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181.
de-Freitas-Junior JC Bastos LG Freire-Neto CA Rocher BD Abdelhay ES Morgado-Díaz JA 《Journal of cellular biochemistry》2012,113(9):2957-2966
During malignant transformation, changes in the expression profile of glycans may be involved in a variety of events, including the loss of cell-cell and cell-matrix adhesion, migration, invasion, and evasion of apoptosis. Therefore, modulation of glycan expression with drugs has promising therapeutic potential for various cancer types. In this study, we investigated the in vitro anticancer activity of the N-glycan biosynthesis inhibitors (swainsonine and tunicamycin) in cells derived from colorectal cancer (CRC). We also examined whether these inhibitors are able to induce radiosensitization and toxicity when used in combination with cisplatin or irinotecan, two current anticancer drugs. Our results show that treatment with tunicamycin inhibits cellular mechanisms related to the malignant phenotype, such as anchorage-dependent and anchorage-independent colony formation, migration and invasion, in undifferentiated HCT-116 colon cancer cells, whereas swainsonine only inhibits cell migration. We also observed that tunicamycin, but not swainsonine, caused radiosensitivity in HCT-116 cells. Moreover, the combination of swainsonine with cisplatin or irinotecan enhanced their toxicity in HCT-116 cells, while the combination of tunicamycin with these drugs had no effect. Given these results, we suggest that the modulation of N-glycan biosynthesis appears to be a potential therapeutic tool for CRC treatment because inhibition of this process induced anticancer activity in vitro. Additionally, the inhibition of the N-glycan biosynthesis in combination with chemotherapic drugs is a promising therapeutic strategy for enhancing radiation therapy. 相似文献
182.
183.
Cavasini CE De Mattos LC Alves RT Couto AA Calvosa VS Domingos CR Castilho L Rossit AR Machado RL 《Human biology; an international record of research》2006,78(2):215-219
We compared the serological phenotypic frequencies of ABO, MNSs, and Duffy in 417 blood donors and 309 malaria patients from four Brazilian Amazon areas. Our results suggest no correlation between ABO phenotype and malaria infection in all areas studied. We observed significant correlation between the S +s +, S +s -, and S -s + phenotypes and malaria infection in three areas. Some of the Duffy phenotypes showed significant correlation between donors and malaria patients in different areas. These data are an additional contribution to the establishment of differential host susceptibility to malaria. 相似文献
184.
185.
Desyreé Murta Jesus Lilian T. Costa Daniela L. Gon?alves Carlos Alberto Achete Marcia Attias Nissin Moussatché Clarissa R. Damaso 《Journal of virology》2009,83(22):11477-11490
Cidofovir (CDV) is one of the most effective antiorthopoxvirus drugs, and it is widely accepted that viral DNA replication is the main target of its activity. In the present study, we report a detailed analysis of CDV effects on the replicative cycles of distinct vaccinia virus (VACV) strains: Cantagalo virus, VACV-IOC, and VACV-WR. We show that despite the approximately 90% inhibition of production of virus progeny, virus DNA accumulation was reduced only 30%, and late gene expression and genome resolution were unaltered. The level of proteolytic cleavage of the major core proteins was diminished in CDV-treated cells. Electron microscopic analysis of virus-infected cells in the presence of CDV revealed reductions as great as 3.5-fold in the number of mature forms of virus particles, along with a 3.2-fold increase in the number of spherical immature particles. A detailed analysis of purified virions recovered from CDV-treated cells demonstrated the accumulation of unprocessed p4a and p4b and nearly 67% inhibition of DNA encapsidation. However, these effects of CDV on virus morphogenesis resulted from a primary effect on virus DNA synthesis, which led to later defects in genome encapsidation and virus assembly. Analysis of virus DNA by atomic force microscopy revealed that viral cytoplasmic DNA synthesized in the presence of CDV had an altered structure, forming aggregates with increased strand overlapping not observed in the absence of the drug. These aberrant DNA aggregations were not encapsidated into virus particles.Vaccinia virus (VACV) is the