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111.
Jennifer Dieppedale Gael Gesbert Elodie Ramond Cerina Chhuon Iharilalao Dubail Marion Dupuis Ida Chiara Guerrera Alain Charbit 《Molecular & cellular proteomics : MCP》2013,12(8):2278-2292
Francisella tularensis is a highly infectious bacterium causing the zoonotic disease tularemia. In vivo, this facultative intracellular bacterium survives and replicates mainly in the cytoplasm of infected cells. We have recently identified a genetic locus, designated moxR that is important for stress resistance and intramacrophage survival of F. tularensis. In the present work, we used tandem affinity purification coupled to mass spectrometry to identify in vivo interacting partners of three proteins encoded by this locus: the MoxR-like ATPase (FTL_0200), and two proteins containing motifs predicted to be involved in protein–protein interactions, bearing von Willebrand A (FTL_0201) and tetratricopeptide (FTL_0205) motifs. The three proteins were designated here for simplification, MoxR, VWA1, and TPR1, respectively. MoxR interacted with 31 proteins, including various enzymes. VWA1 interacted with fewer proteins, but these included the E2 component of 2-oxoglutarate dehydrogenase and TPR1. The protein TPR1 interacted with one hundred proteins, including the E1 and E2 subunits of both oxoglutarate and pyruvate dehydrogenase enzyme complexes, and their common E3 subunit. Remarkably, chromosomal deletion of either moxR or tpr1 impaired pyruvate dehydrogenase and oxoglutarate dehydrogenase activities, supporting the hypothesis of a functional role for the interaction of MoxR and TPR1 with these complexes. Altogether, this work highlights possible links between stress resistance and metabolism in F. tularensis virulence.Francisella tularensis is responsible for the disease tularamia in a large number of animal species. This highly infectious bacterial pathogen can be transmitted to humans in numerous ways (1, 2, 3), including direct contact with sick animals, inhalation, ingestion of contaminated water or food, or by bites from ticks, mosquitoes, or flies. Four different subspecies (subsp.) of F. tularensis that differ in virulence and geographic distribution exist, designated subsp. tularensis (type A), subsp. holarctica (type B), subsp. Novicida, and subsp. mediasiatica, respectively. F. tularensis subsp. tularensis is the most virulent subspecies causing a severe disease in humans, whereas F. tularensis subsp. holarctica causes a similar disease but of less severity (4). Because of its high infectivity and lethality, F. tularensis is considered a potential bioterrorism agent (5).F. tularensis is able to survive and to replicate in the cytoplasm of a variety of infected cells, including macrophages. To resist this stressful environment, the bacterium must have developed stress resistance mechanisms, most of which are not yet well characterized. We recently reported the identification of a novel genetic locus that is important for stress resistance and intracellular survival of F. tularensis (6). This locus was designated moxR because the first gene FTL_0200, encodes a protein belonging to the AAA+ ATPase of the MoxR family ((7) and references therein). The data obtained in that first study had led us to suggest that the F. tularensis MoxR-like protein might constitute, in combination with other proteins of the locus, a chaperone complex contributing to F. tularensis pathogenesis.To further validate this hypothesis and expand our initial observations, we here decided to perform tandem affinity purification (TAP),1 using a dual affinity tag approach coupled to mass spectroscopy analyses (8), to identify proteins interacting in vivo with three proteins encoded by the proximal portion of the moxR locus. For this, we chose as baits: the MoxR-like protein (FTL_0200) and two proteins bearing distinct motifs possibly involved in protein–protein interactions, FTL_0201 (Von Willebrand Factor Type A domain, or VWA) and FTL_0205 (tetratrichopeptide repeat or TPR). The three proteins were designated here for simplification, MoxR, VWA1, and TPR1; and the corresponding genes moxR, vwa1, and tpr1, respectively.VWA domains are present in all three kingdoms of life. They consist of a β-sheet sandwiched by multiple α helices. Frequently, VWA domain-containing proteins function in multiprotein complexes (9). TPR typically contain 34 amino acids. Many three-dimensional structures of TPR domains have been solved, revealing amphipathic helical structures (10). TPR-containing proteins are also found in all kingdoms of life. They can be involved in a variety of functions, and generally mediate protein–protein interactions. In the past few years, several TPR-related proteins have been shown to be involved in virulence mechanisms in pathogenic bacteria ((11) and references therein).Our proteomic approach allowed us to identify a series of protein interactants for each of the three moxR-encoded proteins. Remarkably, the protein TPR1 interacted with all the subunits of the pyruvate dehydrogenase (PDH) and 2-oxoglutarate dehydrogenase (OGDH) complexes. Furthermore, inactivation of tpr1 also severely impaired the activities of these two enzymes. Inactivation of tpr1 affected bacterial resistance to several stresses (and in particular oxidative stress), intramacrophagic bacterial multiplication and bacterial virulence in the mouse model. Functional implications and possible relationship between bacterial metabolism, stress defense, and bacterial virulence are discussed. 相似文献
112.
