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881.
To develop a new strategy to target recombinant proteins to the vacuolar storage system in transgenic plants, the ability
of the transmembrane and cytosolic domains of Arabidopsis receptor homology-transmembrane-RING H2-1 (AtRMR1) was evaluated.
A secreted version of RFP (secRFP) and a fusion of it to the transmembrane and cytosolic domains of AtRMR1 (RFP-TMCT) were
produced and studied both in transient and stable expression assays. Transient expression in leaves of Nicotiana tabacum showed that secRFP is secreted to the apoplast while its fusion to TMCT of AtRMR1 is sufficient to prevent secretion of the
reporter. In tobacco leaves, RFP-TMCT reporter showed an endoplasmic reticulum pattern in early expression stages while in
late expression stages, it was found in the vacuolar lumen. For the first time, the role of TM and CT domains of AtRMR1 in
stable expression in Arabidopsis thaliana is presented; the fusion of TMCT to secRFP is sufficient to sort RFP to the lumen of the central vacuoles in leaves and roots and to the
lumen of PSV in cotyledons of mature embryos. In addition, biochemical studies performed in extract from transgenic plants
showed that RFP-TMCT is an integral membrane protein. Full-length RFP-TMCT was also found in the vacuolar lumen, suggesting
internalization into destination vacuole. Not colocalization of RFP-TMCT with tonoplast and plasma membrane markers were observed.
This membrane vacuolar determinant sorting signal could be used for future application in molecular pharming as an alternative
means to sort proteins of interest to vacuoles. 相似文献
882.
Sporotrichosis is a subcutaneous mycosis caused by Sporothrix schenckii. Zoonotic transmission to man can occur after scratches or bites of animals, mainly cats. In this study, the gamma radiation
effects on yeast of S. schenckii were analyzed with a view of developing a radioattenuated vaccine for veterinary use. The cultures were irradiated at doses
ranging from 1.0 to 9.0 kGy. The reproductive capacity was measured by the ability of cells to form colonies. No colonies
could be recovered above 8.0 kGy, using inocula up to 107 cells. Nevertheless, yeast cells irradiated with 7.0 kGy already were unable to produce infection in immunosuppressed mice.
Evaluation by the FungaLight™ Kit (Invitrogen) indicated that yeast cells remained viable up to 9.0 kGy. At 7.0 kGy, protein
synthesis, estimated by the incorporation of [L-35S] methionine, continues at levels slightly lower than the controls, but a significant decrease was observed at 9.0 kGy. The
DNA of 7.0 kGy irradiated cells, analyzed by electrophoresis in agarose gel, was degraded. Cytoplasmic vacuolation was the
main change verified in these cells by transmission electron microscopy. The dose of 7.0 kGy was considered satisfactory for
yeast attenuation since irradiated cells were unable to produce infection but retained viability, metabolic activity, and
morphology. 相似文献
883.
Gomez C Conejero G Torregrosa L Cheynier V Terrier N Ageorges A 《The Plant journal : for cell and molecular biology》2011,67(6):960-970
In cells, anthocyanin pigments are synthesized at the cytoplasmic surface of the endoplasmic reticulum, and are then transported and finally accumulated inside the vacuole. In Vitis vinifera (grapevine), two kinds of molecular actors are putatively associated with the vacuolar sequestration of anthocyanins: a glutathione-S-transferase (GST) and two MATE-type transporters, named anthoMATEs. However, the sequence of events by which anthocyanins are imported into the vacuole remains unclear. We used MYBA1 transformed hairy roots as a grapevine model tissue producing anthocyanins, and took advantage of the unique autofluorescence of anthocyanins to study their cellular trafficking. In these tissues, anthocyanins were not only visible in the largest vacuoles, but were also present at higher concentrations in several vesicles of different sizes. In the cell, small vesicles actively moved alongside the tonoplast, suggesting a vesicular trafficking to the vacuole. Subcellular localization assays revealed that anthoMATE transporters were closely related with these small vesicles, whereas GST was localized in the cytoplasm around the nucleus, suggesting an association with the endoplasmic reticulum. Furthermore, cells in hairy roots expressing anthoMATE antisense did not display small vesicles filled with anthocyanins, whereas in hairy roots expressing GST antisense, anthocyanins were accumulated in vesicles but not in the vacuole. This suggests that in grapevine, anthoMATE transporters and GST are involved in different anthocyanin transport mechanisms. 相似文献
884.
