pine transcriptome. To assess similarities between angiosperm and gymnosperm embryo development, we examined our EST collection for putative homologs of angiosperm genes implicated in embryogenesis. Out of 108 angiosperm embryogenesis-related genes, homologs were present for 83 of these genes suggesting that pine contains similar genes for embryogenesis and that our RNA sampling methods were successful. We also identified sequences from the pine embryo transcriptome that have no known function and may contribute to the programming of gene expression and embryo development.

Amino Acid Sequence↗

Side population cells isolated from porcine dental pulp tissue with self-renewal and multipotency for dentinogenesis, chondrogenesis, adipogenesis, and neurogenesis.

Dental pulp has the potential to form dentin as a regenerative response to caries. This regeneration is mediated by stem/progenitor cells. Thus, stem cell therapy might be of potential utility in induction of reparative dentin. We isolated side population (SP) cells from dental pulp based on the exclusion of the DNA binding dye Hoechst 33342 by flow cytometry and compared its self-renewal capacities and multipotency with non-SP cells and primary pulp cells. The cumulative cell number of the SP cells was greater than the non-SP cells and primary pulp cells. Bmi1 was continuously expressed in SP cells, suggesting longer proliferative lifespan and self-renewal capacity of SP cells. Next, the maintenance of the multilineage differentiation potential of pulp SP cells was investigated. Expression of type II collagen and aggrecan confirmed chondrogenic conversion (30%) of SP cells. SP cells expressed peroxisome proliferator-activated receptor gamma and adaptor protein 2, showing adipogenic conversion. Expression of mRNA and proteins of neurofilament and neuromodulin confirmed neurogenic conversion (90%). These results demonstrate that pulp SP cells maintain multilineage differentiation potential. We further examined whether bone morphogenetic protein 2 (BMP2) could induce differentiation of pulp SP cells into odontoblasts. BMP2 stimulated the expression of dentin sialophosphoprotein (Dspp) and enamelysin in three-dimensional pellet cultures. Autogenous transplantation of the Bmp2-supplemented SP cells on the amputated pulp stimulated the reparative dentin formation. Thus, adult pulp contains SP cells, which are enriched for stem cell properties and useful for cell therapy with BMP2 for dentin regeneration.

Adipogenesis↗

DIXDC1 isoform, l-DIXDC1, is a novel filamentous actin-binding protein.

Ccd1, a DIX domain containing Zebrafish protein involved in neural patterning, is a positive regulator of the Wnt signaling pathway. DIXDC1, the human homolog of Ccd1, has two predominant isoforms. The short form (s-DIXDC1) has a similar amino acid sequence compared with Ccd1, while the long form (l-DIXDC1) contains an extra N-terminal sequence containing a calponin-homology (CH) domain, suggesting additional interaction with actin that we have performed detailed analysis in this report. We show that mRNA expression of both DIXDC1 isoforms can be detected in various adult tissues by Northern blot analysis and is most abundant in cardiac and skeletal muscles. Both endogenous and ectopically expressed l-DIXDC1, but not s-DIXDC1, in cultured mammalian cells is localized to actin stress fibers at the filament ends in focal adhesion plaques. More importantly, l-DIXDC1 can directly bind to filamentous actin both in vitro and in vivo and the binding is mediated via a novel actin-binding domain (ABD) from amino acid 127 to 300. Thus, our data provide the first evidence that l-DIXDC1 may act as a novel branching component in the Wnt signaling pathway targeting both beta-catenin-TCF complex for gene expression and cytoskeleton for regulating dynamics of actin filaments.

Actin Cytoskeleton↗

The DNA-protein interaction modes of FEN-1 with gap substrates and their implication in preventing duplication mutations.

Flap endonuclease-1 (FEN-1) is a structure-specific nuclease best known for its involvement in RNA primer removal and long-patch base excision repair. This enzyme is known to possess 5'-flap endo- (FEN) and 5'-3' exo- (EXO) nuclease activities. Recently, FEN-1 has been reported to also possess a gap endonuclease (GEN) activity, which is possibly involved in apoptotic DNA fragmentation and the resolution of stalled DNA replication forks. In the current study, we compare the kinetics of these activities to shed light on the aspects of DNA structure and FEN-1 DNA-binding elements that affect substrate cleavage. By using DNA binding deficient mutants of FEN-1, we determine that the GEN activity is analogous to FEN activity in that the single-stranded DNA region of DNA substrates interacts with the clamp region of FEN-1. In addition, we show that the C-terminal extension of human FEN-1 likely interacts with the downstream duplex portion of all substrates. Taken together, a substrate-binding model that explains how FEN-1, which has a single active center, can have seemingly different activities is proposed. Furthermore, based on the evidence that GEN activity in complex with WRN protein cleaves hairpin and internal loop substrates, we suggest that the GEN activity may prevent repeat expansions and duplication mutations.

