Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules...Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules. Despite extensive research on nsLTP in different plant species, mostly angiosperm, and the great diversity of physiological processes in which they seem to be involved, their exact functions still remain unclear. Also, very limited experimental data are available on nsLTP in gymnosperm. In this study, we report for the first time on the molecular cloning of nsLTP, from Pinus sylvestris L.(PsLTP1, GenBank accession JN980402.1) and the expression pattern of PsLTP1 during ontogenesis and in response to environmental stress conditions. Total RNA from roots of 7-day old pine seedlings was used to isolate the cDNA clone, corresponding to Scots pine lipid transfer protein. The open reading frame of PsLTP1 consists of 372 bp encoding a protein of 123 amino acids. Amino acid sequence alignment revealed that mature PsLTP1 shares high level of similarity with nsLTP from other conifers and with well-studied nsLTPs from angiosperms. The PsLTP1 contains a 27-amino-acid N-terminal signal sequence and presents all the features of a plant nsLTP. Amino acid comparison analysis and 3D structure prediction showed that PsLTP1 is a type 1 nsLTP. The results of the expression analysis of Scots pine PsLTP1 gene revealed that its transcripts accumulate in actively growing tissues. Furthermore, transcription of PsLTP1 was upregulated in response to cold and salt treatments, and downregulated during acidic, osmotic and water stresses.展开更多
Ethanol (EtOH) enhances glycinergic currents in the central nervous system (CNS). Because evidence for an interaction between the α1 subunit of the glycine receptor (α1GlyR) and the G protein Gβγ subunit exists in...Ethanol (EtOH) enhances glycinergic currents in the central nervous system (CNS). Because evidence for an interaction between the α1 subunit of the glycine receptor (α1GlyR) and the G protein Gβγ subunit exists in vitro and because cAMP levels are known to increase in response to EtOH, we wanted to investigate the interaction between Gβγ and α1GlyR in response to EtOH treatment in HEK293 cells and to explore the possible sites of interaction between EtOH and the Gαs subunit. His pull-down assays in GlyR-His6-transfected HEK293 cells incubated with ethanol or propofol revealed that only EtOH treatment increased the binding of Gβγ heterodimers to α1GlyR. Using molecular modelling (protein structure prediction), was modelled the hGαs protein for the first time and validated this model by site-directed mutagenesis. By molecular docking, we identified some potential regions of interaction between hGαs and EtOH that are located on the SIII and SI regions of the Gαs. Therefore, we conclude that ethanol increases the interaction between α1GlyR and Gβγ in HEK293 cells, an effect that might be attributed to the interaction between EtOH and hGαs, which consequently stimulates hGαs.展开更多
KDR （kinase insert domain receptor） phosphorylation induces several effects which lead eventually to cell proliferation and survival. The precise mechanisms by which KDR, once it is activated, communicates with the ...KDR （kinase insert domain receptor） phosphorylation induces several effects which lead eventually to cell proliferation and survival. The precise mechanisms by which KDR, once it is activated, communicates with the nucleus are starting to be understood but have not yet been completely unravelled. Two in vitro studies on animal cell lines reported in the literature have demonstrated that, following stimulation with VEGF, KDR is actually translocated within the nucleus. Our aim was to investigate whether this translocation occurs in human cells both in vitro and in vivo. Using laser scanning confocal microscopy, a variable nuclear localization of phosphorylated and total KDR in cell lines and tumour samples was found. In human neoplastic cell lines, hypoxic stimulation greatly increased the nuclear amount of total KDR but less so that of the phosphorylated form. Only after hypoxia and VEGF stimulation there was a comparably increased expression of phosphorylated and total KDR observed in the nuclei of these cells. We conclude that neoplastic cells show a variable expression of total and phosphorylated KDR in the nucleus. The precise functional meaning of nuclear location remains to be established.展开更多