prototypical member of the Poxviridae, a family of large DNA-containing viruses. During infection, a cascade of temporally regulated viral gene expression occurs exclusively in the cell cytoplasm, where viral DNA replication takes place. DNA replication is essential for the onset of the intermediate and late steps of viral gene expression (37). VACV morphogenesis is a complex process that starts within the virus factories, or virosomes, in parallel with the late stage of gene expression. Crescent-shaped virus membranes evolve into immature spherical particles (IV) that subsequently progress to form brick-shaped mature virions (MV) (reviewed in reference 8). Virus assembly and maturation are complex processes requiring telomere resolution of newly replicated DNA (20, 36, 40), genome encapsidation (6, 22, 50), and the proteolytic processing of major structural proteins (38, 51). For the past decade, several reports have analyzed in detail these numerous steps of VACV morphogenesis, unraveling the role of distinct virus late proteins in the progression of viral particle formation (reviewed in reference 8).Cantagalo virus (CTGV) is a strain of VACV isolated from pustular lesions on cows in Brazil (10). Similar outbreaks of vaccinia-like viruses have been reported frequently over the past 8 years (14, 39, 48). Interestingly, the majority of these vaccinia viruses circulating in the wild in Brazil bear a striking similarity to the Brazilian vaccine strain used for systematic vaccination during the eradication campaign, which was produced in Rio de Janeiro (19) and called strain IOC (10). This similarity raises the interesting possibility that the circulating vaccinia viruses represent feral derivatives of IOC or of a closely related ancestor. Little is known about the sensitivity of these novel vaccinia viruses to antiviral compounds. In the absence of an active smallpox vaccination campaign, the spread of these vaccinia viruses in the wild, the prevalence of cowpox infections in Europe and elsewhere (39), and the occurrence of complications from smallpox vaccination (52) make the need for effective antipoxvirus treatment a worldwide concern.Cidofovir (CDV), an acyclic pyrimidine phosphonate analogue, has shown a potent antiviral effect on several poxvirus infections (4, 15, 44, 46). Recently, we have reported the efficacy of CDV in inhibiting the replication of the Brazilian VACV strains CTGV and IOC (26). The mechanism of action of CDV on reactions catalyzed by the VACV DNA polymerase has been studied in vitro. CDV is not a chain-terminating analogue but drastically slows chain extension and inhibits the 3′-5′ exonuclease proofreading activity of the enzyme (34). In addition, templates containing CDV cause inhibition of DNA elongation (33). Differences between the effects of CDV on human cytomegalovirus (55) and VACV enzymes have been observed, but overall it has been widely accepted that CDV acts by inhibiting the process of virus DNA replication. Moreover, most CDV-resistant VACV strains contain mutations in the catalytic domain or in the 3′-5′ exonuclease domain of the DNA polymerase (2, 5, 28, 45).Despite the consensus regarding mechanisms of action, the effects of CDV on the stages of the VACV replicative cycle have never been analyzed. We report here that although CDV led to approximately 90% inhibition of VACV progeny production, we observed only 30% inhibition of DNA replication and normal levels of postreplicative virus gene expression. However, the encapsidation of DNA into virus particles and the proteolytic processing of the major core proteins were inhibited in CDV-treated cells, leading to an impairment of virus morphogenesis. These effects on virus assembly are an indirect result of a primary effect of CDV on VACV DNA synthesis. Atomic force microscopy (AFM) analysis revealed that virus DNA isolated from the cytoplasm of CDV-treated cells formed aggregates of highly entangled and intertwined DNA molecules that were not observed in cytoplasmic viral DNA isolated from untreated cells. In addition, these DNA aggregates were not detected in encapsidated virus genomes isolated from particles purified from untreated or CDV-treated cells. Our data suggest that incorporation of CDV into VACV DNA during the replication process may lead to aberrant DNA structures, which are less able to be packaged into virus particles. 相似文献
186.