Amino Acid Sequence↗

WdChs1p, a class II chitin synthase, is more responsible than WdChs2p (Class I) for normal yeast reproductive growth in the polymorphic, pathogenic fungus Wangiella (Exophiala) dermatitidis.

The chitin synthase gene WdCHS1 was isolated from a partial genomic DNA library of the pathogenic polymorphic fungus Wangiella dermatitidis. Sequencing showed that WdCHS1 encoded a class II chitin synthase composed of 988 amino acids. Disruption of WdCHS1 produced strains that were hyperpigmented in rich media, grew as yeast at wild-type rates at both 25 and 37 degrees C and were as virulent as the wild type in a mouse model. However, detailed morphological and cytological studies of the wdchs1Delta mutants showed that yeast cells often failed to separate, tended to be enriched with chitin in septal regions and, sometimes, were enlarged with multiple nuclei, had broader mother cell-daughter bud regions and had other cell wall defects seen considerably less often than in the wild type or wdchs2 Delta strains. Disruption of WdCHS1 and WdCHS2 in the same background revealed that WdChs1p had functions synergistic to those of WdChs2p, because mutants devoid of both isozymes produced growth that was very abnormal at 25 degrees C and was not viable at 37 degrees C unless osmotically stabilized. These results suggested that WdChs1p was more responsible than WdChs2p for normal yeast cell reproductive growth because strains with defects in the latter exhibited no morphological abnormalities, whereas those with defects in WdChs1p were frequently impaired in one or more yeast developmental processes.

Animals↗

Evidence of focal small-fiber axonal degeneration in complex regional pain syndrome-I (reflex sympathetic dystrophy).

CRPS-I consists of post-traumatic limb pain and autonomic abnormalities that continue despite apparent healing of inciting injuries. The cause of symptoms is unknown and objective findings are few, making diagnosis and treatment controversial, and research difficult. We tested the hypotheses that CRPS-I is caused by persistent minimal distal nerve injury (MDNI), specifically distal degeneration of small-diameter axons. These subserve pain and autonomic function. We studied 18 adults with IASP-defined CRPS-I affecting their arms or legs. We studied three sites on subjects' CRPS-affected and matching contralateral limb; the CRPS-affected site, and nearby unaffected ipsilateral and matching contralateral control sites. We performed quantitative mechanical and thermal sensory testing (QST) followed by quantitation of epidermal neurite densities within PGP9.5-immunolabeled skin biopsies. Seven adults with chronic leg pain, edema, disuse, and prior surgeries from trauma or osteoarthritis provided symptom-matched controls. CRPS-I subjects had representative histories and symptoms. Medical procedures were unexpectedly frequently associated with CRPS onset. QST revealed mechanical allodynia (P<0.03) and heat-pain hyperalgesia (P<0.04) at the CRPS-affected site. Axonal densities were highly correlated between subjects' ipsilateral and contralateral control sites (r=0.97), but were diminished at the CRPS-affected sites of 17/18 subjects, on average by 29% (P<0.001). Overall, control subjects had no painful-site neurite reductions (P=1.00), suggesting that pain, disuse, or prior surgeries alone do not explain CRPS-associated neurite losses. These results support the hypothesis that CRPS-I is specifically associated with post-traumatic focal MDNI affecting nociceptive small-fibers. This type of nerve injury will remain undetected in most clinical settings.

Adolescent↗

The TIGR Maize Database.

Maize is a staple crop of the grass family and also an excellent model for plant genetics. Owing to the large size and repetitiveness of its genome, we previously investigated two approaches to accelerate gene discovery and genome analysis in maize: methylation filtration and high C(0)t selection. These techniques allow the construction of gene-enriched genomic libraries by minimizing repeat sequences due to either their methylation status or their copy number, yielding a 7-fold enrichment in genic sequences relative to a random genomic library. Approximately 900,000 gene-enriched reads from maize were generated and clustered into Assembled Zea mays (AZM) sequences. Here we report the current AZM release, which consists of approximately 298 Mb representing 243,807 sequence assemblies and singletons. In order to provide a repository of publicly available maize genomic sequences, we have created the TIGR Maize Database (http://maize.tigr.org). In this resource, we have assembled and annotated the AZMs and used available sequenced markers to anchor AZMs to maize chromosomes. We have constructed a maize repeat database and generated draft sequence assemblies of 287 maize bacterial artificial chromosome (BAC) clone sequences, which we annotated along with 172 additional publicly available BAC clones. All sequences, assemblies and annotations are available at the project website via web interfaces and FTP downloads.

Chromosome Mapping↗

Unique substitution of CHEK2 and TP53 mutations implicated in primary prostate tumors and cancer cell lines.