Alina Kuzniacka Jolanta Wierzba Magdalena Ratajska Beata S. Lipska Magdalena Koczkowska Monika Malinowska Janusz Limon 《Journal of applied genetics》2013,54(1):27-33
Cornelia de Lange syndrome (CdLS) is a rare multi-system genetic disorder characterised by growth and developmental delay, distinctive facial dysmorphism, limb malformations and multiple organ defects. The disease is caused by mutations in genes responsible for the formation and regulation of cohesin complex. About half of the cases result from mutations in the NIPBL gene coding delangin, a protein regulating the initialisation of cohesion. To date, approximately 250 point mutations have been identified in more than 300 CdLS patients worldwide. In the present study, conducted on a group of 64 unrelated Polish CdLS patients, 25 various NIPBL sequence variants, including 22 novel point mutations, were detected. Additionally, large genomic deletions on chromosome 5p13 encompassing the NIPBL gene locus were detected in two patients with the most severe CdLS phenotype. Taken together, 42 % of patients were found to have a deleterious alteration affecting the NIPBL gene, by and large private ones (89 %). The review of the types of mutations found so far in Polish patients, their frequency and correlation with the severity of the observed phenotype shows that Polish CdLS cases do not significantly differ from other populations. 相似文献
113.
Claudio Ghiglione Maria Chiara Alvaro Huw J. Griffiths Katrin Linse Stefano Schiaparelli 《ZooKeys》2013,(341):37-48
Information regarding the molluscs in this dataset is based on the Rauschert dredge samples collected during the Latitudinal Gradient Program (LGP) on board the R/V “Italica” in the Ross Sea (Antarctica) in the austral summer 2004. A total of 18 epibenthic dredge deployments/samplings have been performed at four different locations at depths ranging from 84 to 515m by using a Rauschert dredge with a mesh size of 500μm. In total 8,359 specimens have been collected belonging to a total of 161 species. Considering this dataset in terms of occurrences, it corresponds to 505 discrete distributional records (incidence data). Of these, in order of abundance, 5,965 specimens were Gastropoda (accounting for 113 species), 1,323 were Bivalvia (accounting for 36 species), 949 were Aplacophora (accounting for 7 species), 74 specimens were Scaphopoda (3 species), 38 were Monoplacophora (1 species) and, finally, 10 specimens were Polyplacophora (1 species). This data set represents the first large-scale survey of benthic micro-molluscs for the area and provides important information about the distribution of several species, which have been seldom or never recorded before in the Ross Sea. All vouchers are permanently stored at the Italian National Antarctic Museum (MNA), Section of Genoa, enabling future comparison and crosschecking. This material is also currently under study, from a molecular point of view, by the barcoding project “BAMBi” (PNRA 2010/A1.10). 相似文献
114.
115.
Ivana Sreckovic Ruth Birner-Gruenberger Britta Obrist Tatjana Stojakovic Hubert Scharnagl Michael Holzer Monika Scholler Sonia Philipose Gunther Marsche Uwe Lang Gernot Desoye Christian Wadsack 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2013,1831(4):737-746
In human high-density lipoprotein (HDL) represents the major cholesterol carrying lipoprotein class in cord blood, while cholesterol is mainly carried by low-density lipoprotein in maternal serum. Additionally, to carrying cholesterol, HDL also associates with a range of proteins as cargo. We tested the hypothesis that fetal HDL carries proteins qualitatively and quantitatively different from maternal HDL. These differences then contribute to distinct HDL functionality in both circulations. Shotgun proteomics and biochemical analyses were used to assess composition/function of fetal and maternal HDL isolated from uncomplicated human pregnancies at term of gestation. The pattern of analyzed proteins that were statistically elevated in fetal HDL (apoE, proteins involved in coagulation, transport processes) suggests a particle characteristic for the light HDL2 sub-fraction. In contrast, proteins that were enriched in maternal HDL (apoL, apoF, PON1, apoD, apoCs) have been described almost exclusively in the dense HDL3 fraction and relevant to its anti-oxidative function and role in innate immunity. Strikingly, PON1 mass and activity were 5-fold lower (p < 0.01) in the fetus, which was accompanied by attenuation of anti-oxidant capacity of fetal HDL. Despite almost equal quantity of CETP in maternal and fetal HDL, its enzymatic activity was 55% lower (p < 0.001) in the fetal circulation, whereas LCAT activity was not altered. These findings indicate that maternally derived HDL differs from fetal HDL with respect to its proteome, size and function. Absence of apoA-1, apoL and PON1 on fetal HDL is associated with decreased anti-oxidative properties together with deficiency in innate immunity collectively indicating distinct HDLs in fetuses. 相似文献
116.
