Depolarization-dependent Induction of Site-specific Changes in Sialylation on N-linked Glycoproteins in Rat Nerve Terminals.
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ABSTRACT:
SUBMITTER: Boll I
PROVIDER: S-EPMC8143646 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Synaptic transmission is an ultra-fast process that aims at transducing a signal from one neuron to another neuron or cell by the release of neurotransmitters. This process requires rapid response which can be transmitted through fast molecular switches such as post-translational modifications of proteins. We aimed at investigating the potential importance in dynamic modulation of sialic acids on N-linked glycoproteins that are essential for the synaptic transmission process. For this, we characterize site-specific alteration in sialylated N-linked glycosylation in the active zone of rat nerve terminals (synaptosomes) after depolarization by using quantitative sialiomics.
2020-06-12 | PXD016230 | Pride
Project description:Glycoproteins containing N-linked oligosaccharides were prepared from plasma and liver microsomes of rats aged 0-5 weeks, and galactose and sialic acid content were determined. The sialic acid/galactose ratios in plasma membrane N-glycans remained at about 1 throughout the postnatal period, suggesting that most of the galactose residues are sialylated. In the same way, it was suggested that most of the galactose residues of microsomal N-glycans were sialylated at 0, 4 and 5 weeks of age, but that the degree of sialylation was lower at the other ages, with a minimum at 2 weeks. When the activities of sialyltransferase and galactosyltransferase in liver Golgi membranes were determined, age-dependent changes were found, not only in the specific activities of the enzymes, but also in the Golgi membrane content per g of liver. The activity of galactosyltransferase per g of liver increased immediately after birth, whereas that of sialyltransferase remained at a low level for 2 weeks and then increased to a constant level at 4 weeks. It is probable that this delayed increase in the activity of sialyltransferase results in the decreased sialylation of microsomal N-glycans at 1, 2 and 3 weeks. Sialyltransferase was solubilized from the liver microsomes of rats aged 2, 3 and 4 weeks and characterized. Phosphocellulose column chromatography separated the activity into two subfractions, designated transferase I and transferase II in the order of elution. The increase in total sialyltransferase activity during this period was caused mainly by an increase in transferase I. Rechromatography of each transferase from 3-week-old rats after neuraminidase treatment showed that transferase I but not transferase II contained sialic acid residue(s) and that desialylated transferase I was eluted in a similar way as transferase II. Although the apparent Km value for CMP-N-acetylneuraminic acid and the heat stability of transferase I were different from those of transferase II, the difference was abolished by treating transferase I with neuraminidase, suggesting that transferase II may be a desialylated form of transferase I. These changes in the sialylation of membrane glycoproteins, including sialyltransferase, may be related to the control of liver growth during postnatal development.
| S-EPMC1130617 | biostudies-other
Project description:The effect of palmitoylethanolamide (PEA), an endogenous fatty acid amide displaying neuroprotective actions, on glutamate release from rat cerebrocortical nerve terminals (synaptosomes) was investigated. PEA inhibited the Ca²⁺-dependent release of glutamate, which was triggered by exposing synaptosomes to the potassium channel blocker 4-aminopyridine. This release inhibition was concentration dependent, associated with a reduction in cytosolic Ca²⁺ concentration, and not due to a change in synaptosomal membrane potential. The glutamate release-inhibiting effect of PEA was prevented by the Ca(v)2.1 (P/Q-type) channel blocker ω-agatoxin IVA or the protein kinase A inhibitor H89, not affected by the intracellular Ca²⁺ release inhibitors dantrolene and CGP37157, and partially antagonized by the cannabinoid CB1 receptor antagonist AM281. Based on these results, we suggest that PEA exerts its presynaptic inhibition, likely through a reduction in the Ca²⁺ influx mediated by Ca(v)2.1 (P/Q-type) channels, thereby inhibiting the release of glutamate from rat cortical nerve terminals. This release inhibition might be linked to the activation of presynaptic cannabinoid CB1 receptors and the suppression of the protein kinase A pathway.