Genetic defects in CHEK2 and TP53 have been implicated in prostate cancer development. However, the interaction of these two genes in prostate cancer tumorigenesis has not been investigated. We previously described 11 CHEK2 mutations in a group of 84 primary prostate tumors. In this report, we screened the same group of tumors for TP53 mutations and revealed nine somatic and two germline mutations. One germline TP53 mutation (c.408A > T/p.Gln136His) and two somatic mutations (c.1022T > G/p.Phe341Cys and c.108-109ins22/p.His37fsX13) are novel to human cancer. More interestingly, CHEK2 and TP53 mutations were observed to be mutually substituted in these tumors. Analysis of five commonly used prostate cancer cell lines revealed that four cell lines harboring TP53 mutations carry no CHEK2 mutation while the only cell line (LNCaP) carrying wild-type TP53 harbors a CHEK2 mutation. The novel CHEK2 mutation (c.1160C>T/p.Thr387Asn) identified in LNCaP cells changes amino acid Thr387 to Asn which has been shown to impair CHEK2 autophosphorylation and activation. Our data suggest that the CHEK2 and TP53 mutations can substitute each other in at least 25% (21/84) of prostate cancers and that DNA damage-signaling pathway plays an important role in prostate cancer tumorigenesis.

Aged↗

Identification of norharman as the cytotoxic compound produced by the sponge (Hymeniacidon perleve)-associated marine bacterium Pseudoalteromonas piscicida and its apoptotic effect on cancer cells.

Marine invertebrates harbour a wealth of micro-organisms in their bodies. Most of these micro-organisms can catabolize antibiotic compounds as chemical-defence compounds. These compounds not only play an important protective role for their producer and for their hosts, but also have high potential in medicinal applications. In order to discover natural anticancer products, 29 marine bacterial strains have been isolated from the sponge Hymeniacidon perleve, samples of which were collected from the intertidal zone during low tide off Nanji island in Eastern China. By means of a cytotoxicity bioassay, one strain, NJ6-3-1, with significant cytotoxic activity, was selected for culture in a 30-litre fermentation tank. The major cytotoxic compound in the metabolites of NJ6-3-1, separated by means of a bioassay-guided fractionation process, has been identified as norharman (a beta-carboline alkaloid) by electron-impact MS and NMR analyses. Norharman showed cytotoxicity towards both the HeLa cervical-cancer cell line and the BGC-823 stomach-cancer cell line, with an IC(50) of 5 microg/ml. Several methods were used to study the mechanism by which norharman is cytotoxic to HeLa cells. By means of an Acridine Orange/ethidium bromide dual-staining assay, condensation of chromatin was observed. A TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling) assay showed degradation of DNA. Flow-cytometric analysis indicated that norharman could arrest cells at the G(2)/M phase of the cell cycle. These results demonstrate the cytotoxic mechanism of norharman involves the induction of apoptosis in HeLa cells.

Animals↗

Gene therapy for dentin regeneration with bone morphogenetic proteins.

Recent advances in stem cell biology and gene therapy technology have provided the great potential of adult stem cells for therapeutic use in regeneration of lost tissue due to diseases including cancer, trauma, and even caries. Dental pulp tissues harbor mesenchymal stem/progenitor cells and have potential to regenerate and/or repair dentin-pulp complex after injury such as caries. There are two main methods, in vivo and ex vivo gene therapy. In in vivo gene therapy the healing potential of pulp tissue is enhanced by genes inducing dentin directly applied on the exposed/amputated dental pulp. In ex vivo gene therapy, pulp stem/progenitor cells transfected with some therapeutically proven genes to induce differentiation into odontoblasts which are transplanted on the exposed/amputated pulp. In the inflamed pulp under deep caries or trauma, possibly due to the limited supply of pulp stem/progenitor cells, it might be useful to apply cell-based ex vivo gene therapy compared to in vivo gene therapy. Before clinical use of ex vivo gene therapy for dentin regeneration in endodontics, there is a need for establishment of isolation, identification and expansion of the pulp stem cells. A safe and efficient gene delivery system also needs to be optimized. In this review we provide an overview of our current knowledge in the biology and function of adult pulp stem cells. This is followed by a discussion of the challenges of translating basic cellular and molecular biology of differentiation of pulp stem cells to safe and efficient gene therapy for dentin regeneration.

Bone Morphogenetic Proteins↗

Identification of an essential glycoprotease in Staphylococcus aureus.

The emergence of multi-drug resistant bacterial pathogens is generating enormous public health concern, and highlights an urgent need for new, alternative agents for treating multi-drug-resistant pathogens. The gene products essential for bacterial growth in vitro and survival during infection constitute an initial set of protein targets for the development of antibacterial agents. In this study, we employed regulated gene expression approaches and demonstrated that a putative glycoprotease (Gcp) is required for staphylococcal growth in the culture. We found that Staphylococcus aureus becomes more sensitive to the Zn(2+) ion under the downregulation of Gcp expression in vitro. Bioinformatic analyses demonstrated that Gcp is conserved in many Gram-positive pathogens and exists in a variety of Gram-negative pathogens. Our results indicate that Gcp is a potential novel target for the development of antimicrobials against S. aureus infection.

Anti-Bacterial Agents↗