Nguyen Thuy Duong

Nesprins: Tissue-Specific Expression of Epsilon and Other Short Isoforms

Nesprin-1-giant and nesprin-2-giant regulate nuclear positioning by the interaction of their C-terminal KASH domains with nuclear membrane SUN proteins and their N-terminal calponin-homology domains with cytoskeletal actin. A number of short isoforms lacking the actin-binding domains are produced by internal promotion. We have evaluated the significance of these shorter isoforms using quantitative RT-PCR and western blotting with site-specific monoclonal antibodies. Within a complete map of nesprin isoforms, we describe two novel nesprin-2 epsilon isoforms for the first time. Epsilon isoforms are similar in size and structure to nesprin-1-alpha. Expression of nesprin isoforms was highly tissue-dependent. Nesprin-2-epsilon-1 was found in early embryonic cells, while nesprin-2-epsilon-2 was present in heart and other adult tissues, but not skeletal muscle. Some cell lines lack shorter isoforms and express only one of the two nesprin genes, suggesting that either of the giant nesprins is sufficient for basic cell functions. For the first time, localisation of endogenous nesprin away from the nuclear membrane was shown in cells where removal of the KASH domain by alternative splicing occurs. By distinguishing between degradation products and true isoforms on western blots, it was found that previously-described beta and gamma isoforms are expressed either at only low levels or with a limited tissue distribution. Two of the shortest alpha isoforms, nesprin-1-alpha-2 and nesprin-2-alpha-1, were found almost exclusively in cardiac and skeletal muscle and a highly conserved and alternatively-spliced exon, available in both nesprin genes, was always included in these tissues. These “muscle-specific” isoforms are thought to form a complex with emerin and lamin A/C at the inner nuclear membrane and mutations in all three proteins cause Emery-Dreifuss muscular dystrophy and/or inherited dilated cardiomyopathy, disorders in which only skeletal muscle and/or heart are affected.

Specific localization of nesprin-1-α2, the short isoform of nesprin-1 with a KASH domain, in developing, fetal and regenerating muscle, using a new monoclonal antibody

BackgroundNesprin-1-giant (1008kD) is a protein of the outer nuclear membrane that links nuclei to the actin cytoskeleton via amino-terminal calponin homology domains. The short nesprin-1 isoform, nesprin-1-α2, is present only in skeletal and cardiac muscle and several pathogenic mutations occur within it, but the functions of this short isoform without calponin homology domains are unclear. The aim of this study was to determine mRNA levels and protein localization of nesprin-1-α2 at different stages of muscle development in order to shed light on its functions.ResultsmRNA levels of all known nesprin-1 isoforms with a KASH domain were determined by quantitative PCR. The mRNA for the 111 kD muscle-specific short isoform, nesprin-1-α2, was not detected in pre-differentiation human myoblasts but was present at significant levels in multinucleate myotubes. We developed a monoclonal antibody against the unique amino-terminal sequence of nesprin-1-α2, enabling specific immunolocalization for the first time. Nesprin-1-α2 protein was undetectable in pre-differentiation myoblasts but appeared at the nuclear rim in post-mitotic, multinucleate myotubes and reached its highest levels in fetal muscle. In muscle from a Duchenne muscular dystrophy biopsy, nesprin-1-α2 protein was detected mainly in regenerating fibres expressing neonatal myosin. Nesprin-1-giant was present at all developmental stages, but was also highest in fetal and regenerating fibres. In fetal muscle, both isoforms were present in the cytoplasm, as well as at the nuclear rim. A pathogenic early stop codon (E7854X) in nesprin-1 caused reduced mRNA levels and loss of protein levels of both nesprin-1-giant and (unexpectedly) nesprin-1-α2, but did not affect myogenesis in vitro.ConclusionsNesprin-1-α2 mRNA and protein expression is switched on during myogenesis, alongside other known markers of muscle differentiation. The results show that nesprin-1-α2 is dynamically controlled and may be involved in some process occurring during early myofibre formation, such as re-positioning of nuclei.

Regulation of dendritic cell function by insulin/IGF-1/PI3K/Akt signaling through klotho expression

Abstract Insulin or insulin-like growth factor 1 (IGF-1) promotes the activation of phosphoinositide 3 kinase (PI3K)/Akt signaling in immune cells including dendritic cells (DCs), the most potent professional antigen-presenting cells for naive T cells. Klotho, an anti-aging protein, participates in the regulation of the PI3K/Akt signaling, thus the Ca2+-dependent migration is reduced in klotho-deficient DCs. The present study explored the effects of insulin/IGF-1 on DC function through klotho expression. To this end, the mouse bone marrow cells were isolated and cultured with GM-CSF to attain bone marrow-derived DCs (BMDCs). Cells were treated with insulin or IGF-1 and followed by stimulating with lipopolysaccharides (LPS). Tumor necrosis factor (TNF)-α formation was examined by enzyme-linked immunosorbent assay (ELISA). Phagocytosis was analyzed by FITC-dextran uptake assay. The expression of klotho was determined by quantitative PCR, immunoprecipitation and western blotting. As a result, treatment of the cells with insulin/IGF-1 resulted in reducing the klotho expression as well as LPS-stimulated TNF-α release and increasing the FITC-dextran uptake but unaltering reactive oxygen species (ROS) production in BMDCs. The effects were abolished by using pharmacological inhibition of PI3K/Akt with LY294002 and paralleled by transfecting DCs with klotho siRNA. In conclusion, the regulation of klotho sensitive DC function by IGF-1 or insulin is mediated through PI3K/Akt signaling pathway in BMDCs.