| S-EPMC4394492 | biostudies-literature
Project description:[3H]Mannose-labelled glycopeptides in the slices of livers from neonatal and 1-, 2-, 3- and 5-week-old rats were characterized by column chromatographies on Sephadex G-50 and concanavalin A-Sepharose and by endo-beta-N-acetylglucosaminidase H digestion. The proportion of complex-type glycopeptides was increased with time until 2 weeks post partum and then returned to the neonatal level. This was mainly due to the increased proportion of concanavalin A-bound (biantennary) species. These changes were accompanied by consistent changes in the activities of processing enzymes in liver microsomal fraction, especially of N-acetylglucosaminyltransferase I. Complex-type glycopeptides from neonatal and 2- and 5-week-old rat livers were further characterized by column chromatographies on Bio-Gel P-6 and DE 52 DEAE-cellulose in combination with neuraminidase digestion. No significant difference was found between concanavalin A-bound species from neonatal liver and those from liver 5 weeks post partum, most of which were sialylated. Concanavalin A-bound species 2 weeks post partum were comparatively smaller in size and less sialylated. On the other hand, there was no significant difference among concanavalin A-unbound species from the three different sources, most of which were sialylated. Since glycoproteins from regenerating rat liver also contain a higher proportion of complex-type oligosaccharides, as previously reported, such changes in N-linked oligosaccharides of glycoproteins may be related to control of the growth of liver cells.
| S-EPMC1149257 | biostudies-other
Project description:Nerve terminals are generally considered to be the destination points for electrical signals, which propagate unidirectionally from the soma to nerve terminals. We found that small hyperpolarizations or depolarizations ( approximately 10 mV) generated under physiological conditions in rat nerve terminals backpropagated up the axon ( approximately 400-800 microm) and changed the threshold for initiating action potentials and thus firing patterns. These results suggest a mechanism for information processing in neurons and neuronal circuits.
| S-EPMC3086371 | biostudies-literature
Project description:Excessive release of glutamate induces excitotoxicity and causes neuronal damage in several neurodegenerative diseases. Natural products have emerged as potential neuroprotective agents for preventing and treating neurological disorders. Dehydrocorydaline (DHC), an active alkaloid compound isolated from Corydalis yanhusuo, possesses neuroprotective capacity. The present study investigated the effect of DHC on glutamate release using a rat brain cortical synaptosome model. Our results indicate that DHC inhibited 4-aminopyridine (4-AP)-evoked glutamate release and elevated intrasynaptosomal calcium levels. The inhibitory effect of DHC on 4-AP-evoked glutamate release was prevented in the presence of the vesicular transporter inhibitor bafilomycin A1 and the N- and P/Q-type Ca2+ channel blocker ω-conotoxin MVIIC but not the intracellular inhibitor of Ca2+ release dantrolene or the mitochondrial Na+/Ca2+ exchanger inhibitor CGP37157. Moreover, the inhibitory effect of DHC on evoked glutamate release was prevented by the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) inhibitor PD98059. Western blotting data in synaptosomes also showed that DHC significantly decreased the level of ERK1/2 phosphorylation and synaptic vesicle-associated protein synapsin I, the main presynaptic target of ERK. Together, these results suggest that DHC inhibits presynaptic glutamate release from cerebrocortical synaptosomes by suppressing presynaptic voltage-dependent Ca2+ entry and the MAPK/ERK/synapsin I signaling pathway.
| S-EPMC8838318 | biostudies-literature
Project description:BackgroundPeripheral nerves can self-regenerate after traumatic injury, although their self-regeneration ability is limited after severe nerve injury. After peripheral nerve injury, the distal nerve stumps undergo Wallerian degeneration while the proximal nerve stumps undergo a regeneration process.MethodsHere, to decipher genetic changes and involved biological processes in the proximal nerve stumps after peripheral nerve injury, microarray data (GSE30165) were analyzed. Differentially expressed genes in the proximal nerve stumps at 0.5 h, 1 h, 3 h, 6 h, 9 h, 1 d, 4 d, 7 d, and 14 d after rat sciatic nerve transection were subjected to Ingenuity pathway analysis (IPA) bioinformatic analysis.ResultsCytokine signaling, cellular immune response, nuclear receptor signaling, disease-specific pathways, and organismal growth and development were significantly activated in the proximal nerve stumps after nerve transection. Organ development, inflammation and immune response, diseases and organ abnormalities, and cellular behavior-related biological functions were highly involved.ConclusionsThe expression levels of differentially expressed genes in biological function "Organismal Injury and Abnormalities" were displayed and validated. Our current study helps to obtain a better understanding of the biological processes of peripheral nerve regeneration, especially the regeneration process in the proximal nerve stumps, and thus may help to discover new therapeutic methods that can promote nerve regeneration.