The aim of this paper is to study the disaccharidase profile in GD （Gaucher disease） patients treated or not with miglustat and compare it with a healthy control group. Miglustat is an iminosugar used as substrate i...The aim of this paper is to study the disaccharidase profile in GD （Gaucher disease） patients treated or not with miglustat and compare it with a healthy control group. Miglustat is an iminosugar used as substrate inhibitor in the therapy of some lysosomal disorders, its main side effects resembling carbohydrate maldigestion symptoms and cause more than 50% of medication discontinuation among GD patients. In-vitro studies have revealed that miglustat can act as an inhibitor of some digestive enzymes. An exploratory non-interventional study was designed to compare the disaccharidase profile assessed by MHBT （methane hydrogen breath test） and to analyze the correlation with the reported gastrointestinal symptoms in GD patients （40） and healthy subjects （20）. MHBT was performed following the ingestion of lactose, sucrose and maltose on different days. Each participant completed two detailed surveys about dietary habits, medications and gastrointestinal symptoms previous and during the test. Twenty-one GD were receiving miglustat, 10 （47.6%） of them reported gastrointestinal side effects, and 7/10 （70%） recorded a positive MHBT （lactose 5, maltose 2, and sucrose 1）. In 6/19 （31.6%） patients that never been exposed to miglustat and 7/20 （35%） controls a positive MHBT were detected. The comparison of the malabsorption phenotype between GD patients exposed and not exposed to miglustat （p = 0.028） and control group （p 〈 0.04） showed high statistical significance for the group of patients treated with miglustat. These results suggest that miglustat therapy induces persistent changes in digestive enzyme activity in GD patients.展开更多
The K/HDEL receptor ERD2 mediates the transport of soluble endoplasmic reticulum （ER）-resident proteins containing a C-terminal K/HDEL signal from the Golgi apparatus back to the ER via COPI （COat Protein I）-coate...The K/HDEL receptor ERD2 mediates the transport of soluble endoplasmic reticulum （ER）-resident proteins containing a C-terminal K/HDEL signal from the Golgi apparatus back to the ER via COPI （COat Protein I）-coated vesicles. Sorting of ERD2 within COPI vesicles is facilitated by p24 proteins. In Arabidop- sis, p24δ5 has been shown to interact directly with ERD2 via its luminal GOLD （GOLgi Dynamics） domain and with COPI proteins via its cytoplasmic C-terminal tail at the acidic pH of the Golgi apparatus. Several members of the p24 family in mammals and yeast have been shown to be glycosylated, but whether Arabidopsis p24 proteins are glycosylated and the role of the sugar moiety in p24 function remain unclear. Here, we show that Arabidopsis p24δ5 protein is N-glycosylated in its GOLD domain. Furthermore, we demonstrate that this post-translational modification is important for its coupled transport with p241β2 at the ER-Golgi interface, for its interaction with the K/HDEL receptor ERD2, and for retrograde transport of ERD2 and K/HDEL ligands from the Golgi apparatus back to the ER.展开更多
Dear Editor, During recent decades, a novel mechanism of secretion has been identified in a wide range of mammalian cells. It involves the release of bioactive membrane nanovesicles （30-100 nm）, termed exosomes, up...Dear Editor, During recent decades, a novel mechanism of secretion has been identified in a wide range of mammalian cells. It involves the release of bioactive membrane nanovesicles （30-100 nm）, termed exosomes, upon the fusion of multivesicular bodies with the plasma membrane （Thery et al., 2009）. Exosomes are implicated in diverse functions, such as scavenging of archaic proteins, intercellular messengers delivering cell-specific signals, and vehicles for transmissible pathogens. Exosomes have also been described in other organisms such as bacte- ria, Drosophila, and fungi.展开更多
基金supported by grants from the Ministry of Education and Science of Ukraine(0116U003593)grant from cieA3(Campus de Excelencia Internacional Agroalimentario)-UCO,Spain
文摘Plant nonspecific lipid transfer proteins (nsLTPs) are widely distributed through plant kingdom and are characterized by the presence of a central hydrophobic cavity, suitable for binding various hydrophobic molecules. Despite extensive research on nsLTP in different plant species, mostly angiosperm, and the great diversity of physiological processes in which they seem to be involved, their exact functions still remain unclear. Also, very limited experimental data are available on nsLTP in gymnosperm. In this study, we report for the first time on the molecular cloning of nsLTP, from Pinus sylvestris L.(PsLTP1, GenBank accession JN980402.1) and the expression pattern of PsLTP1 during ontogenesis and in response to environmental stress conditions. Total RNA from roots of 7-day old pine seedlings was used to isolate the cDNA clone, corresponding to Scots pine lipid transfer protein. The open reading frame of PsLTP1 consists of 372 bp encoding a protein of 123 amino acids. Amino acid sequence alignment revealed that mature PsLTP1 shares high level of similarity with nsLTP from other conifers and with well-studied nsLTPs from angiosperms. The PsLTP1 contains a 27-amino-acid N-terminal signal sequence and presents all the features of a plant nsLTP. Amino acid comparison analysis and 3D structure prediction showed that PsLTP1 is a type 1 nsLTP. The results of the expression analysis of Scots pine PsLTP1 gene revealed that its transcripts accumulate in actively growing tissues. Furthermore, transcription of PsLTP1 was upregulated in response to cold and salt treatments, and downregulated during acidic, osmotic and water stresses.