Moreira A Arsati F de Oliveira Lima-Arsati YB de Freitas CG de Araújo VC 《Journal of strength and conditioning research / National Strength & Conditioning Association》2011,25(7):1932-1936
The purpose of this study was to investigate the responses of salivary immunoglobulin A (SIgA) in 10 professional top-level Brazilian futsal players after 2 highly competitive games separated by 7 days. Unstimulated saliva was collected over a 5-minute period at PRE- and POST-match. The SIgA was measured by an enzyme-linked immunosorbent assay and expressed as the absolute concentration (SIgAabs) and secretion rate of IgA (SIgArate). Rate of perceived exertion and heart rate were used to monitor the exercise intensity. A 2-way analysis of variance with repeated measures showed nonsignificant differences between matches to SIgAabs, SIgArate, and saliva flow rate (p > 0.05). However, significant time differences were observed for all these parameters. In summary, we showed that a competitive training match induced a decrease in SIgA levels in top-level futsal players, which suggests an increment of the vulnerability to infections meditated by the training stimulus. This decrease suggests that the athletes were at an increased risk of developing an upper respiratory tract infection, and therefore, it could be necessary to take protective actions to minimize contact with cold viruses or even reduce the training load for athletes. 相似文献
885.
Camila A. Wilkens Claudia Altamirano Ziomara P. Gerdtzen 《Biotechnology and Bioprocess Engineering》2011,16(4):714-724
t-PA producing CHO cells have been shown to undergo a metabolic shift when the culture medium is supplemented with a mixture
of glucose and galactose. This metabolic change is characterized by the reincorporation of lactate and its use as an additional
carbon source. The aim of this work is to understand lactate metabolism. To do so, Chinese hamster ovary cells were grown
in batch cultures in four different conditions consisting in different combinations of glucose and galactose. In experiments
supplemented with glucose, only lactate production was observed. Cultures with glucose and galactose consumed glucose first
and produced lactate at the same time, after glucose depletion galactose consumption began and lactate uptake was observed.
Comparison of the metabolic state of cells with and without the shift by metabolic flux analysis show that the metabolic fluxes
distribution changes mostly in the reactions involving pyruvate metabolism. When not enough pyruvate is being produced for
cells to support their energy requirements, lactate dehydrogenase complex changes the direction of the reaction yielding pyruvate
to feed the TCA cycle. The slow change from high fluxes during glucose consumption to low fluxes in galactose consumption
generates intracellular conditions that allow the influx of lactate. Lactate consumption is possible in cell cultures supplemented
with glucose and galactose due to the low rates at which galactose is consumed. Evidence suggests that an excessive production
and accumulation of pyruvate during glucose consumption leads to lactate production and accumulation inside the cell. Other
internal conditions such as a decrease in internal pH, forces the flow of lactate outside the cell. After metabolic shift
the intracellular pool of pyruvate, lactate and H+ drops permitting the reversal of the monocarboxylate transporter direction, therefore leading to lactate uptake. Metabolic
analysis comparing glucose and galactose consumption indicates that after metabolic shift not enough pyruvate is produced
to supply energy metabolism and lactate is used for pyruvate synthesis. In addition, MFA indicates that most carbon consumed
during low carbon flux is directed towards maintaining energy metabolism. 相似文献
886.
887.