SELECTION OF INDIGENOUS STRAINS (Paecilomyces lilacinus) PARASITIZE Meloidogyne spp. ISOLATED FROM BA RIA – VUNG TAU PROVINCE

Twenty two Paecilomyces lilacinus strains were isolated from forest soils and black pepper rhizospheres in Ba Ria –Vung Tau Province. The ability to degrade chitin of PB 3.3, PB 2.9, QT2, and QT5 strains was high. The ability to degrade casein of PB 1.3, PB 2.10, KL5, and KL6 strains was efficient. And then, these strains were parasitized females and egg masses of Meloidogyne spp. in vitro . In female parasitism test, the rates of parasitizing female nematodes reached more than 50 % after treating for 2 days. Four strains of PB 2.10, PB 1.3, KL6 and QT5 belonged to the first group achieved the highest parasitic (> 90 %) effects on female after 3 days of incubation. In egg masses parasitism test, three strains of PB 1.3, PB 2.10 and QT5 exhibited 83.33 %, 75 % and 75 % parasitism on egg masses after 11 days of incubation. The rates of parasitizing female were higher than egg masses. Three selected strains from the experiments were PB 1.3, PB 2.10 and QT5.

CORROSION INHIBITION OF CARBON STEEL BY LDH/GO HYBRID INTERCALATED WITH 2-BENZOTHIAZOLYTHIO-SUCCINIC ACID

Layered double hydroxide/graphene oxide hybrid (LDH/GO) intercalated with corrosion inhibitor 2-benzothiazolylthio-succinic acid (BTSA) was prepared using co-precipitation method. The synthesized LDH/GO-BTSA was characterized by FTIR, XRD and SEM. The inhibitive action of LDH/GO-BTSA on carbon steel was evaluated and compared with LDH-BTSA by electrochemical measurement. It was shown that the GO and BTSA were intercalated in LDH structure. The obtained results showed that LDH/GO-BTSA is anodic corrosion inhibitors, and the inhibition efficiency was 94 % at concentration of 1 g/l.

Complete human mtDNA genome sequences from Vietnam and the phylogeography of Mainland Southeast Asia

Vietnam is an important crossroads within Mainland Southeast Asia (MSEA) and a gateway to Island Southeast Asia, and as such exhibits high levels of ethnolinguistic diversity. However, comparatively few studies have been undertaken of the genetic diversity of Vietnamese populations. In order to gain comprehensive insights into MSEA mtDNA phylogeography, we sequenced 609 complete mtDNA genomes from individuals belonging to five language families (Austroasiatic, Tai-Kadai, Hmong-Mien, Sino-Tibetan and Austronesian) and analyzed them in comparison with sequences from other MSEA countries and Taiwan. Within Vietnam, we identified 399 haplotypes belonging to 135 haplogroups; among the five language families, the sequences from Austronesian groups differ the most from the other groups. Phylogenetic analysis revealed 111 novel Vietnamese mtDNA lineages. Bayesian estimates of coalescence times and associated 95% HPD for these show a peak of mtDNA diversification around 2.5–3 kya, which coincides with the Dong Son culture, and thus may be associated with the agriculturally-driven expansion of this culture. Networks of major MSEA haplogroups emphasize the overall distinctiveness of sequences from Taiwan, in keeping with previous studies that suggested at most a minor impact of the Austronesian expansion from Taiwan on MSEA. We also see evidence for population expansions across MSEA geographic regions and language families.

Whole genome sequencing and mutation rate analysis of trios with paternal dioxin exposure

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin (TCDD) or dioxin, is commonly considered the most toxic man‐made substance. Dioxin exposure impacts human health and diseases, birth defects and teratogenesis were frequently observed in children of persons who have been exposed to dioxin. However, the impact of dioxin on human mutation rate in trios has not yet been elucidated at the whole genome level. To identify and characterize the genetic alterations in the individuals exposed to dioxin, we performed whole genome sequencing (WGS) of nine Vietnamese trios whose fathers were exposed to dioxin. In total, 846 de novo point mutations, 26 de novo insertions and deletions, 4 de novo structural variations, and 1 de novo copy number variation were identified. The number of point mutations and dioxin concentrations were positively correlated (P‐value < 0.05). Considering the substitution pattern, the number of A > T/T > A mutation and the dioxin concentration was positively correlated (P‐value < 0.05). Our analysis also identified one possible disease‐related mutation in LAMA5 in one trio. These findings suggested that dioxin exposure might affect father genomes of trios leading to de novo mutations in their children. Further analysis with larger sample sizes would be required to better clarify mutation rates and substitution patterns in trios caused by dioxin.

Genetic variation of mitochondrial sequence-hv2 in Vietnamese populations

HV1 and HV2, which located in control D-loop region of mitochondrial, are the hyper-variable parts of mitochondrial DNA (mtDNA). Therefore, these two sequences were often used for evolutional and forensic science. In order to determine the genetic variation of populations belong to Nam-A and Nam-Dao language systems, we sequenced and analyzed the mtDNA HV2 region of individuals from four ethic groups: Kinh, Muong, Ede and Jarai which represent for those language systems. Study subjects are peripheral blood of 169 individuals belonging to four populations: Kinh, Muong, Ede and Jarai. HV2 regions were sequenced by Sanger sequencing. As a result, the analyzed mitochondrial control region sequences could be assigned to 79 different haplogroups, with the dominant proportion of three haplogroups: R, B and F. Donors representing Kinh or Muong population revealed higher haplogroup composition variation in comparison with those affiliated with Jarai or Ede. In addition, it was indicated that the 4 populations shared the genetic analogy with other populations inhabited in Southeast Asia and South Asia.