文摘Ethanol (EtOH) enhances glycinergic currents in the central nervous system (CNS). Because evidence for an interaction between the α1 subunit of the glycine receptor (α1GlyR) and the G protein Gβγ subunit exists in vitro and because cAMP levels are known to increase in response to EtOH, we wanted to investigate the interaction between Gβγ and α1GlyR in response to EtOH treatment in HEK293 cells and to explore the possible sites of interaction between EtOH and the Gαs subunit. His pull-down assays in GlyR-His6-transfected HEK293 cells incubated with ethanol or propofol revealed that only EtOH treatment increased the binding of Gβγ heterodimers to α1GlyR. Using molecular modelling (protein structure prediction), was modelled the hGαs protein for the first time and validated this model by site-directed mutagenesis. By molecular docking, we identified some potential regions of interaction between hGαs and EtOH that are located on the SIII and SI regions of the Gαs. Therefore, we conclude that ethanol increases the interaction between α1GlyR and Gβγ in HEK293 cells, an effect that might be attributed to the interaction between EtOH and hGαs, which consequently stimulates hGαs.
文摘KDR （kinase insert domain receptor） phosphorylation induces several effects which lead eventually to cell proliferation and survival. The precise mechanisms by which KDR, once it is activated, communicates with the nucleus are starting to be understood but have not yet been completely unravelled. Two in vitro studies on animal cell lines reported in the literature have demonstrated that, following stimulation with VEGF, KDR is actually translocated within the nucleus. Our aim was to investigate whether this translocation occurs in human cells both in vitro and in vivo. Using laser scanning confocal microscopy, a variable nuclear localization of phosphorylated and total KDR in cell lines and tumour samples was found. In human neoplastic cell lines, hypoxic stimulation greatly increased the nuclear amount of total KDR but less so that of the phosphorylated form. Only after hypoxia and VEGF stimulation there was a comparably increased expression of phosphorylated and total KDR observed in the nuclei of these cells. We conclude that neoplastic cells show a variable expression of total and phosphorylated KDR in the nucleus. The precise functional meaning of nuclear location remains to be established.
文摘The aim of this paper is to study the disaccharidase profile in GD （Gaucher disease） patients treated or not with miglustat and compare it with a healthy control group. Miglustat is an iminosugar used as substrate inhibitor in the therapy of some lysosomal disorders, its main side effects resembling carbohydrate maldigestion symptoms and cause more than 50% of medication discontinuation among GD patients. In-vitro studies have revealed that miglustat can act as an inhibitor of some digestive enzymes. An exploratory non-interventional study was designed to compare the disaccharidase profile assessed by MHBT （methane hydrogen breath test） and to analyze the correlation with the reported gastrointestinal symptoms in GD patients （40） and healthy subjects （20）. MHBT was performed following the ingestion of lactose, sucrose and maltose on different days. Each participant completed two detailed surveys about dietary habits, medications and gastrointestinal symptoms previous and during the test. Twenty-one GD were receiving miglustat, 10 （47.6%） of them reported gastrointestinal side effects, and 7/10 （70%） recorded a positive MHBT （lactose 5, maltose 2, and sucrose 1）. In 6/19 （31.6%） patients that never been exposed to miglustat and 7/20 （35%） controls a positive MHBT were detected. The comparison of the malabsorption phenotype between GD patients exposed and not exposed to miglustat （p = 0.028） and control group （p 〈 0.04） showed high statistical significance for the group of patients treated with miglustat. These results suggest that miglustat therapy induces persistent changes in digestive enzyme activity in GD patients.
文摘The K/HDEL receptor ERD2 mediates the transport of soluble endoplasmic reticulum （ER）-resident proteins containing a C-terminal K/HDEL signal from the Golgi apparatus back to the ER via COPI （COat Protein I）-coated vesicles. Sorting of ERD2 within COPI vesicles is facilitated by p24 proteins. In Arabidop- sis, p24δ5 has been shown to interact directly with ERD2 via its luminal GOLD （GOLgi Dynamics） domain and with COPI proteins via its cytoplasmic C-terminal tail at the acidic pH of the Golgi apparatus. Several members of the p24 family in mammals and yeast have been shown to be glycosylated, but whether Arabidopsis p24 proteins are glycosylated and the role of the sugar moiety in p24 function remain unclear. Here, we show that Arabidopsis p24δ5 protein is N-glycosylated in its GOLD domain. Furthermore, we demonstrate that this post-translational modification is important for its coupled transport with p241β2 at the ER-Golgi interface, for its interaction with the K/HDEL receptor ERD2, and for retrograde transport of ERD2 and K/HDEL ligands from the Golgi apparatus back to the ER.
文摘Dear Editor, During recent decades, a novel mechanism of secretion has been identified in a wide range of mammalian cells. It involves the release of bioactive membrane nanovesicles （30-100 nm）, termed exosomes, upon the fusion of multivesicular bodies with the plasma membrane （Thery et al., 2009）. Exosomes are implicated in diverse functions, such as scavenging of archaic proteins, intercellular messengers delivering cell-specific signals, and vehicles for transmissible pathogens. Exosomes have also been described in other organisms such as bacte- ria, Drosophila, and fungi.