Longbiao Guo ;Lilian Ma ;Hua Jiang ;Dali Zeng ;Jiang Hu ;Liwen Wu ;Zhenyu Gao ;Guangheng Zhang ;Qian Qian 《Acta Botanica Sinica》2009,(1):45-51
To identify genetic loci controlling grain weight, an elite indica rice variety, Baodali, with large grains was identified and used in this study. Its derived F2, F3 and BC2 F2 with another japonica rice variety Zhonghua 11 were used as mapping populations. Linkage analyses demonstrated that two genes controlling grain weight, designated as GW3 and GW6, were mapped to chromosome 3 and chromosome 6, respectively. Fine mapping delimited GW3 to a 122 kb physical distance between two sequence tagged site markers (WGWt6 and WGW19) containing 16 open reading frames annotated by The Institute for Genomic Research (http://www.tigr.org). GW6 was further mapped between two simple sequence repeat markers (RM7179 and RM3187). These results are useful for both marker assisted selection of grain weight, and for further cloning of GW genes, which will contribute to the dissection of the molecular mechanism underlying grain weight in rice. 相似文献
187.
Lilian S. Catenacci Kristel M. De Vleeschouwer Sérgio L. G. Nogueira-Filho 《Biotropica》2009,41(6):744-750
Seed dispersal by small primates may be particularly relevant in areas where populations of larger frugivores have been reduced or extinguished by hunting and/or habitat disturbance. In this context, the aim of this study was to evaluate the role of the golden-headed lion tamarin Leontopithecus chrysomelas as a seed disperser in Atlantic forest remnants in Brazil. To this end, we opportunistically collected feces deposited during observations on the feeding behavior of two groups of golden-headed lion tamarins ranging in the degraded areas of the Una Biological Reserve, Bahia, Brazil, from February 2006 to January 2007. We collected 587 fecal samples, of which 524 contained seeds from 24 plant species, distributed over 13 families. Disregarding seeds of<3 mm, the majority of seeds recovered were bromeliad seeds. In general, ingestion of seeds by golden-headed lion tamarins did not improve the germination proportion or decrease the germination delay of seeds, with the exception of Aechmea spp. seeds. The tamarins encountered different habitats during their daily activity period, while feeding and defecating. Consequently, some seeds were transported to different habitats including disturbed areas. Thus, the role of seed dispersal in combination with the daily movement pattern of L. chrysomelas contributes to the persistence of fruit plants and epiphyte species and to the natural regeneration process within Atlantic forest remnants. 相似文献
188.
Thaís Leal-Silva Flaviane Vieira-Santos Fabrício Marcus Silva Oliveira Luiza de Lima Silva Padro Lucas Kraemer Pablo Hemanoel da Paixo Matias Camila de Almeida Lopes Ana Cristina Loiola Ruas Isabella Carvalho de Azevedo Denise Silva Nogueira Milene Alvarenga Rachid Marcelo Vidigal Caliari Remo Castro Russo Ricardo Toshio Fujiwara Lilian Lacerda Bueno 《PLoS neglected tropical diseases》2021,15(7)
Toxocariasis is a neglected disease that affects people around the world. Humans become infected by accidental ingestion of eggs containing Toxocara canis infective larvae, which upon reaching the intestine, hatch, penetrate the mucosa and migrate to various tissues such as liver, lungs and brain. Studies have indicated that Th2 response is the main immune defense mechanism against toxocariasis, however, there are still few studies related to this response, mainly the IL-33/ST2 pathway. Some studies have reported an increase in IL-33 during helminth infections, including T. canis. By binding to its ST2 receptor, IL-33 stimulating the Th2 polarized immune cell and cytokine responses. Thus, we aimed to investigate the role of the IL-33/ST2 pathway in the context of T. canis larval migration and the immunological and pathophysiological aspects of the infection in the liver, lungs and brain from Wild-Type (WT) BALB/c background and genetically deficient mice for the ST2 receptor (ST2-/-). The most important findings revealed that the IL-33/ST2 pathway is involved in eosinophilia, hepatic and cerebral parasitic burden, and induces the formation of granulomas related to tissue damage and pulmonary dysfunction. However, ST2-/- mice, the immune response was skewed to Th1/Th17 type than Th2, that enhanced the control of parasite burden related to IgG2a levels, tissue macrophages infiltration and reduced lung dysfunction. Collectively, our results demonstrate that the Th2 immune response triggered by IL-33/ST2 pathway mediates susceptibility to T. canis, related to parasitic burden, eosinophilia and granuloma formation in which consequently contributes to tissue inflammation and injury. 相似文献
189.