Elena De Falco Gaia Scafetta Chiara Napoletano Rosa Puca Enzo Maria Vingolo Giuseppe Ragona Olga Iorio Giacomo Frati 《Cell and tissue banking》2013,14(2):277-287
Good manufacturing practices guidelines require safer and standardized cell substrates especially for those cell therapy products to treat ocular diseases where fibroblasts are used as feeder layers. However, if these are unavailable for stem cells culturing, murine fibroblasts are regularly used, raising critical issues as accidentally transplanting xenogenous graft and adversely affecting stem cell clinical trials. Moreover, human fibroblasts play a significant role in testing novel ophthalmologic drugs. Accordingly, we developed a standardized laboratory and surgical approach to isolate normal and undamaged Tenon’s fibroblasts to implement the setting up of banks for both stem cells-based ocular cell therapy and in vitro drug testing. A 2–3 cm2 undamaged Tenon’s biopsy was surgically obtained from 28 patients without mutually correlated ocular diseases. Nineteen dermal biopsies were used as control. Fibroblasts were isolated with or without collagenase, cultured in autologous, fetal bovine or AB serum, tested for viability by trypan blue, vimentin expression and standardized until passage 6. Successful Tenon’s fibroblasts isolation was age dependent (P = 0.001) but not sex, pathology or surgery related. A significant rate of successful cultures were obtained when biopsies were not digested by collagenase (P = 0.013). Moreover, cultures in autologous or fetal bovine serum had comparable proliferative properties (P = 0.77; P = 0.82). Through our surgical and laboratory standardized procedure, we elucidated for the first time key points of this human primary culture system, the role of the autologous serum, comparing Tenon’s and dermal fibroblasts. Our protocol may be clinically useful to reduce the risk above mentioned and may be potentially more effective for ophthalmological clinical purposes. 相似文献
117.
Francesca Borgo Aristodemo Carpen Chiara Ferrario Stefania Iametti Maria Grazia Fortina 《Journal of industrial microbiology & biotechnology》2013,40(5):489-494
Through the analysis of the recently available genome shotgun sequence of Enterococcus italicus DSM 15952T type strain (Accession PRJNA61487, ID 61487), we found the presence of a gene encoding a bifunctional enzyme, termed γ-GCS-GS or GshF, involved in glutathione production and not influenced by feedback inhibition. The gshF gene exhibited high nucleotide and amino acid sequence similarity to other reported sequences from the Enterococcus genus and was constitutively expressed both in osmotic shock or in common cultural conditions. Several experimental studies concerning the culture medium, physiological stress, cell extract obtainment, and scaling-up showed that in selected conditions E. italicus was able to accumulate up to 250 μM of intracellular glutathione, which represented the main thiol group present into the cells. This is the first report regarding the production of glutathione by E. italicus, a species that could be used as a safe adjunct culture for glutathione-enriched dairy foods. 相似文献
118.
Monika Dybicz Anna Gierczak Julia Dąbrowska Łukasz Rdzanek Bogdan Michałowicz 《Parasitology international》2013,62(4):364-367
The identity of the causative agent of cystic echinococcosis (CE) in humans from central Poland receiving treatment between 2000 and 2010 was determined. A total of 47 samples obtained after hepatectomy were examined and protoscoleces were identified in wet preparations in 27 cases. Using DNA extracted from the samples, two mitochondrial regions (nad1 and cox1 genes) were amplified and the nad1 fragment was sequenced. This PCR analysis confirmed the presence of Echinococcus species in 30 cases and nad1 sequence alignments showed identity with the G7 (pig) strain, Echinococcus canadensis. These data demonstrate that the pig strain of this parasite is the most frequent causative agent of human cystic echinococcosis in central Poland. 相似文献
119.
Nematodes are very common in the deep sea and are an important component of deep-sea hydrothermal vent communities. In early 2006, the eruption of the underwater volcano at 9°50’N East Pacific Rise wiped out almost the entire faunal communities of the area. This provided us with the opportunity to study nematode primary succession at vents as well as on adjacent seafloor basalt. Nematode abundance and richness were extremely low at all studied sites in late 2006 and 2007, and increased only slightly in 2009. Interestingly, the most abundant species during early succession were also prominent in this area prior to the eruption. Our results show that nematodes are extremely influenced by volcanic eruptions and need a long period of time to colonize the lava-flooded area in greater numbers and richness. We hypothesize that low food availability on the young bare basalt and harsh environmental conditions at early succession vent sites might hinder a more successful nematode establishment. In addition to the newly established active vent sites we also studied an inactive vent site that was not directly hit by the eruption but whose vent fluid had ceased after the eruption. At this inactive and older vent, diversity was also relatively low but was higher than at the younger, newly established sites. In addition to the ecological analyses, we here describe the two most abundant species found at inactive vents, namely Neochromadora aff. poecilosoma De Mann 1893 and Linhomoeus caudipapillosus sp. n. 相似文献
120.
Jan Plue Pieter De Frenne Kamal Acharya Jrg Brunet Olivier Chabrerie Guillaume Decocq Martin Diekmann Bente J. Graae Thilo Heinken Martin Hermy Annette Kolb Isgard Lemke Jaan Liira Tobias Naaf Anna Shevtsova Kris Verheyen Monika Wulf Sara A. O. Cousins 《Global Ecology and Biogeography》2013,22(10):1106-1117