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101.
102.
Darío FernándezSusana Larrucea Adam NowakowskiMiguel Pericacho Roberto Parrilla Matilde S. Ayuso 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2011,1813(8):1504-1510
Podocalyxin (PODXL) is a type I membrane mucoprotein abundantly presented in the epithelial cells (podocytes) of kidney glomeruli where it plays an important role in maintaining the plasma filtration. PODXL is also expressed in other types of cells but its function is ignored. A recombinant soluble fragment of the PODXL ectodomain modifies the signaling of the membrane bound PODXL. Based on this antecedent, we aimed at investigating whether PODXL could be cleaved and released into the extracellular space as a soluble peptide. In this study, we used a fusion protein of human PODXL and green fluorescent protein expressed in CHO cells (CHO-PODXL-GFP) and a human tumor cell (Tera-1) inherently expressing PODXL. PODXL was detected by wide-field microscopy in the Golgi, the plasma membrane and in a vesicular form preferentially located at the leading edges of the cell and also progressing along the filopodium. We detected PODXL in the insoluble and soluble fractions of the extracellular medium of CHO-PODXL-GFP cells. Stimulation of protein kinase C (PKC) by Phorbol-12-myristate-13-acetate (PMA) enhanced the release of PODXL to the extracellular space whereas this effect was prevented either by inhibitors of PKC or specific inhibitors of matrix metalloproteinases. It is concluded that intact PODXL is released to the extracellular space as a cargo of microvesicles and also as a soluble cleaved fragment of ectodomain. 相似文献
103.
Many techniques have been applied to understand viral cell-to-cell movement in host plants, but little progress has been made in understanding viral vascular transport mechanisms. We propose the use of chlorophyll fluorescence imaging techniques, not only to diagnose the viral infection, but also to follow the movement of the virus through the vascular system and its subsequent spread into the leaves. In Nicotiana benthamiana plants, imaging of chlorophyll fluorescence parameters such as ФPSII and NPQ proved useful to follow infections with Pepper mild mottle virus. The results demonstrate a correlation between changes in the chlorophyll fluorescence parameters and the viral distribution analyzed by tissue printing. 相似文献
104.
Marracci S Michelotti V Guex GD Hotz H Uzzell T Ragghianti M 《Journal of molecular evolution》2011,72(4):368-382
RrS1-like sequences of water frogs (genus Pelophylax) display varied genomic organization, whereas the centromeric hybridization pattern reveals species-specific differences.
Using fluorescent in situ hybridization, Pelophylax cf. bedriagae, Pelophylax
kurtmuelleri, and Pelophylax
ridibundus showed a hybridization signal at centromeres of chromosomes 1–5, but in P. kurtmuelleri the medium-small chromosome labeled was 10 rather than 8. Pelophylax
cretensis had almost 16 of 26 centromeres labeled, as did Pelophylax
lessonae from Poland when its chromosomes are hybridized with a homologous probe. When StuI-digested genomic DNA was hybridized with RrS1 probe, hybridization ladders for P. ridibundus from Poland have evenly spaced steps (about 100 bp) of uniform intensity from about 200 bp upward. Steps in hybridization
ladders from circum-Aegean taxa vary in intensity: larger, odd-numbered steps are often fainter. A strong double band (800/900 bp)
in Anatolian P. cf. bedriagae, emphasized by a weak 700 bp band, distinguishes them from P. kurtmuelleri from the Peloponnisos, in which the 900 bp band is almost absent. The ladder in P. cretensis lacks odd-numbered steps. A and B repeats, observed originally within the RrS1 satellite of P. ridibundus, occur also in the circum-Aegean frogs and in P. lessonae, Pelophylax
epeiroticus, Pelophylax
saharicus, and Pelophylax
shqipericus. It is plausible that AB dimers or ABB trimers rather than A or B monomers correspond to functional/evolutionary units. The
presence of regions similar to yeast CDEs and mammalian CENP-B boxes suggests a role for RrS1 sequences in centromere organization. 相似文献
105.
Tuñon MJ San Miguel B Crespo I Riezu-Boj JI Larrea E Alvarez M González I Bustos M González-Gallego J Prieto J 《Journal of virology》2011,85(24):13124-13132
Rabbit hemorrhagic disease virus (RHDV) causes lethal fulminant hepatitis closely resembling acute liver failure (ALF) in humans. In this study, we investigated whether cardiotrophin-1 (CT-1), a cytokine with hepatoprotective properties, could attenuate liver damage and prolong survival in virus-induced ALF. Twenty-four rabbits were infected with 2 × 10(4) hemagglutination units of RHDV. Twelve received five doses of CT-1 (100 μg/kg) starting at 12 h postinfection (hpi) (the first three doses every 6 h and then two additional doses at 48 and 72 hpi), while the rest received saline. The animals were analyzed for survival, serum biochemistry, and viral load. Another cohort (n = 22) was infected and treated similarly, but animals were sacrificed at 30 and 36 hpi to analyze liver histology, viral load, and the expression of factors implicated in liver damage and repair. All infected rabbits that received saline died by 60 hpi, while 67% of the CT-1-treated animals survived until the end of the study. Treated animals showed improved liver function and histology, while the viral loads were similar. In the livers of CT-1-treated rabbits we observed reduction of oxidative stress, diminished PARP1/2 and JNK activation, and decreased inflammatory reaction, as reflected by reduced expression of tumor necrosis factor alpha, interleukin-1β, Toll-like receptor 4, VCAM-1, and MMP-9. In addition, CT-1-treated rabbits exhibited marked upregulation of TIMP-1 and increased expression of cytoprotective and proregenerative growth factors, including platelet-derived growth factor B, epidermal growth factor, platelet-derived growth factor receptor β, and c-Met. In conclusion, in a lethal form of acute viral hepatitis, CT-1 increases animal survival by attenuating inflammation and activating cytoprotective mechanisms, thus representing a promising therapy for ALF of viral origin. 相似文献
106.