| S-EPMC6990034 | biostudies-literature
Project description:BackgroundIn approximately 80% of patients, ovarian cancer is diagnosed when the patient is already in the advanced stages of the disease. CA125 is currently used as the marker for ovarian cancer; however, it lacks specificity and sensitivity for detecting early stage disease. There is a critical unmet need for sensitive and specific routine screening tests for early diagnosis that can reduce ovarian cancer lethality by reliably detecting the disease at its earliest and treatable stages.ResultsIn this study, we investigated the N-linked sialylated glycopeptides in serum samples from healthy and ovarian cancer patients using Lectin-directed Tandem Labeling (LTL) and iTRAQ quantitative proteomics methods. We identified 45 N-linked sialylated glycopeptides containing 46 glycosylation sites. Among those, ten sialylated glycopeptides were significantly up-regulated in ovarian cancer patients' serum samples. LC-MS/MS analysis of the non-glycosylated peptides from the same samples, western blot data using lectin enriched glycoproteins of various ovarian cancer type samples, and PNGase F (+/-) treatment confirmed the sialylation changes in the ovarian cancer samples.ConclusionHerein, we demonstrated that several proteins are aberrantly sialylated in N-linked glycopeptides in ovarian cancer and detection of glycopeptides with abnormal sialylation changes may have the potential to serve as biomarkers for ovarian cancer.
| S-EPMC3488482 | biostudies-literature
Project description:BACKGROUND:The use of immunodeficient mice transplanted with human hematopoietic stem cells is an accepted approach to study human-specific infectious diseases such as HIV-1 and to investigate multiple aspects of human immune system development. However, mouse and human are different in sialylation patterns of proteins due to evolutionary mutations of the CMP-N-acetylneuraminic acid hydroxylase (CMAH) gene that prevent formation of N-glycolylneuraminic acid from N-acetylneuraminic acid. How changes in the mouse glycoproteins' chemistry affect phenotype and function of transplanted human hematopoietic stem cells and mature human immune cells in the course of HIV-1 infection are not known. RESULTS:We mutated mouse CMAH in the NOD/scid-IL2R?c-/- (NSG) mouse strain, which is widely used for the transplantation of human cells, using the CRISPR/Cas9 system. The new strain provides a better environment for human immune cells. Transplantation of human hematopoietic stem cells leads to broad B cells repertoire, higher sensitivity to HIV-1 infection, and enhanced proliferation of transplanted peripheral blood lymphocytes. The mice showed no effect on the clearance of human immunoglobulins and enhanced transduction efficiency of recombinant adeno-associated viral vector rAAV2/DJ8. CONCLUSION:NSG-cmah-/- mice expand the mouse models suitable for human cells transplantation, and this new model has advantages in generating a human B cell repertoire. This strain is suitable to study different aspects of the human immune system development, provide advantages in patient-derived tissue and cell transplantation, and could allow studies of viral vectors and infectious agents that are sensitive to human-like sialylation of mouse glycoproteins.
| S-EPMC6322283 | biostudies-literature
Project description:Hypertension is the most prevalent risk factor for cardiovascular disease caused by a persistent increase in arterial blood pressure that has lasting effects on the mechanical properties of affected tissues like myocardium and blood vessels. Our group recently discovered that gut dysbiosis is linked to hypertension in several animal models and humans; however, whether hypertension influences the gut's mechanical properties remains unknown. In this study, we evaluated the hypothesis that hypertension increases fibrosis and thus mechanical properties of the gut. A custom indentation system was used to test colon samples from Wistar Kyoto (WKY) normotensive rats and Spontaneously Hypertensive Rats (SHR). Using force-displacement data, we derived an steady-state modulus metric to quantify mechanical properties of gastrointestinal tissue. We observed that SHR proximal colon has a mean steady-state modulus almost 3 times greater than WKY control rat colon (5.11±1.58kPa and 18.17±11.45kPa, respectively). These increases were associated with increase in vascular smooth muscle cells layer and collagen deposition in the intestinal wall in the SHR.Mechanical characterization of biological materials can provide insight into health and disease of tissue. Recent investigations into a variety of cardiovascular pathologies show coincident changes in the microbiome and pathology of the gut. In this study, we sought to quantify changes in the gut in hypertension through mechanical characterization. Our methods and simple models for characterization, adapted from Hertz indentation models, prove useful to identify a meaningful steady-state modulus metric for small and irregular tissues from laboratory animals. Our data, for the first time, establish a stiffening of the gut wall in Spontaneously Hypertensive Rats. This observation suggests significant structural and functional changes in the gut correlate with hypertension, and future experiments are warranted to explore the specific causal relationship between dysbiosis, fibrosis, and stiffening in the gut during the development and maintenance of hypertension.
| S-EPMC5069177 | biostudies-literature