Denise A. Berti Cain Morano Lilian C. Russo Leandro M. Castro Fernanda M. Cunha Xin Zhang Juan Sironi Cl��cio F. Klitzke Emer S. Ferro Lloyd D. Fricker 《The Journal of biological chemistry》2009,284(21):14105-14116
Thimet oligopeptidase (EC 3.4.24.15; EP24.15) is an intracellular enzyme
that has been proposed to metabolize peptides within cells, thereby affecting
antigen presentation and G protein-coupled receptor signal transduction.
However, only a small number of intracellular substrates of EP24.15 have been
reported previously. Here we have identified over 100 peptides in human
embryonic kidney 293 (HEK293) cells that are derived from intracellular
proteins; many but not all of these peptides are substrates or products of
EP24.15. First, cellular peptides were extracted from HEK293 cells and
incubated in vitro with purified EP24.15. Then the peptides were
labeled with isotopic tags and analyzed by mass spectrometry to obtain
quantitative data on the extent of cleavage. A related series of experiments
tested the effect of overexpression of EP24.15 on the cellular levels of
peptides in HEK293 cells. Finally, synthetic peptides that corresponded to 10
of the cellular peptides were incubated with purified EP24.15 in
vitro, and the cleavage was monitored by high pressure liquid
chromatography and mass spectrometry. Many of the EP24.15 substrates
identified by these approaches are 9–11 amino acids in length,
supporting the proposal that EP24.15 can function in the degradation of
peptides that could be used for antigen presentation. However, EP24.15 also
converts some peptides into products that are 8–10 amino acids, thus
contributing to the formation of peptides for antigen presentation. In
addition, the intracellular peptides described here are potential candidates
to regulate protein interactions within cells.Intracellular protein turnover is a crucial step for cell functioning, and
if this process is impaired, the elevated levels of aged proteins usually lead
to the formation of intracellular insoluble aggregates that can cause severe
pathologies (1). In mammalian
cells, most proteins destined for degradation are initially tagged with a
polyubiquitin chain in an energy-dependent process and then digested to small
peptides by the 26 S proteasome, a large proteolytic complex involved in the
regulation of cell division, gene expression, and other key processes
(2,
3). In eukaryotes, 30–90%
of newly synthesized proteins may be degraded by proteasomes within minutes of
synthesis (3,
4). In addition to proteasomes,
other extralysosomal proteolytic systems have been reported
(5,
6). The proteasome cleaves
proteins into peptides that are typically 2–20 amino acids in length
(7). In most cases, these
peptides are thought to be rapidly hydrolyzed into amino acids by
aminopeptidases
(8–10).
However, some intracellular peptides escape complete degradation and are
imported into the endoplasmic reticulum where they associate with major
histocompatibility complex class I
(MHC-I)3 molecules and
traffic to the cell surface for presentation to the immune system
(10–12).
Additionally, based on the fact that free peptides added to the intracellular
milieu can regulate cellular functions mediated by protein interactions such
as gene regulation, metabolism, cell signaling, and protein targeting
(13,
14), intracellular peptides
generated by proteasomes that escape degradation have been suggested to play a
role in regulating protein interactions
(15). Indeed, oligopeptides
isolated from rat brain tissue using the catalytically inactive EP24.15 (EC
3.4.24.15) were introduced into Chinese hamster ovarian-S and HEK293 cells and
were found capable of altering G protein-coupled receptor signal transduction
(16). Moreover, EP24.15
overexpression itself changed both angiotensin II and isoproterenol signal
transduction, suggesting a physiological function for its intracellular
substrates/products (16).EP24.15 is a zinc-dependent peptidase of the metallopeptidase M3 family
that contains the HEXXH motif
(17). This enzyme was first
described as a neuropeptide-degrading enzyme present in the soluble fraction
of brain homogenates (18).