Luciana Dalla Rosa Aleksandro S. Da Silva Jader B. Ruchel Lucas T. Gressler Camila B. Oliveira Raqueli T. França Sonia T.A. Lopes Daniela B.R. Leal Silvia G. Monteiro 《Experimental parasitology》2013
This study aimed to verify the effect of 3′-deoxyadenosine and deoxycoformycin on hematologic parameters and adenosine deaminase (ADA) activity in plasma and brain of mice infected with Trypanosoma evansi. Seventy animals were divided into seven groups, which were divided into two subgroups each for sampling on days 4 and 8 post-infection (PI). The groups were composed of three uninfected groups (A–C), namely, not-treated (A), treated with 3′-deoxyadenosine (B), and treated with deoxycoformycin (C) and four infected groups, mice with T. evansi (D–G), namely, not-treated (D), treated with 3′-deoxyadenosine (E), treated with deoxycoformycin (F), and treated with a combination 3′-deoxyadenosine and deoxycoformycin (G). Hematological parameters and ADA activity were evaluated in plasma and brain. Animals in groups B and C exhibited a reduction in the levels of plasma total protein compared group A. Animals in groups D and F showed changes in the hematological parameters. The ADA activity significantly reduced in the animals of groups C, D, F and G. Mice in the group E presented increased ADA activity in plasma. Therefore, we conclude that the treatment interferes significantly in the hematologic parameters in mice infected with T. evansi. On the other hand, when the ADA inhibitor was used we observed a significant decrease in the values of hematocrit, total erythrocytes, and hemoglobin concentration. The deoxycoformycin was able to inhibit the ADA activity of parasite thus it may be one of the mechanisms of efficacy of this treatment. 相似文献
888.
889.
Valentina Parra Francisco Moraga Jovan Kuzmicic Camila López-Crisosto Rodrigo Troncoso Natalia Torrealba Alfredo Criollo Jessica Díaz-Elizondo Beverly A. Rothermel Andrew F.G. Quest Sergio Lavandero 《生物化学与生物物理学报:疾病的分子基础》2013,1832(8):1334-1344
Ceramides are important intermediates in the biosynthesis and degradation of sphingolipids that regulate numerous cellular processes, including cell cycle progression, cell growth, differentiation and death. In cardiomyocytes, ceramides induce apoptosis by decreasing mitochondrial membrane potential and promoting cytochrome-c release. Ca2 + overload is a common feature of all types of cell death. The aim of this study was to determine the effect of ceramides on cytoplasmic Ca2 + levels, mitochondrial function and cardiomyocyte death. Our data show that C2-ceramide induces apoptosis and necrosis in cultured cardiomyocytes by a mechanism involving increased Ca2 + influx, mitochondrial network fragmentation and loss of the mitochondrial Ca2 + buffer capacity. These biochemical events increase cytosolic Ca2 + levels and trigger cardiomyocyte death via the activation of calpains. 相似文献
890.
Mauricio Rodriguez‐Lanetty Camila Granados‐Cifuentes Albert Barberan Anthony J. Bellantuono Carolina Bastidas 《Molecular ecology》2013,22(16):4349-4362
The functional role of the bacterial organisms in the reef ecosystem and their contribution to the coral well‐being remain largely unclear. The first step in addressing this gap of knowledge relies on in‐depth characterization of the coral microbial community and its changes in diversity across coral species, space and time. In this study, we focused on the exploration of microbial community assemblages associated with an ecologically important Caribbean scleractinian coral, Porites astreoides, using Illumina high‐throughput sequencing of the V5 fragment of 16S rRNA gene. We collected data from a large set of biological replicates, allowing us to detect patterns of geographical structure and resolve co‐occurrence patterns using network analyses. The taxonomic analysis of the resolved diversity showed consistent and dominant presence of two OTUs affiliated with the order Oceanospirillales, which corroborates a specific pattern of bacterial association emerging for this coral species and for many other corals within the genus Porites. We argue that this specific association might indicate a symbiotic association with the adult coral partner. Furthermore, we identified a highly diverse rare bacterial ‘biosphere’ (725 OTUs) also living along with the dominant bacterial symbionts, but the assemblage of this biosphere is significantly structured along the geographical scale. We further discuss that some of these rare bacterial members show significant association with other members of the community reflecting the complexity of the networked consortia within the coral holobiont. 相似文献