The E3 ubiquitin ligase RNF168 is a DNA damage response (DDR) factor that promotes monoubiquitination of H2A/H2AX at K13/15, facilitates recruitment of other DDR factors (e.g. 53BP1) to DNA damage, and inhibits homologous recombination (HR) in cells deficient in the tumor suppressor BRCA1. We have examined the domains of RNF168 important for these DDR events, including chromosomal HR that is induced by several nucleases (I-SceI, CAS9-WT and CAS9-D10A), since the inducing nuclease affects the relative frequency of distinct repair outcomes. We found that an N-terminal fragment of RNF168 (1-220/N221*) efficiently inhibits HR induced by each of these nucleases in BRCA1 depleted cells, and promotes recruitment of 53BP1 to DNA damage and H2AX monoubiquitination at K13/15. Each of these DDR events requires a charged residue in RNF168 (R57). Notably, RNF168-N221* fails to self-accumulate into ionizing radiation induced foci (IRIF). Furthermore, expression of RNF168 WT and N221* can significantly bypass the role of another E3 ubiquitin ligase, RNF8, for inhibition of HR in BRCA1 depleted cells, and for promotion of 53BP1 IRIF. We suggest that the ability for RNF168 to promote H2A/H2AX monoubiquitination and 53BP1 IRIF, but not RNF168 self-accumulation into IRIF, is important for inhibition of HR in BRCA1 deficient cells. 相似文献
107.
Linda Avesani Matilde Merlin Elisa Gecchele Stefano Capaldi Annalisa Brozzetti Alberto Falorni Mario Pezzotti 《Transgenic research》2014,23(2):281-291
The 65-kDa isoform of human glutamic acid decarboxylase (hGAD65) is a major diabetes autoantigen that can be used for the diagnosis and (more recently) the treatment of autoimmune diabetes. We previously reported that a catalytically-inactive version (hGAD65mut) accumulated to tenfold higher levels than its active counterpart in transgenic tobacco plants, providing a safe and less expensive source of the protein compared to mammalian production platforms. Here we show that hGAD65mut is also produced at higher levels than hGAD65 by transient expression in Nicotiana benthamiana (using either the pK7WG2 or MagnICON vectors), in insect cells using baculovirus vectors, and in bacterial cells using an inducible-expression system, although the latter system is unsuitable because hGAD65mut accumulates within inclusion bodies. The most productive of these platforms was the MagnICON system, which achieved yields of 78.8 μg/g fresh leaf weight (FLW) but this was substantially less than the best-performing elite transgenic tobacco plants, which reached 114.3 μg/g FLW after six generations of self-crossing. The transgenic system was found to be the most productive and cost-effective although the breeding process took 3 years to complete. The MagnICON system was less productive overall, but generated large amounts of protein in a few days. Both plant-based systems were therefore advantageous over the baculovirus-based production platform in our hands. 相似文献
108.
109.
Lázaro Molina Zulema Udaondo Estrella Duque Matilde Fernández Carlos Molina-Santiago Amalia Roca Mario Porcel Jesús de la Torre Ana Segura Patrick Plesiat Katy Jeannot Juan-Luis Ramos 《PloS one》2014,9(1)
Environmental microbes harbor an enormous pool of antibiotic and biocide resistance genes that can impact the resistance profiles of animal and human pathogens via horizontal gene transfer. Pseudomonas putida strains are ubiquitous in soil and water but have been seldom isolated from humans. We have established a collection of P. putida strains isolated from in-patients in different hospitals in France. One of the isolated strains (HB3267) kills insects and is resistant to the majority of the antibiotics used in laboratories and hospitals, including aminoglycosides, ß-lactams, cationic peptides, chromoprotein enediyne antibiotics, dihydrofolate reductase inhibitors, fluoroquinolones and quinolones, glycopeptide antibiotics, macrolides, polyketides and sulfonamides. Similar to other P. putida clinical isolates the strain was sensitive to amikacin. To shed light on the broad pattern of antibiotic resistance, which is rarely found in clinical isolates of this species, the genome of this strain was sequenced and analysed. The study revealed that the determinants of multiple resistance are both chromosomally-borne as well as located on the pPC9 plasmid. Further analysis indicated that pPC9 has recruited antibiotic and biocide resistance genes from environmental microorganisms as well as from opportunistic and true human pathogens. The pPC9 plasmid is not self-transmissible, but can be mobilized by other bacterial plasmids making it capable of spreading antibiotic resistant determinants to new hosts. 相似文献
110.
Carles Gaig Dolores Vilas Jon Infante María Sierra Inés García-Gorostiaga Mariateresa Buongiorno Mario Ezquerra Maria José Martí Francesc Valldeoriola Miquel Aguilar Matilde Calopa Jorge Hernandez-Vara Eduardo Tolosa 《PloS one》2014,9(10)