Whereas EP24.15 can be secreted
(19,
20), its predominant location
in the cytosol and nucleus suggests that the primary function of this enzyme
is not the extracellular degradation of neuropeptides and hormones
(21,
22). EP24.15 was shown in
vivo to participate in antigen presentation through MHC-I
(23–25)
and in vitro to bind
(26) or degrade
(27) some MHC-I associated
peptides. EP24.15 has also been shown in vitro to degrade peptides
containing 5–17 amino acids produced after proteasome digestion of
β-casein (28). EP24.15
shows substrate size restriction to peptides containing from 5 to 17 amino
acids because of its catalytic center that is located in a deep channel
(29). Despite the size
restriction, EP24.15 has a broad substrate specificity
(30), probably because a
significant portion of the enzyme-binding site is lined with potentially
flexible loops that allow reorganization of the active site following
substrate binding (29).
Recently, it has also been suggested that certain substrates may be cleaved by
an open form of EP24.15 (31).
This characteristic is supported by the ability of EP24.15 to accommodate
different amino acid residues at subsites S4 to S3′, which even includes
the uncommon post-proline cleavage
(30). Such biochemical and
structural features make EP24.15 a versatile enzyme to degrade structurally
unrelated oligopeptides.Previously, brain peptides that bound to catalytically inactive EP24.15
were isolated and identified using mass spectrometry
(22). The majority of peptides
captured by the inactive enzyme were intracellular protein fragments that
efficiently interacted with EP24.15; the smallest peptide isolated in these
assays contained 5 and the largest 17 amino acids
(15,
16,
22,
32), which is within the size
range previously reported for natural and synthetic substrates of EP24.15
(18,
30,
33,
34). Interestingly, the
peptides released by the proteasome are in the same size range of EP24.15
competitive inhibitors/substrates
(7,
35,
36). Taken altogether, these
data suggest that in the intracellular environment EP24.15 could further
cleave proteasome-generated peptides unrelated to MHC-I antigen presentation
(15).Although the mutated inactive enzyme “capture” assay was
successful in identifying several cellular protein fragments that were
substrates for EP24.15, it also found some interacting peptides that were not
substrates. In this study, we used several approaches to directly screen for
cellular peptides that were cleaved by EP24.15. The first approach involved
the extraction of cellular peptides from the HEK293 cell line, incubation
in vitro with purified EP24.15, labeling with isotopic tags, and
analysis by mass spectrometry to obtain quantitative data on the extent of
cleavage. The second approach examined the effect of EP24.15 overexpression on
the cellular levels of peptides in the HEK293 cell line. The third set of
experiments tested synthetic peptides with purified EP24.15 in vitro,
and examined cleavage by high pressure liquid chromatography and mass
spectrometry. Collectively, these studies have identified a large number of
intracellular peptides, including those that likely represent the endogenous
substrates and products of EP24.15, and this original information contributes
to a better understanding of the function of this enzyme in vivo. 相似文献
190.
Ernesto S Nakayasu Lilian L Nohara Ana Claudia T Torrecilhas Andrei V Nikolaev Igor C Almeida 《Molecular systems biology》2009,5(1)
Glycosylphosphatidylinositol (GPI) anchoring is a common, relevant posttranslational modification of eukaryotic surface proteins. Here, we developed a fast, simple, and highly sensitive (high attomole‐low femtomole range) method that uses liquid chromatography‐tandem mass spectrometry (LC‐MSn) for the first large‐scale analysis of GPI‐anchored molecules (i.e., the GPIome) of a eukaryote, Trypanosoma cruzi, the etiologic agent of Chagas disease. Our genome‐wise prediction analysis revealed that approximately 12% of T. cruzi genes possibly encode GPI‐anchored proteins. By analyzing the GPIome of T. cruzi insect‐dwelling epimastigote stage using LC‐MSn, we identified 90 GPI species, of which 79 were novel. Moreover, we determined that mucins coded by the T. cruzi small mucin‐like gene (TcSMUG S) family are the major GPI‐anchored proteins expressed on the epimastigote cell surface. TcSMUG S mucin mature sequences are short (56–85 amino acids) and highly O‐glycosylated, and contain few proteolytic sites, therefore, less likely susceptible to proteases of the midgut of the insect vector. We propose that our approach could be used for the high throughput GPIomic analysis of other lower and higher eukaryotes. 相似文献